Hepatocellular carcinoma (HCC) occurs primarily in patients who have sustained years of chronic liver injury. Unfortunately, most patients are diagnosed with advanced disease when potentially curative surgical resection or liver transplant is no longer a therapeutic option and medical treatments for HCC are largely ineffective. Consequently, the average 5-year survival for HCC is less than 15%. The global burden of HCC is expected to increase in the coming decades as a result of an increasing population and the global rise of obesity and fatty liver disease as emerging HCC etiologies. Given this context, prevention efforts for HCC are urgently needed to reduce the incidence of this deadly disease. Primary prevention involves avoidance of exposure to risk factors for chronic liver disease, including toxins such as ethanol and certain chemicals, viral hepatitis through vaccination, and overweight or metabolic syndromes via lifestyle modification. National neonatal hepatitis B vaccination and reduced aflatoxin exposure have successfully reduced HCC incidence in traditionally endemic regions such as East Asia. Secondary prevention encompasses mitigation of risk factors in patients with baseline chronic liver disease. Multiple dietary and pharmaceutical agents have been suggested to have preventative potential in retrospective analyses, though few clinical trials have been performed. Finally, tertiary prevention represents interventions to reduce the risk of de novo HCC recurrence after curative resection of primary HCC tumor. In this chapter, we provide a comprehensive overview of chemopreventive strategies and discuss future directions.

Rationale for HCC Prevention

Hepatocellular carcinoma (HCC) is the fifth most common cancer with rising global incidence [1]. Surgical resection, liver transplant, and ablation provide the only potential curative therapies, though over 70% of patients are diagnosed at advanced stages of disease. Unfortunately, medical therapies have been largely ineffective, and HCC mortality remains high, with a 5-year survival of less than 15% in the United States [2]. Moreover, despite rigorous evaluation of screening strategies, surveillance noncompliance remains a limiting factor for early diagnosis and curative treatment [3–5]. HCC arises from a milieu of chronic inflammation and persistent liver injury that progresses over decades and occurs 40 times more often among cirrhotics [6, 7]. Therefore, there is potential for identification of at-risk individuals and development of preventative strategies. For these reasons, prevention is an attractive alternative approach for HCC management.

There are three main approaches to HCC prevention, primary, secondary, and tertiary, which are distinguished by a previous history of chronic liver disease and HCC [8]. The purpose of primary HCC prevention is to identify and abolish risk factors for chronic liver disease, and among patients with already established cirrhosis, to treat underlying chronic liver conditions prior to the onset of HCC [9]. Multiple primary and secondary preventative strategies have been proposed, though few have been validated in prospective, randomized controlled trials (RCTs). Particular strategies have increased relevance based on geography and context. Endemic HBV in East Asia and Sub-Saharan Africa presents a major source of cirrhosis and HCC, and therefore, implementation of vaccination programs stand to reduce the burden of hepatitis-mediated HCC in these regions [10]. Similarly, aflatoxin exposure remains a public health issue in East Asia, and policy changes to reduce dietary toxin exposure is another potential high-yield intervention [11]. In contrast, in Western nations, obesity-related nonalcoholic steatohepatitis (NASH) is the fastest rising cause of liver failure and HCC [12, 13]. In this regard, more focus has been placed on bariatric surgery and anti-inflammatory pharmacologic agents that are used to treat metabolic syndrome-related disorders, including aspirin, statins, cyclooxygenase inhibitors, and anti-diabetic agents [14]. There is also great interest in dietary supplements, notably coffee and green tea polyphenols, both of which are thought to have anti-inflammatory and anticancer effects. Hepatitis C virus (HCV) is highly prevalent on a global scale, and the recent development of effective direct acting antivirals (DAAs) offers great potential not only for curative therapy in patients with chronic viral hepatitis but secondarily to reduce the transmission and incidence of HCV [15].

Tertiary prevention strategies are designed to reduce the risk of HCC recurrence after curative resection of primary disease, typically in patients with baseline chronic liver disease or cirrhosis. Many of the same dietary and pharmacologic agents that are under investigation for secondary prevention are also being studied in the tertiary context. In addition, adjuvant chemotherapeutic agents targeting cancer are also under investigation. In this chapter, we systematically review HCC prevention strategies, their indications and limitations, and future directions for HCC prevention.

Primary and Secondary Prevention Strategies

HCC is a malignancy borne from chronic hepatic insults, and approximately 90% of cases occur in the setting of cirrhosis. The annual incidence of HCC among cirrhotics is approximately 2–5% [16, 17]. The most common etiologies for chronic liver disease and cirrhosis include viral infection, alcohol consumption, obesity and metabolic syndromes, autoimmune and genetic disorders, and toxin exposure [18]. Despite each having unique molecular mechanisms, these processes result in a common pathogenic pathway characterized by persistent hepatocyte injury, chronic inflammation, progressive hepatic fibrosis, and ultimately hepatocarcinogenesis and HCC development [19]. In the sections below, we review these common etiologies of chronic liver disease and targeted efforts for HCC prevention.

Viral Hepatitis

Approximately 80% of new HCC cases occur in developing countries, of which 80% are attributable to HCV and HBV infection [20]. In the United States and Europe, untreated HCV infection accounts for 25–75% of HCC cases [21]. Among HCV and HBV carriers, the risk of HCC is increased by approximately 17-fold and 15-fold, respectively [22]. Viral oncogenesis is a complex multistep process that initiates with hepatocyte infection, injury, and subsequent inflammation. Autocrine and paracrine signaling feedback loops between hepatocytes and stellate cells establish a pro-survival, chronically inflamed microenvironment with reduced immune surveillance that is optimal for hepatocyte transformation [23]. HCV and HBV have unique molecular mechanisms that contribute to this process, as discussed below.

