Theranostics and Precision Medicine for the Management of Hepatocellular Carcinoma, Volume 1: Biology and Pathophysiology

Theranostics and Precision Medicine for the Management of Hepatocellular Carcinoma, Volume One: Biology and Pathophysiology provides comprehensive information about ongoing research and clinical data surrounding liver cancer. The book presents detailed descriptions about diagnostics and therapeutic options for easy understanding, with a focus on precision medicine approaches to improve treatment outcomes. This volume discusses topics such as tumor microenvironment in hepatocellular carcinoma, endoplasmic reticulum stress and unfolded protein response, effects of cirrhosis and hepatitis on the prognosis of HCC, mitochondrial metabolism, next generation sequencing, and telomerase. In addition, it discusses exosomes role in HCC progression, metastasis and chemokines.

This is a valuable resource for cancer researchers, oncologists, graduate students, hepathologists and members of biomedical research who need to understand more about liver cancer for their research work or clinical setting.

• Provides an updated literature review and detailed understanding of liver cancer tumor biology
• Discusses abnormal changes in the liver caused, resulting from, or associated with hepatocellular carcinoma from a holistic view
• Presents the content with fully colored images and summarizing tables for easy understanding of complex topics

• Language: English
• Publisher: Academic Press
• Release date: Feb 16, 2022
• ISBN: 9780323993647

Book preview

Preface

Sujatha Peela and Ganji Purnachandra Nagaraju

Hepatocellular carcinoma (HCC) is a form of primary liver cancer that has emerged as a most frequently diagnosed human malignancy worldwide. It predominantly targets individuals with underlying conditions such as cirrhosis, hepatitis B, and hepatitis C infections. Despite widespread occurrences of HCC, its underlying mechanisms that lead to tumor progression are still unclear. In contrast to various other cancers, systematic therapies for HCC, including chemotherapy and radiotherapy, are not effective. Currently, the only viable options for managing advanced HCC are surgical resection and transplantation. Therefore it is crucial to investigate pathways and factors that lead to HCC tumor suppression, thereby advancing novel therapeutic options of HCC.

In the current series we have three different volumes. Volume 1 discusses the biology, pathophysiology, and progression of liver cancer. Volume 2 discusses multiple signaling and molecular mechanisms associated with HCC progression and metastasis, including renowned and novel biomarkers as well as diagnostic approaches for advanced HCC. Volume 3 discusses several therapeutic targets for HCC and therapeutic molecules such as phytochemicals, and small molecules as well as clinical assessment, management, and precision medicine.

Volume 1 includes a detailed discussion about HCC cell origin, biology, and pathophysiology. It also includes the risk factors and pathogenic mechanisms associated with HCC development. The effect of hepatitis and cirrhosis on liver cancer prognosis is further explained in detail. This volume provides an overview of the mechanism by which aflatoxin b1 activates HCC and ultimately leads to tumor progression. The role of G protein–coupled receptor in HCC is discussed. Furthermore, the volume discusses the role of HCC stem cells, progression, and various therapeutic options.

In-depth explanation of the role of tumor microenvironment and polymorphism in HCC advancement and progression is scrutinized by the authors with detailed understanding of the impact of environmental pollution on HCC. Furthermore, the association between mitochondrial metabolism, succinate dehydrogenase, and telomerase is discussed in addition to the role of glutamine metabolism progression in liver cancer and its efficacy as a therapeutic target. It further analyzes the influence of endoplasmic reticulum stress and unfolded protein response during the commencement and development of HCC.

Finally, HCC induced by the comparative genomics and molecular epidemiology on hepatitis virus is elaborated. New opportunities for gene therapy are explained, specifically, the role of genetic insights and tumor microenvironment in liver cancer. Lastly, the roles of exosomes, noncoding RNAs, and microRNAs in HCC are discussed.

The aim of this series is not only to illustrate the experimental biochemistry and clinical significance of HCC but also to stress the functional and pathophysiological role of different signaling pathways. The knowledge of these roles could represent an essential step to understand the tumor markers in a way that their application will result in undeniable medical and economic advantages. This series will provide novel ideas to the researchers and scholars as well as innovative future prospective in the field of research and clinical applications.

Chapter 1

Cell origin, biology, and pathophysiology of hepatocellular carcinoma

Begum Dariya¹, Sujatha Peela² and Ganji Purnachandra Nagaraju³,    ¹Department of Biosciences and Biotechnology, Banasthali University, Banasthali, India,    ²Department of Biotechnology, Dr. BR Ambedkar University, Srikakulam, India,    ³School of Medicine, Division of Hematology and Oncology, University of Alabama, Birmingham, AL, United States

Abstract

Hepatocellular carcinoma (HCC) is a heterogeneous disease, ranking third for the cancer-related mortalities worldwide. Hepatocytes and progenitor cells are the cells of origin and play a major role in transforming into mature HCC cells. A better knowledge of the cell origin is essential as it aids in identifying tumor growth-associated molecular mechanisms and therapeutic options. Recent advances such as genome profiling and next-generation sequencing have uncovered various genomic alterations in HCC and profiled aberrantly behaving genes in HCC. The aberrantly behaving genes in HCC are TP53, CCND1, AXIN1, TERT promoter, CTNNB1, ARID1A, and CDKN2A. The tumor heterogeneity promotes tumor progression, metastasis, disease recurrence, and chemoresistance and limits patient prognosis. Thus a better understanding of the characterization of heterogeneity in HCC is significant for clinical practice and better survival of the patient. In addition, the biomarkers profiling for HCC is very limited and is essential for clinical decision-making. This chapter provides an overview about the cell of origin and biology of HCC, cellular heterogeneity, and associated pathways that contribute to HCC progression.

Keywords

HCC; heterogeneity; hepatocytes; progenitor cells; CTNNB1; TP53

Abbreviations

AAV2 Adeno-associated virus type 2

ARID1A AT rich interactive domain-containing protein 1A

CCNA2 Cyclin A2

CK19 Cytokeratin 19

CSC Cancer stem cells

CTNNB1 Catenin beta-1

EGFR Epidermal growth factor receptor

FGF Fibroblast growth factor

HBV Hepatitis B virus

HCC Hepatocellular carcinoma

MDM2 Mouse double minute 2 homolog

NGS Next-generation sequencing

NSAID Nonsteroidal anti-inflammatory drugs

TERT Telomerase reverse transcriptase

TGF Transforming growth factor

VEGF Vascular endothelial growth factor

YAP-1 Yes-associated protein

Introduction

Hepatocellular carcinoma (HCC) is determined as the third leading cause for cancer-related deaths worldwide [1]. The major risk factors include chronic liver damage caused due to fibrosis and inflammation, infection by hepatitis C or hepatitis B virus [1]. The therapeutic regimen like surgical resection, hepatic transplantation, and techniques like local ablation with radiofrequency remain as the mainstay for HCC therapy [2]. However, only 30% cases are only eligible for the surgery as most of the patients are diagnosed at their advanced or metastatic stages and thus require more efficient therapies. The conventional interventions including chemo-, immune-, and targeted therapy are now widely developed to prevent primary and secondary recurrence of the disease. The molecular heterogeneity of HCC progression is associated with the prognosis and therapeutic efficiency [3]. Therefore understanding the origin of heterogeneity is the principal approach to better progress in diagnosis and therapeutic strategies. Additionally, a deeper insight of molecular mechanisms responsible for cellular origin, tumor progression, and maintenance enables the detection of associated tumor markers for diagnosis and prognosis as well as promote developing new targeted therapies for HCC. In this chapter, we provide an overview for cell origin and pathophysiology of HCC and their importance in diagnostic and therapeutic approaches.

Cell of origin of HCC

The hepatic structures are made of parenchymal (cholangiocytes and hepatocytes) and nonparenchymal cells (endothelial, fibroblasts, Kupffer, and stellate) [4]. The hepatic progenitor cells (hepatoblasts) are the common progenitors for hepatocytes and cholangiocytes that further initiate hepatocarcinogenesis as revealed from various hypotheses of cell of origin. However, the most common liver cancer HCCs and intrahepatic cholangiocarcinoma are distinct and are developed from respective matured hepatocytes and cholangiocytes [5,6]. In the further sections, we have enclosed evidences for progenitor cells and hepatocytes as the major cellular origin of HCC.