HCV Epidemiology and Pathogenesis

Approximately 3% of the world’s population is infected with HCV, which is a single-strand RNA virus from the Flaviviridae family. The annual HCC incidence among patients with cirrhosis and active HCV infection ranges from 1% to 8%; however, with effective treatment, evidenced by sustained viral response (SVR), this incidence decreases to 1% or less [24, 25]. Unlike HBV, the risk of HCV-related HCC increases with the degree of cirrhosis, and HCC is rarely observed in the absence of hepatic fibrosis [26]. There is conflicting evidence regarding the association between serum viral RNA load and HCC risk [27, 28]. Due to a high viral mutation rate, attempts at vaccine development against HCV have been unsuccessful but remain an active area of investigation even after the successful development of effective direct acting antivirals (DAAs).

HCV often presents with a mild acute phase and the majority of patients are unaware of their infection status until signs of chronic liver disease present several decades later. In the United States, approximately 1.6% of the populace is infected with HCV; however, less than 25% of individuals are aware of their infection status, and this rate decreases to less than 5% globally [29]. In the United States, the majority of HCV infections occurred among the “baby boomer” generation (prevalence of nearly 3%), and therefore the Centers for Disease Control (CDC) recommends HCV testing for anyone born between the years 1945 and 1965 [30]. Approximately 80% of HCV infections progress to chronic hepatitis, of which 15% will progress to cirrhosis [31]. It is projected that without treatment, 14% of HCV patients will eventually develop HCC [32]. Given the chronic nature of HCV, there is theoretically ample opportunity for preventive intervention. However, because the disease is frequently silent for many years and regular HCV testing is not utilized in most countries, there is still the hurdle of identifying patients who are infected.

The mechanism of HCV-induced hepatocarcinogenesis is not well established, though the oncogenic effects are likely secondary to viral proteins as the virus is unable to stably integrate into the genome. HCV core protein is capable of inhibiting multiple tumor suppressors including p21, p53, and Rb, and has also been associated with CDH1 downregulation via promoter hypermethylation, TGF-β upregulation, and activation of MEK/ERK phosphosignaling [33–35]. On a physiologic level, HCV core protein has been shown to influence multiple cellular processes, including proliferation, survival, lipid metabolism, reactive oxygen species (ROS) generation, and immune interactions [36]. In particular, HCV core protein, as well as another HCV protein, NS5A, has been shown to impair β-oxidation, resulting in reduced mitochondrial electron transport chain function, endoplasmic reticulum (ER) stress, impaired lipid metabolism leading to hepatic steatosis, and increased ROS formation [37, 38]. Given the interdependence of these metabolic functions, dysregulation of any given process can further deregulate cellular homeostasis in a feed-forward manner. Over time, HCV-induced hepatocyte dysfunction and cell death establishes an inflammatory microenvironment that drives stellate-cell activation, fibrogenesis, and altered immune surveillance, which are conditions that have been shown to support hepatocyte transformation.

HCV-Targeted Prevention

Interferon (IFN)-based therapy was introduced in 1986 and served as the mainstay of HCV treatment prior to the development of DAAs. IFN activates the immune system, which frequently results in enhanced viral detection and clearance. SVR rates of 50–70% in genotype 1/4 HCV patients without advanced fibrosis were achievable using pegylated interferon (PegIFN) in combination with ribavirin, a nucleoside inhibitor [39]. However, among patients with bridging fibrosis or cirrhosis, SVR was observed in only 51% and 33%, respectively. Given the immune-stimulating effects of IFN, treatment is often associated with severe adverse effects. Thus, low efficacy and high toxicity limit the utility of IFN-based HCV treatment.

Advances in our understanding of molecular virology led to the development of DAAs, which were introduced in 2011, creating a paradigmatic shift in our ability to safely and effectively treat HCV. Initially, first-generation NS3-4A protease inhibitors, telaprevir and boceprevir, in combination with PegIFN and ribavirin were shown to achieve SVR rates of 65–75%, resulting in their Food and Drug Administration (FDA) approval for HCV genotype 1 treatment [40, 41]. This was followed by approval of Simeprevir, another NS3-4A inhibitor, in 2013 [42]. However, development of NS5B polymerase inhibitors led to a critical breakthrough in achieving SVR. Sofosbuvir, a nucleotide analogue that causes early chain termination of viral RNA, was shown to induce SVR rates of 90% in combination with PegIFN and ribavirin triple therapy [43]. Multiple additional inhibitors targeting NS3-4A, NS5B, and NS5A (replication complex protein) have since been developed. Importantly, two-drug DAA combination therapy, of which there are multiple permutations, has been shown to induce SVRs of greater than 90% [44–46]. A list of currently used DAAs is included in Table 13.1.

Table 13.1

Current DAAs for HCV and their efficacy, genotype specificity, resistance rates, and combination regiments

HCV SVR is associated with significant reductions in all-cause mortality, hepatic fibrosis progression, and hepatocarcinogenesis [60]. In a recent meta-analysis evaluating 25,906 HCV patients treated with IFN-based regimens, the incidence of HCC was 1.5% in patients who achieved SVR compared to 6.2% among nonresponders [61]. Loannou et al. recently performed a meta-analysis evaluating the effect of DAA-associated SVR on HCC incidence. The study included 62,354 patients treated with IFN-only regimens (58%), DAA plus IFN regimens (7%), or DAA-only regimens (35%). In all instances, they found an HCC risk reduction of 71–76% in patients who achieved SVR [62]. The risk reduction did not vary by treatment received. However, even among patients who achieve SVR with therapy, the risk for HCC remains elevated compared to healthy controls. The risk of HCC after SVR appears highest in patients with advanced liver fibrosis and diabetes mellitus, and therefore, these patients should be enrolled into HCC screening protocols and additional preventive measures should be considered [63].

HBV Epidemiology and Pathogenesis

As of 2017, 291 million people are infected with hepatitis B (HBV), which is responsible for 887,000 deaths per year globally [64]. HBV is endemic in developing regions including Southeast Asia and Africa, where approximately 8% of the population is infected, which is largely due to a lack of healthcare resources and established neonatal vaccination programs. In contrast, carrier rates in developed Western nations including the United States are below 2%. Two-thirds of patients who develop acute HBV infection exhibit minimal symptoms causing the illness to go undetected, which is a contributing factor to high transmission rates in regions where HBV screening is rarely available [65]. HBV is most frequently vertically transmitted during infancy; therefore, the World Health Organization (WHO) and the Centers for Disease Control (CDC) recommend HBV vaccination within the first 12–24 h after birth. HBV is highly infectious by requiring less than ten viral particles to establish hepatocyte infection [66, 67]. The likelihood of developing chronic infection, and thereby increased cancer risk is related to the age of HBV exposure. In newborns, the risk of chronic HBV infection is 90%, while conversion from acute to chronic infection in adults, who have more well-established immune function, is less than 5% [68]. Chronic HBV is the leading risk factor for HCC development globally (approximately 15-fold increased risk), and approximately 80% of new HCC arise in regions of endemic HBV [69]. Unlike HCV-related HCC, which rarely develops in the absence of cirrhosis, HBV-related HCC can occur in the absence of chronic liver disease, indicating a unique oncogenic mechanism [70].

The pathogenesis for HBV-related HCC is not well established, though is thought to include direct effects from viral DNA integration and translated protein products, as well as indirect effects from secondary hepatic inflammation and fibrosis. Integration of HBV DNA into the host hepatocyte genome is observed in 84.6% of HBV-related HCC [71]. The location and frequency of integrations determines the degree of subsequent genomic instability. Moreover, insertion mutations within tumor suppressor genes have been shown to support aberrant hepatocyte proliferation and survival [71, 72]. Unlike HCV, HBV serum DNA levels directly correlate with risk of HCC development; however, the HBV replication cycle is not directly cytotoxic to hepatic cells [73]. Translation of the HBV genome yields multiple protein products including the viral envelope core, polymerase proteins, and preC and hepatitis B X (HBx) polypeptides [74]. HBx has been associated with multiple oncogenic processes including activation of the Ras-Raf-MAPK pathway, upregulation of miR-181a, and inhibition of p53 and PTEN tumor suppressors [75–77]. Host immune responses to HBV antigens are critical determinants of hepatocellular injury [74]. HBV-induced hepatic inflammation drives local ROS generation and increased hepatocyte oxidative stress, inducing damage to lipids, proteins, and DNA, as well as alteration of multiple cell signaling pathways including MAPK and PI3K. Chronic HBV infection also upregulates hepatic NF-kB and STAT3 signaling and circulating IL-6 levels [78]. These signaling changes stimulate prosurvival gene networks, supporting hepatocyte accumulation of mutational burden while also dampening immune surveillance and thereby increasing the likelihood of malignant transformation.

HBV-Targeted Prevention

Vaccination is the most impactful form of HBV prevention. Establishment of immunity early in life is critical as the likelihood of developing chronic infection, and thus increased HCC risk, is greatest among infants and children. In the United States, initiation of a national neonatal vaccination program in 1981 has led to a substantial drop in the infection rate (9.6 per 100,000 persons in 1982 to 1.1 per 100,000 persons in 2015). The first of 3 vaccine doses are given within 24 h of birth and provides full protection to greater than 90% of infants, children, and adults who receive the entire series [79]. Over 175 countries have now implemented HBV vaccination programs, with the greatest impact occurring in regions with endemic HBV, and it is estimated that over 210 million new chronic infections have been prevented worldwide as of 2015 [80]. For example, Taiwan reported an 80% reduction in HCC incidence upon implementation of a nationwide vaccination program in 1984. Moreover, the childhood/adolescent HCC incidence decreased by 51%, which translated to a 90% reduction in HCC mortality in individuals aged 5–29 years old [81]. However, there are still developing regions with endemic HBV, notably Southeast Asia and Africa, where vaccination programs are lacking, and the burden of HBV-related HCC and mortality remains high.

Multiple challenges hinder successful implementation of birth dose vaccination in low-income and middle-income countries, including high out-of-hospital birth rates, monetary limitations, and healthcare misconceptions regarding vaccine safety [82]. Implementation of successful vaccinations programs in these challenging settings requires understanding of specific regional limitations. For example, Bangladesh, Myanmar, Nepal, and Timor-Leste have low health facility birth rates; therefore, strategies to promote institutional deliveries and out-of-clinic immunizations might provide tailored benefit. Similar principles have been championed in India where a trained healthcare professional is present for approximately 52% of births. Accordingly, programs were instituted to train midwives in vaccination, and standardized neonatal and maternal care protocols were developed, which increased HBV vaccination coverage by twofold [83].

Drug treatment for acute HBV infection is rarely needed for adults, as the vast majority of individuals clear the virus with only mild hepatitis. However, for individuals who develop fulminant hepatitis or chronic infection, antiviral therapy is indicated. Among chronically infected HBV patients, outcomes have improved over the last three decades due to the development of IFN-based therapies and nucleoside/nucleotide analogues (NAs) [84, 85]. Antiviral treatment, regardless of drug type, has been shown to reduce 3-year HCC incidence rate from 4% to 1.5% and 5-year HCC incidence rate from 12% to 5.1% [86]. Blood-based viral load and HBe-Ag titer correlate with viral suppression and future HCC risk; therefore, reductions in these markers are used as surrogates for treatment efficacy [87–89]. Although previous trials have shown that PegIFN/Lamivudine combination therapy is superior to monotherapy for achieving SVR (PegIFN/Lamivudine 57%, Lamivudine 31%, PegIFN 20%), the development of newer and more effective NAs has limited the use of PegIFN, which has a toxic side effect profile [90, 91].

NAs act as viral DNA chain terminators by inhibiting HBV polymerase and reducing HBV replication. Clinically available NAs include lamivudine (Epivir), adefovir (Hepsera), tenofovir (Viread), telbivudine (Tyzeka), and entecavir (Baraclude). First-generation agents were limited by weak antiviral activity and high susceptibility to drug resistance (76% genotypic resistance after 8 years of treatment for lamivudine and 29% resistance rate after 5 years of adefovir treatment) [92]. Entecavir and tenofovir are newer NA agents that have shown superior efficacy to lamivudine and adefovir, achieving SVR in 95% of patients and regression of cirrhosis in 71– 96% of patients at 3-year follow-up [93, 94]. These drugs are also associated with lower rates of resistance (entecavir resistance between 0.5% and 1.2% after 5 years) [89, 95]. Hosaka et al. showed that among chronically infected HBV patients, the cumulative 5-year HCC incidence was significantly reduced with entecavir monotherapy compared to no treatment (3.7% vs. 13.7%). Furthermore, entecavir reduced HCC incidence by fourfold in cirrhotic patients, who are at greatest risk HCC development [96]. In 2015, the WHO published updated guidelines recommending tenofovir or entecavir as first-line agents [97]. Unfortunately, these newer and more effective NAs are prohibitively expensive for many poor, underdeveloped regions with high rates of endemic HBV infection [98, 99].