The previous researchers determined that the progenitor cells isolated have the ability to develop into hepatic carcinoma after they are transplanted into recipient mice and performed consecutive ex vivo genetic modifications [7]. For instance, a driven transformation of IL-6 accompanied with transforming growth factor-beta (TGF-β) in the progenitor cells of a mice liver have developed cancer [8]. Additionally, a constant stimulation of TGF-β in cirrhotic liver promotes neoplastic transformation of progenitor cells to tumor-initiating cells that further facilitate hepatocarcinogenesis via activating miR216a and PTEN pathway [9]. Similarly, a mice attenuated with Hippo signaling cascade resulted in increased expression of YAP protein have shown liver tumor transformation [10,11]. NOTCH signaling is also found activated in almost one-third of the HCC human samples [12]. The NOTCH targeting genes SOX9 were also found overexpressed in HCC and are taken as marker of progenitor cells [13]. A unique population of progenitor cells is developed into HCC with lung metastasis with the stabilization of somatic β-catenin protein and Wnt pathway [14]. However, in certain cases, the activation of Wnt pathway is insufficient and additional alterations of oncogenic Ras and AKT mutations were essential for cancer transformation [15–17]. Alternatively, the mice cells with deficient hepatocytes acquired cancer stem cells-like characteristics and developed into HCC cells via activation of STAT3 protein. The depiction of neoplastic development led to the identification of progenitor markers like cytokeratin (CK19) in preneoplastic lesions, potentially developed into tumor [18,19]. In addition, other markers including CD133+ CD44+ were also found as a part of progenitor cells [20]. Thus the markers are potentially used for therapeutic and diagnostic purpose for the benefit of the patient.

The other major source for HCC is hepatocytes. These cells are polyploid and constitute a total of 60%–80% of the liver mass [4]. They possess additional properties such as remarkable plasticity that entails the capacity to dedifferentiate into progenitors. This eventually transforms into hepatocyte pool via genetic alterations, contributing to phenotypical and molecular diversity. For instance, the mature hepatocyte develops into Nestin-positive progenitor like cells develop into primary liver cancers with the loss of tumor suppressor TP53 function and acquiring mutations in Wnt pathway [21]. In addition, the hepatocyte-specific p62 expression also showed HCC initiation via activation of mTOR1 and induction of c-MYC [22]. Moreover, the hepatic cells that show the expression of activated oncogenes like H-RAS and growth factors FGF19 also promote HCC transformation [22,23]. STAT3 is activated through several cytokines, including interleukin 6 (IL-6) in tumor microenvironment contribute to FGF-19-associated tumorigenesis. The mesenchymal-epithelial transition proteins that play a crucial role in stem cell renewal also induce dedifferentiation of hepatocytes and activation of epidermal growth factor receptor (EGFR) and NOTCH [24,25], leading to HCC development [26].

Thus both hepatocytes and progenitor cells contribute to intratumoral heterogeneity. However, the HCC originated majorly from hepatocytes, while the progenitor cells are activated and proliferate into benign lesions [27].

Biology and pathophysiology

The cells are the essential elements of heterogeneity that influences tumor growth. The healthy, transformed, and malignant cells possess the property of heterogeneity to a certain extent. The hepatocytes originated from the central vital vein to the portal nodal cells of a liver show a regularity in the profile of gene expression [28]. For instance, the hepatocytes markers including Cyp2e1, Glul, Cyp2f2, and Alb from different sites are expressed differently. The renal cells transformed into precancerous nodules show heterogeneous genetic and epigenetics variation [29].

Earlier with minimal technologies, identification of tumor diversity was a vast process to the pathologist and took at least a century to describe the heterogeneity of the tumor via immune-histological staining but evaluated less quantitatively [30]. However, the current emerging techniques such as single-cell sequencing, spectrometry, immunoblotting, and multiomics techniques have potentiated the development of a clear landscape of cellular and tumor heterogeneity [31–34]. Triple omics sequencing techniques such as scTrio-seq have provided evidences for developing genetic heterogeneity of HCC [35]. A subpopulation of cancer cells having the property of stemness is known as cancer stem cells (CSCs) grab the attention of researchers for featuring single-cell heterogeneity. For instance, the CSCs such as EpCAM [36,37], CD133 [36,38–40], CD44 [36,41,42], CD90 [36,43], CD47 [42], CD13 [39,42], and CD90 [36,43] are identified as markers of hepatic CSCs using single-cell sequencing. The CSCs are not a druggable subset but are influenced or mediated by various environmental factors such as hypoxia, DNA damage, and inflammation phenotypically in various cancers [44,45]. For instance, an oxygen gradient between veins and arteries in the renal lobules induced vascular perfusion in HCC [46]. This promoted differential activation of hypoxia-inducible factor signaling cascade in an oxygen-dependent way and subsequently induced stemness. Thus targeting CSCs would be an efficient therapeutic target. However, improved understanding of CSCs and identifying novel opportunities must be developed to overcome the heterogeneity.

Molecular consensus of HCC

The molecular classification of HCC is characterized based on the genomic landscape and is correlated with etiological, pathological, and clinical features of the patient’s outcome. The two major subgroups include proliferation and nonproliferation classes.