Finally, despite improved viral suppression and low resistance rates associated with newer NAs, these medications must be taken indefinitely due to HBV integration into the host genome and continued replication [100]. Only 10% of patients on NA therapy have complete clearance of HBs-Ag after 5 years of treatment [101]. Consequently, although these agents reduce the risk of HCC development, the incidence does not decrease to the level of healthy controls [102]. Therefore, novel approaches targeting multiple steps of the HBV lifecycle are currently under investigation (Table 13.2), including HBV viral entry, formation of covalently closed circular DNA (cccDNA), pregenomic RNA (pgRNA) packing, and virion assembly and secretion [114]. In particular, approaches targeting cccDNA might allow for complete eradication of viral genomic material. In this regard, multiples research teams are investigating the use of CRISPR/Cas9 technology for selective destruction of cccDNA [115, 116]. Other approaches involve epigenetic silencing of cccDNA via HBx targeting and exogenous cytokine therapy to stimulate pathways that drive cccDNA degradation [117, 118].

Table 13.2

Novel therapies for chronic HBV infection

Metabolic Insult and Disorders

Alcohol Epidemiology and Pathogenesis

Alcoholic cirrhosis affects over 16 million people globally and is responsible for 80% of deaths secondary to liver failure [119]. Of chronic alcohol users, 10–20% will develop cirrhosis, of which 10% will develop HCC [120]. In the United States, approximately 7% of the adult population meets criteria for alcohol abuse/dependence, which is five times higher than the prevalence of HCV [119]. The National Institute on Alcohol Abuse and Alcoholism estimates that 26.9% of Americas greater than 18 years old engage in binge drinking each month [121]. Although alcohol intake information is prone to bias, error, and underreporting, studies have shown that the risk development for HCC increases when daily alcohol intake is chronic and exceeds six drinks per day for more than 10 years [120]. Moreover, heavy drinkers have a 2–23-fold increased risk of death from cirrhosis compared to the general population [122]. Females are more sensitive to alcohol-mediated hepatotoxicity, possibly due to slower rates of alcohol metabolism and greater exposure to hepatotoxic alcohol byproducts [119, 123].

Multiple pathogenic mechanisms contribute to the development of alcohol-related HCC [9]. In hepatocytes, alcohol is initially metabolized by alcohol dehydrogenase (ADH) to acetaldehyde, which in excess quantities can induce hepatocyte DNA adducts [124]. The ADH reaction also increases the NADH/NAD+ ratio, causing an intracellular redox shift that transiently disrupts the TCA cycle, inducing lipogenesis and hepatocyte steatosis [125]. Over time, chronic alcohol intake also activates a supplementary ethanol breakdown pathway called the microsomal ethanol oxidizing system (MEOS), which generates high quantities of reactive oxygen species, resulting in oxidative stress, lipid peroxidation, and DNA mutations [126]. Collectively, these processes overwhelm the hepatic glutathione system and establish a milieu of hepatocyte death, chronic inflammation, and fibrogenesis, which may ultimately progress to steatohepatitis and eventually cirrhosis and HCC [126]. Finally, when chronic excessive alcohol intake occurs concomitantly with underlying metabolic liver disease or viral infection, the risk of cirrhosis and HCC are magnified presumably due to a synergistic injurious effect on hepatocytes [127, 128].

Alcohol Prevention

Alcohol cessation is the most effective treatment for alcohol-related hepatic injury, which may be accomplished through a combination of psychosocial interventions and pharmacological therapy [129]. Sobriety is challenging to both obtain and maintain, though each subsequent year of abstinence provides additional protection from alcohol-related comorbidities. It has been difficult to quantify the absolute HCC risk reduction with abstinence, which has been estimated at 4–7% per year, though individuals who maintain sobriety for greater than 10 years have a significantly lower risk of developing HCC compared to continued drinkers [130–132].

Critical to the abstinence process is first identifying at risk individuals and providing intervention. Screening and brief interventions (SBIs) in primary care clinics have been shown to be both effective and cost-effective in this regard [133–135]. Multiple intervention styles have been developed and trialed, though in general they involve advice counseling regarding the risks of heavy alcohol consumption by a healthcare professional. Kaner et al. recently published a meta-analysis of 34 studies (n = 15,197) providing moderate-quality evidence that patients receiving brief interventions in the primary care setting consume less alcohol than individuals receiving minimal or no intervention after 1 year [136]. However, the long-term beneficial health implications are more challenging to quantify at a population level. In a recent multi-institutional analysis of SBI practices in multiple European countries, it was observed that screening was executed in only 5.3% of over 6000 participants, which varied greatly by region (mean 1.7% Poland, 9.8% Sweden) [137]. Of the screened positive, interventions were also variably administered, ranging from 59.2% in Catalonia to 94.2% in Poland. The authors concluded that despite policy changes, increased awareness education, and SBI training among healthcare professionals in the studied regions over the last decade, screening and intervention frequency has unfortunately remained largely unchanged, suggesting improvements in this regard are still greatly needed.

Another historically common approach to alcohol cessation has been the use of pharmacologic agents that target the metabolism of ethanol. Disulfiram was the first FDA-approved medication for alcohol abuse, which functions by inhibiting acetaldehyde dehydrogenase, resulting in a buildup of the caustic metabolite, acetaldehyde, with alcohol consumption. Elevated acetaldehyde concentrations are associated with nausea, vomiting, headache, and severe physical discomfort. Thus, the function of disulfiram is to associate an undesirable physiologic response with alcohol consumption in order to establish a psychologic aversion [129]. Disulfiram has been shown to effectively reduce relapse rates; however, patient adherence to therapy is poor, limiting its widespread utility.

Currently, there are over 700 clinical trials registered in the United States for the treatment of alcoholism, many of which involve a combined approach of pharmacologic therapy with cognitive behavioral therapy (CBT). Pharmacologic agents under investigation include naltrexone, acamprosate, prazosin, doxazosin, propranolol, psilocybin, varenicline, odansetron, antidepressants (sertraline, escitalopram, duloxetine, mirtazapine), anticonvulsants (topiramate, zonisamide, levetiracetam), antipsychotics (aripriprazole, quetiapine, olanzapine), and analgesics (pregabalin, gabapentin, baclofen, ketamine). Many abstinence interventions are effective for fractional subsets of the population, and one of the challenges going forward is tailoring therapy based on the likelihood of success. Unfortunately, many of these drugs have potential hepatoxicity and are contraindicated in patients with advanced liver disease [129]. Baclofen, a selective GABA_b receptor agonist, is the only pharmacological therapy for alcoholism that has been approved for use in patients with advanced liver disease [138]. Baclofen was shown to be superior to placebo for maintaining abstinence in alcoholics with cirrhosis, which was associated improved liver function tests after 12 weeks of treatment [139]. Baclofen has also been shown to have a HCC preventive effect via induction of hepatocyte cell cycle arrest [140].

NAFLD, Metabolic Syndrome, Obesity Epidemiology, and Pathogenesis

Nonalcoholic fatty liver disease (NAFLD) is the most common form of chronic liver disease in developed nations with an estimated prevalence of 30%. NAFLD is defined as hepatic steatosis (HS) in the absence of excessive alcohol consumption. Although it may occur in lean patients, this condition is highly associated with obesity (up to 90% of patients) [141]. NAFLD is also associated with HCC risk, and the incidences of both conditions are rising such that the decreases in virus-related liver cancer might be offset by the rise of fatty liver disease [142, 143].

A subset of NAFLD patients will go on to develop hepatic inflammation in association with steatosis, a condition referred to as nonalcoholic steatohepatitis (NASH). In the United States, NASH effects approximately 3–8% of the population [141]. The transition from NAFLD to NASH is poorly understood, though NASH is a more severe condition in which patients are at increased risk for the development of cirrhosis or HCC. Liver biopsy is required for a diagnosis of NASH, which few patients undergo. Unfortunately, there are currently no reliable noninvasive tests to establish a diagnosis of NASH, and thus the true prevalence of the disease remains unknown, though one study proposed that NASH affects 20–25% of NAFLD patients [144]. Due to these diagnostic limitations, the NAFLD/NASH global burden is likely underestimated. In a more recent meta-analysis, it was estimated that the NASH prevalence among biopsied NAFLD patients is approximately 59.1%, nearly three times higher than what the previous studies have shown [143]. In addition, NASH has been proposed to account for a large proportion of idiopathic or cryptogenic cirrhosis (CC) cases, in which most of the histological hallmarks of NASH are no longer present by the time of presentation, often because patients have lost considerable weight with progressive liver failure. In CC patients who undergo liver transplantation, 25% develop NAFLD and 16% develop NASH within 26 months of transplant [145].

Most patients diagnosed with NASH have stable disease (34–50%) or might even observe improvements in their NAS score (18–29%) over time [146]. However, approximately 9–20% of NASH patients develop an aggressive form of disease that will progress to cirrhosis, and 4–27% go on to develop HCC [147]. Furthermore, common NASH comorbidities including elevated BMI and diabetes have been shown to be independent risk factors for HCC development. Historically, in the United States, the majority of HCC cases have been attributed to chronic HCV infection. However, 15–50% of new HCC cases are idiopathic, and there is reason to believe that NASH accounts for a large proportion of these [148]. A study examining 641 patients with HCC revealed that that clinical features of NASH, including obesity, diabetes, dyslipidemia, and elevated glucose were all significantly associated with CC-related HCC [149]. It is important to note that many of these clinical features, e.g., obesity and diabetes, are themselves independent risk factors for HCC and reports have stated that their coexistence with NASH-related cirrhosis has an additive effect on HCC risk.

The development of HCC in the context of NAFLD/NASH is a complex process with many contributing factors that can be broadly categorized into three main pathogenic mechanisms: cytokine and hormone dysregulation, lipotoxicity, and fibrosis. Obesity is highly prevalent in the NAFLD population and is also an independent risk factor for carcinogenesis by fostering a low-grade chronic inflammatory state throughout the body [150]. Hormonal dysregulation of leptin, adiponectin, and insulin has been implicated with adipose tissue expansion. Leptin is a proinflammatory, proangiogenic, and profibrogenic hormone with growth-promoting effects mediated by JAK/STAT, PI3K/AKT, and ERK signaling pathways [151]. Adipose content and leptin levels are positively correlated, and leptin has been shown to activate hepatic Kupffer and stellate cells, both of which have been connected to NAFLD/NASH fibrotic disease progression and HCC development [152]. In contrast, adiponectin is an anti-inflammatory hormone that is reduced in NAFLD. Low levels of adiponectin reduce its regulatory impact on inflammatory cell signaling, the mTOR mitogenic pathway, and angiogenesis, all of which promote hepatic oncogenesis [153]. Finally, insulin resistance and subsequent hyperinsulinemia are commonly observed in states of obesity and NAFLD. Elevated insulin levels upregulate the production of IGF1, which stimulates hepatic cellular proliferation and inhibits apoptosis through downstream activation of various oncogenic pathways including MAPK, PI3K/AKT, and VEGF [154]. Taken together, NAFLD-associated hormonal dysregulation supports proinflammatory and prosurvival signaling while downregulating checkpoint signals, establishing an environment in which hepatocyte malignant transformation is likely to occur unchecked.