The tumors that come under proliferation class are heterogeneous and enriched with signaling cascades associated with proliferation such as insulin-like growth factor, mammalian target of rapamycin, and NOTCH [46,47]. These classes also involve epigenetic characteristics that include expression of DNA methylation and miRNA [48,49]. The gene expression associated with tumor recurrence is also included in the proliferation class [50]. Additionally, the chromosomal aberrations that are associated with transcriptome bases subclasses are highly detected in HCC specifically at chromosome 1 and chromosome 8. Similarly, high-level amplifications detected in tumors from the proliferation class were reported in Chr. 6p21 of vascular endothelial growth factor A and Chr. 11q13 of FGF19 and CCND1.

The tumors of nonproliferation class retain the features of hepatocytes and are related to the activation of Wnt pathway that occurs via mutation in CTNNB1 [51]. This subclass includes well-differentiated but less aggressive tumors having decreased levels of α-fetoprotein. In addition, they also include aberrant expression of EGFR pathway and activation of inflammatory pathways involving NF-κB and IL-6 [52,53].

The recent advances in next-generation sequencing (NGS) have now given a complete picture for molecular interactions in HCCs [54] that potentiate the development of mutation signatures. On the basis of this, the researchers have developed mutation signatures for HCC that are associated with aflatoxinB, smoking, and alcohol [55]. The clinical data of patient cohorts have promoted exome sequencing of the prevalent mutations associated with HCC pathogenesis such as TP53, ARID1A, TERT promoter, and CTNNB1 [56,57]. A map of viral integrations including hepatitis B virus (HBV) [58] and adeno-associated virus type 2 (AAV2) [59] correlated with HCC progression were also provided by NGS. The HBV integrations involve the effect of genes MLL4, TERT, and CCNE1 and are determined as poor prognosis of patients and affect the survival of patient [58]. Similarly, AAV2 was reported in almost 5% of HCC cases affecting CCNA2, TERT, and CCNE1 [59].

Signaling pathways

The HCC resulted from the abnormal signaling pathways contributes as the novel molecular therapeutic targets.

The Wnt pathway-associated genes including AXIN1 and CTNNB are found dysregulated in HCC. CTNNB1 is a β-actin protein and is a crucial protein in the Wnt pathway that regulates cell adhesion, growth, and differentiation. The mutation in CTNNB1 harbors about 11%–41% of HCC [60–63]. Furthermore, Wnt inhibitors and nonsteroidal anti-inflammatory drugs such as sulindac and celecoxib targets Wnt pathway and activated CTNNB1 [64]. Sorafenib also targets Wnt cascade and decreases the expression of CTNNB1 [65–67]. AXINI is the negative regulator of Wnt and controls the expression levels of β-catenin. However, the mutation of AXIN1 harbors 5%–19% of HCC [60–62]. Thus targeting AXIN1 would accelerate apoptosis and inhibit HCC. XAV939 is a molecular inhibitor, suppresses poly-ADP ribosylating enzymes (tankyrase 1 and 2), and stabilizes AXIN [68], thus signifies as the molecular targets of HCC therapy.

The genomic aberration of p53 pathway resulted due to the mutation in TP53 and harbors about 13%–48% of HCC [60–62]. TP53 is a tumor suppressor protein and induces apoptosis and regulates vascular endothelial growth factor (VEGF) expression [69]. Anti-angiogenic agents such as bevacizumab improved survival of the patients that harbored tumor progression with TP53 mutation [70,71]. Similarly, the cell cycle-associated genes including loss or deletion of CDKN2A harbor 8% of HCC progression. It is a tumor suppressor and promotes cell cycle arrest at G1–G2 phases. In addition, it also inhibits the expression of oncogenes CDK4/6 and MDM2 [63]. However, the mutated CDKN2A induces upregulation of CDK4 and CDK6.

Chromatic remodeling

ARID1A and ARID2 are the two genes that play crucial role in chromatin remodeling. The commonly detected ARID1A harbors 4%–17% of HCC [60–62,72]. Their alterations are significantly associated with mutation in CTNNB1 and activation of Akt/PI3K/mTOR signaling pathway. ARID2 is a tumor suppressor gene, however, mutational changes in the gene harbors 5%–7% of HCC [56,61,72]. ARID2 plays a major role in selectively promoting transcriptional activation and suppression of oncogenes via chromatin remodeling.