Hepatocytes are capable of de novo lipogenesis (DNL), both as a means of lipid production for other tissues and for hepatocyte metabolism. Aberrantly elevated DNL, which is observed in NASH, has recently been implicated as a pathogenic mechanism that contributes to HCC carcinogenesis. Specifically, it has been postulated that upregulated DNL might provide necessary energy for the growing microenvironment of pre-cancerous lesions [155, 156]. Moreover, tumor mRNA expression of prolipogenic genes such as acetyl CoA carboxylase and fatty acid synthase correlates with cell proliferation rates and poor HCC prognosis [156, 157]. Another consequence of aberrant DNL is lipotoxicity, in which excessive mitochondrial B-oxidation and degradation of free fatty acid intermediates lead to reactive oxygen species generation, subsequently inducing ER stress, inflammation, and DNA damage. Obese mouse models have demonstrated that ROS production increases with hepatic fatty infiltration and is associated with the development of hepatic hyperplasia and dysplasia, both of which precede development of invasive malignancy [158, 159]. Lipid accumulation also increases lipid breakdown products, some of which can cause direct damage to hepatocytes. Trans-4-hydroxy-2-noneal is a byproduct of lipid peroxidation and has been shown to cause mutations within the TP53 tumor suppressor gene, a common mutation involved in over 50% of HCC cases [160]. NRF1 is an essential transcription factor that mediates oxidative stress in hepatocytes, which is downregulated in animal models of NASH. Decreased NRF1 expression correlated with hepatic steatosis, inflammation, fibrosis, and HCC development [161].

Persistent lipotoxic hepatocyte injury is associated with constitutive activation of tumor necrosis factor alpha (TNFα) and interleukin-6 (IL-6), driving chronic hepatic inflammation [150]. TNFα is a potent activator of multiple pro-oncotic pathways that involve mTOR, JNK, and NF-κB [162]. IL-6 is an inflammatory cytokine that exhibits antiapoptotic and cell-proliferative phenotypes through the activation of STAT3, a transcriptional factor linked to malignant transformation and HCC aggressiveness [163, 164]. Experimental mouse models of dietary obesity have been linked to the activation of IL-6, TNFα, and their associated oncogenic signaling pathways, which was associated with increased HCC risk [162, 163]. Chronic hepatic inflammation is also associated with hepatic stellate cell activation and fibrogenesis, which may progress to cirrhosis. It is unclear whether hepatic fibrosis is an independent risk factor for HCC development, or rather that fibrosis correlates with advanced parenchymal disease. However, more than one-third of the NASH-related HCC occurs in the absence of cirrhosis [144]. Noncirrhotic HCC patients are typically older, suggesting there may be differences in disease biology [165].

NASH Prevention

A major challenge in engineering therapies for metabolic disorders is creating a treatment that targets a multifaceted disease. Although multiple medications targeting lipid homeostasis and glucose metabolism are currently in use to treat obesity and diabetes, there are currently no FDA-approved drugs that specifically target NASH. To date, the only accepted treatments for NASH are weight loss by lifestyle modification or bariatric surgery. In a recent analysis of patients treated with weight loss interventions, individuals capable of losing at least 5% of their body weight showed a significant reduction of hepatic steatosis. Weight loss of greater than 7% was associated with improvement in NALFD Activity (NAS) score, and weight loss greater than 10% was associated with improvement in histologic features of NASH, including portal inflammation and fibrosis [166, 167]. In addition, the type of diet appears to be less important than sustained weight loss for the resolution of NAFLD [168]. Although the majority of obese and NAFLD patients participate in minimal physical activity, large randomized control trials assessing the direct effect of exercise on NASH are missing. Retrospective evidence has suggested that exercising five times per week for a minimum of 10 min was linked to decreases in new fatty liver deposition and improvements in existing liver disease [169].

Bariatric surgery has emerged as an effective alternative weight loss treatment across the spectrum of NAFLD/NASH patients. A meta-analysis of 15 studies researching the effects of bariatric surgery in NASH showed improvements in steatosis (91.6%), steatohepatitis (81.3%), fibrosis (65.5%), and disease resolution (69.5%) after surgery [170]. Two subsequent prospective studies in obese patients with NASH saw disease resolution in 85% of cases at 1 year after surgery and 69.7% after 5 years [171, 172]. Despite evidence to support its efficacy, bariatric surgery is currently restricted to patients with a body mass index ≥40 or ≥35 with obesity-related comorbidities. Most patients who receive bariatric surgery retain weight loss rates between 14% and 25% below their preoperative weight after 10 years, and they show improvements in diabetes, insulin resistance, and cardiovascular events [173–175]. Given the strong association between weight loss and improvement in NASH severity, expanding indications for bariatric surgery to include NASH in patients with elevated BMI might be of value. The hesitation in doing so is partly due to a lack of high-level evidence that establishes a direct connection between bariatric surgery and NASH [176]. The effect of bariatric surgery on hepatic fibrosis is also controversial. Initial studies suggested that hepatic fibrosis increased after surgery due to increases in proinflammatory cytokines [177]. However, recent evidence has shown that 80–95% of patients see no change or regression in their hepatic fibrosis status at 5 years post-op [172]. This may be in part due to a reduction in hepatic profibrogenic cytokine gene expression, thereby attenuating hepatitis and subsequent fibrosis [178]. Finally, there is concern regarding the risk of surgery in patients with intrinsic liver disease. A recent study by Jan et al. showed no difference in complication rate after bariatric surgery in patients with compensated cirrhosis compared to those without the evidence of liver disease [179–182]. However, decompensated cirrhosis remains a contraindication to bariatric surgery [180, 182, 183].

Similar to bariatric surgery, it is thought that medications targeting metabolic disorders may have secondary benefits on HCC prevention. Metformin, an insulin sensitizer used as first-line treatment for hyperglycemia and noninsulin-dependent type 2 diabetes, is currently under investigation as a possible HCC preventative agent. Retrospective studies and pooled analysis have shown that metformin decreased HCC incidence up to 50% when compared to observation alone [184, 185]. However, RCT examining the effects of metformin on HCC development does not support the claim that metformin is superior to other antidiabetic agents in reducing HCC [186].

Pioglitazone , a peroxisome proliferator-activated receptor gamma (PPARδ) activator, is another diabetes medication that showed promising results as an anti-NASH agent. Several RCTs in diabetic and nondiabetic populations consistently showed improvements in NASH histological parameters (steatosis, parenchymal and lobular inflammation, hepatocyte ballooning, and NAS score), which are believed to indirectly reduce the risk of HCC [187–191]. Despite these promising results, multiple studies have associated pioglitazone with an unfavorable side effect profile including fluid retention and weight gain, limiting its widespread use [192–194].

Statins are commonly used to reduce the risk for cardiovascular disease and diabetes; however, there has been evidence to support their use as a chemopreventive agent in a variety of cancers [195]. Statins reduce blood cholesterol by inhibiting HMG-CoA reductase, which consequently decreases the production of mevalonate pathway metabolites, a pathway recently implicated in cell growth and cancer transformation [196]. Singh et al. investigated the effects of statins on the risk of HCC by conducting a meta-analysis that included 26 randomized control trials and over 1.4 million patients [197]. They found that statin use was associated with a 37% decrease in the risk of HCC after adjusting for confounding variables. Furthermore, a recent retrospective study of nearly 10,000 patients showed that the beneficial effect of statin use on HCC chemoprevention was greater in patients with diabetes or cirrhosis [198]. However, one RCT did not demonstrate a significant difference in HCC incidence between statin and placebo groups [199]. Further investigation regarding the mechanism of statin-mediated anticancer effects remains ongoing.

More recently, a plethora of drugs designed to specifically target NASH pathogenesis are in clinical development (Table 13.3). Given the complex pathophysiology of NASH, these drugs have been designed to have either metabolic, antisteatotic, anti-inflammatory, or antifibrotic effects. It is expected that drugs that can impede several of these disease mechanisms or drug combinations that can successfully inhibit multiple pathways will have the most beneficial effects for patients. Obeticholic acid (OCA) was the first such drug designed for NASH patients and is a Farnesoid X Receptor agonist that functions to reduce the conversion of cholesterol to bile acids in the liver, which has been associated with reductions in cholestasis, hepatic inflammation, hepatocyte injury, and HCC development. In a phase II clinical trial, OCA was associated with significant reductions in histologic markers of NASH [200, 201]. OCA is being subsequently investigated in a phase III clinical trial (REGENERATE; NCT02548351), examining the long-term effects of OCA on fibrosis and mortality in NASH patients. In the 18-month interim analysis, OCA 25 mg once daily met the primary endpoint of fibrosis improvement (≥1 stage) without worsening NASH.

Table 13.3

Clinical trials for the pharmacological therapies for NASH

Elafibranor is a dual PPARα/δ agonist that functions by decreasing serum glucose and triglyceride levels and has also been shown to have anti-inflammatory properties. Elafibranor was studied in a phase IIa clinical trial (GOLDEN; NCT01694849) of NASH patients in which it was associated with reductions in hepatocyte ballooning, lobar inflammation, fibrosis staging, and cardiometabolic risk profile [202]. These improvements were most evident in patients with NAS scores ≥4. Importantly, unlike some PPARα/δ agonists, there has been no evidence to suggest that Elafibranor causes weight gain. Elafibranor is currently in a phase III clinical trial (RESOLVE-IT; NCT02704403) investigating its impact on fibrosis staging after 72 weeks of treatment, as well as rates of mortality, cirrhosis, and liver-related comorbidities up to 4 years after therapy.

Cenicriviroc is a CCR2/5 inhibitor which efficiently inhibits monocyte infiltration and chemokine activation. CCR2 is upregulated in fibrotic human livers alongside an accumulation of monocyte-derived phagocytes [203]. In obese NASH patients, proportions of CCR2+ macrophages in visceral adipose tissue are associated with histological disease severity [204]. Clinical trials testing cenicriviroc in the NASH population showed decreases in hepatic fibrosis and slowing NASH progression [205]. Cenicriviroc is in a phase III clinical trial (AURORA; NCT03028740) examining its impact on fibrosis staging after 12 months of treatment and its effect on cirrhosis rates, liver related clinical outcomes, and mortality 5 years after therapy.

Selonsertib is an apoptosis signal-regulating kinase 1 (ASK-1) inhibitor designed to treat NASH. Activation of ASK-1 is normally achieved by TNF-alpha or cellular stress (oxidative or ER), leading to activation of p38/JNK pathway and subsequent hepatic fibrosis and apotosis [206]. In a murine model of NASH, selonsertib improved metabolic parameters (serum cholesterol and bile acid), hepatic steatosis, inflammation, and fibrosis [207]. Moreover, combination therapy with simtuzumab, a lysyl oxidase-like molecule 2 (LOXL2) inhibitor, potentiated the antifibrotic effects of anti-ASK and anti-LOXL2 monotherapy in mice [208]. This evidence warranted a phase II clinical trial testing the effect of selonsertib in NASH which showed significant reduction in hepatic fibrosis, lobular inflammation, and serum biomarkers for apoptosis (cytokeratin-18 M30) and necrosis (cytokeratin-18 M65) after 24 weeks of treatment [209]. Selonsertib is being evaluated in a phase III clinical trial (STELLAR 4; NCT03053063) evaluating its ability to improve hepatic fibrosis and survival in patients with NASH-related compensated cirrhosis, although the primary endpoint for fibrosis was not met.

Tertiary Prevention

Curative treatment options for HCC are limited due to the lack of effective chemotherapy and radiotherapy. Therefore, tumor resection and orthotopic liver transplantation (OLT) are the only options for long-term cure. Although advances in these surgical procedures have significantly improved perioperative morbidity, over 70% of patients still develop intrahepatic recurrence within 5 years of their first hepatectomy [221]. HCC recurrence rates among liver transplant recipients is varied between centers (6.4–56.5%; 5-year recurrence rates); however, tumor size, nodule count, vascular invasion, presence of cirrhosis, and tumor grade have emerged as the most clinically predictive characteristics for recurrence [222].