Other activated genomic alterations include TERT promoter mutation, which accounts for 60%–90% of HCC [73–75]. It plays major role in telomerase maintenance pathway, via adding telomerase repeats at the chromosomal ends and protects telomeric ends; however, mutation in TERT promotes the cell to divide continuously and is thus involved in malignant transformation of HCC. Similarly, loss of PTEN activates kinase-promoting pathways including PI3K/Akt/mTOR [76]. PTEN deletion accounts for 53% of HCC cancers [77]. Thus these dysregulated or aberrantly behaving genes can be targeted to enhance the therapies. However, further investigation with appropriate biomarkers is highly warranted.

Conclusion

Over the last few decades, with the development of NGS, there has been an enormous knowledge of molecular and cellular level complexity of HCC. The current NGS studies have efficiently contributed information about the cells and proteins involved in signaling cascades that promote carcinogenesis. Additionally, the in vivo lineage tracing model and cell tracing models have extensively revealed the plasticity of mature hepatocytes and progenitor cells that further transformed into HCC with progenitor biomarkers.

Detecting the origin of cells that develop into tumor and characterizing these subclasses of tumors basing on their molecular level features could develop a novel approach for therapy and prognosis. Furthermore, incorporating molecular signatures or biomarker studies into the therapies would efficiently bring a better prognosis. Thus learning more about tumor microenvironment and tumor heterogeneity to characterize the genetic and molecular level features are highly essential to potentiate the advanced personalized treatment.

Conflict of interest

None.

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Chapter 2

Hepatocellular carcinoma—An updated review

Varimadugu Aruna, A. Sneha and D. Sai Harshitha,    Chaitanya Bharathi Institute of Technology, Hyderabad, India

Abstract

Cancer is referred to as the abnormal proliferation of cells. This occurs due to combined genetic and nongenetic alterations that are usually induced by environmental factors, which trigger inappropriate expression of specific genes leading to neoplastic transformations. There are many types of cancer among which the sixth most frequently detected is the liver cancer. The occurrence of hepatocellular carcinoma (HCC) has been steadily rising year by year all over the world. This review deals with various reasons for HCC stating from lifestyle to mutations. The literature suggested that the cause for the occurrence of this cancer is due to aflatoxin, alcohol, and hepatitis virus. Though the detailed mechanism of progression of HCC is not exactly clear, some experimental evidence that provides partial information about the mechanism of HCC are discussed here. In this chapter various methods that are available for the diagnosis of HCC and its further treatment are discussed. With technological advancements in the recent years, many new methods are designed for the detection of HCC as well as to treat it. This can further increase the life span of a person to some more extent.

Keywords

Aflatoxin; cancer diagnosis; cancer treatment; chemotherapy; cholangiocarcinoma; hepatectomy; hepatocellular carcinoma; hepatitis; liver cancer; liver transplantation; oncogenic drivers; radiation therapy; surgical resection; tumor suppressor

Abbreviations

AFP Alpha-fetoprotein

CT Computed tomography

GPI Glycosylphosphatidylinositol

HBV Hepatitis B virus

HCC Hepatocellular carcinoma

HCV Hepatitis C virus

HSPGs Heparan sulfate proteoglycans

MRI Magnetic resonance Imaging

NAFLD Nonalcoholic fatty liver disease

ROS Reactive oxygen species

Introduction

In many areas of the world, the leading cause of death is cancer [1]. For many years, cancer incidence has been increased when compared with other diseases [2]. Cancer results in abnormal cell proliferation due to many changes in genetic expression and involves many steps. Cancer can be benign or malignant, but malignant cancers lead to lethal situations [3]. Evidence has proven that cancer can be linked with lifestyle, epigenetic factors, gene aberrations, and many more [4]. Different types of cancer can be seen in people at any age, but especially persons with older age—disease of aging [5].

The cancer complexity which can be understood by a few rules that describe the conversion of a general cell into malignant cells is called hallmarks of malignancy, such as independence in growth signals, insensitivity to antigrowth signals, overcoming apoptosis, unlimited replicative potential, maintained new blood vessel formation, and invasion and hallmarks of malignancy [6,7]. It was stated in a study by Globocon in 2020 that globally the number of cancer cases was approximately 19.3 million and that of deaths due to cancer was 9.9 million. The most leading cancer type occurring in males is lung cancer with 14.3% of total cases of cancer and 21.5% of total deaths due to cancer. In females, it is breast cancer with 24.5% of cancer cases and 15.5% of cancer deaths [8]. Treatment of cancer can be done using various methods such as chemotherapy, radiation therapy, surgery, cancer vaccinations, or combinations, but people prefer alternative ways than conventional medicine [9]. It was found that after the 1970s, there is a rise in mortality rate by 20% in both men and women