Tertiary HCC prevention aims to prevent cancer recurrence in patients curatively treated for initial cancer as adjuvant therapy [24]. Various strategies to prevent the recurrence of HCC have been tried, such as vitamin K2, retinoids, and systemic chemotherapy, although none were proven to be effective when tested in large-scale randomized control trials [223–225]. Interferon is the most widely used adjuvant tertiary prevention therapy, although evidence to support its effectiveness in reducing recurrence in conflicting [226]. Creating tertiary prevention strategies for HCC is a major challenge due to its intrinsic chemoresistance which has been linked to various mechanisms including overexpression of drug efflux pumps (MDR1 and MRP2), enhanced DNA repair mutations (ERCC-1, FENs, Chk2, ATM, APE1), impairment of apoptotic machinery (CD95, FADD, FLICE), and activation of cell survival signaling (Hedgehog, Hippo, Wnt/beta-catenin) [227]. Unlike other cancers where metastasis is the primary method for recurrence, HCC has a heightened risk of de novo carcinogenesis especially patients with cirrhosis or viral infection. Moreover, the majority of HCC recurrence occurs in the liver remnant, making it nearly impossible to differentiate its origin. For these reasons, it has been difficult to study and find an agent that can inhibit both HCC recurrence mechanisms.

A multityrosine kinase inhibitor, sorafenib, inhibits cell proliferation and angiogenesis in murine models and therefore may reduce HCC recurrence [228]. A phase III clinical trial (STORM) showed sorafenib did not reduce recurrence-free survival compared to control and deemed it ineffective as an adjuvant intervention for HCC following resection or ablation [229]. Although HCC is not normally considered an immunogenic tumor, it has been reported that patients with higher levels of lymphocytes within their tumors have longer survival rates and are at lower risk of recurrence [230, 231]. Patients expressing higher numbers of tumor-associated antigens (TAA) also had better survival rates than those with fewer TAAs indicating a role for a patient’s own immune system in fending off HCC-related comorbidities [231]. These results provide the rationale for immunotherapy as a tertiary prevention against HCC recurrence. Immune checkpoint blockade by anti-PD-1 antibodies, nivolumab and (CHECKMATE 040; NCT01658878) and pembrolizumab (KEYNOTE-224; NCT02702414), demonstrated objective response rates of up to 20% in the setting of oncology treatment [232, 233]. Based on the result, immune checkpoint inhibition is under evaluation as adjuvant therapy following HCC resection or ablation (Checkmate 9DX; NCT03383458). A combination of a VEGF inhibitor, bevacizumab, and an anti-PD-L1 antibody, atezolizumab, yielded a high objective response rate of 32% and a 6-month progression free survival of 65% (NCT02715531) [233]. This suggests that combination therapy may also serve as adjuvant therapy in HCC. Cytokine-induced killer (CIK) cell-based therapy is one of the promising adjuvant immunotherapies tested in HCC. CIK cells are a mixture of T cells (CD3+/CD56+ cells and CD3+/CD56− T cells and CD3−/CD56+ natural killer cells) that are ex vivo expanded using cytokines. Preclinical studies showed CIK cells have multiple favorable characteristics including potent in vitro HCC-targeted damage, ability to localize within hepatic cancer mass in vivo, and no major side effects with repeated therapies [234–236]. These encouraging preclinical studies allowed for the phase III clinical trial showing that activated CIK treatment in patients previously treated with HCC surgical resection, radiofrequency ablation, or percutaneous ethanol injection extended median recurrence-free and overall survival compared to placebo [237]. These results have continued to hold as Lee and colleagues recently reported significant improvements in 5-year recurrence-free and overall survival rates [238]. Other immunotherapy treatments attempt to treat HCC using more targeted forms of therapy including utilizing CAR-T cells, HCC vaccines, and altering immune checkpoint inhibitors. These types of therapies have been designed to treat existing HCC; however, it is conceivable for them to be used to attenuate the risk of recurrence due to their anticarcinogenesis effects [239].

Conclusions and Future Directions

Given the lack of successful treatment options for HCC, prevention should be given more attention. Primary prevention strategies have already shown remarkable success in reducing HCC incidence. For example, after decades of research, DAAs have emerged as a promising approach to reduce HCC incidence in the majority of HCV patients. Risk of HCC does still persist especially in those patients with advanced fibrosis at the time of treatment, so screening strategies will need to be implemented for these patients. While a vaccine for HCV remains elusive, efforts in this area remain ongoing and could offer an additional solution for cure. Likewise, HBV vaccination has proven to be a very effective strategy for reducing HCC incidence in places where HBV is endemic. Unfortunately, socioeconomic concerns have lagged behind the science in these instances as not all people at risk for HBV infection have access to adequate healthcare with HBV vaccination programs and similarly not all HCV patients currently have access to DAAs given their high cost. Solutions for these problems, like midwife vaccination programs, will need to be continually evaluated in the coming years.

It is not as clear whether alcohol cessation or changes in eating habits and weight loss will be as sustainable in reducing HCC risk as a result of ASH and NASH, respectively. Alternative solutions like liver transplantation and bariatric surgery exist but also come with a high socioeconomic cost. A plethora of trials are underway to evaluate new drugs that target different pathways in the disease pathogenesis including insulin resistance, de novo lipogenesis, inflammation, and fibrogenesis, but more efforts should be deployed for repurposing current drugs, like metformin, which have been associated with decreased HCC incidence in retrospective analyses.

Given the long duration between exposure to insult and development of primary HCC, investigation of preventative therapies will most likely occur in the tertiary setting and/or secondary setting with enrichment of high-risk cirrhosis population [8]. Trial designs could include DNA sequencing to verify that recurrent tumors are in fact de novo cancers as opposed to regrowth of a previous unsuccessfully treated tumor. In addition, both invasive, like gene signatures, and noninvasive biomarkers, like serum proteins or DNA, could be evaluated for their ability to predict successful therapies. Such biomarkers would be instrumental in the subsequent evaluation of promising therapies into the primary setting.

In conclusion, given the readily identifiable population at risk, HCC prevention is an achievable goal as supported by numerous successful programs to date. Given the increasing incidence and the lack of effective treatments, more efforts in HCC prevention should be undertaken to improve the prognosis of this deadly disease.

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