"Complimenting the Complement": Mechanistic Insights and Opportunities for Therapeutics in Hepatocellular Carcinoma

Abstract

Hepatocellular carcinoma (HCC) is the most common primary malignancy of the liver and a leading cause of death in the US and worldwide. HCC remains a global health problem and is highly aggressive with unfavorable prognosis. Even with surgical interventions and newer medical treatment regimens, patients with HCC have poor survival rates. These limited therapeutic strategies and mechanistic understandings of HCC immunopathogenesis urgently warrant non-palliative treatment measures. Irrespective of the multitude etiologies, the liver microenvironment in HCC is intricately associated with chronic necroinflammation, progressive fibrosis, and cirrhosis as precedent events along with dysregulated innate and adaptive immune responses. Central to these immunological networks is the complement cascade (CC), a fundamental defense system inherent to the liver which tightly regulates humoral and cellular responses to noxious stimuli. Importantly, the liver is the primary source for biosynthesis of >80% of complement components and expresses a variety of complement receptors. Recent studies implicate the complement system in liver inflammation, abnormal regenerative responses, fibrosis, carcinogenesis, and development of HCC. Although complement activation differentially promotes immunosuppressive, stimulant, and angiogenic microenvironments conducive to HCC development, it remains under-investigated. Here, we review derangement of specific complement proteins in HCC in the context of altered complement regulatory factors, immune-activating components, and their implications in disease pathogenesis. We also summarize how complement molecules regulate cancer stem cells (CSCs), interact with complement-coagulation cascades, and provide therapeutic opportunities for targeted intervention in HCC.

Keywords: HCC and COVID-19; complement activation; complement proteins; complement-based therapeutics; hepatocellular carcinoma; immunotherapy; inflammatory factors; prognostic markers.

Figure 1

Immunopathogenic complement activation regulates progression to hepatocellular carcinoma. Exposure of the hepatic milieu to several triggers linked either to viral infections (Hepatitis B virus, Hepatitis C virus, etc.), severe obstructive and cholestatic diseases (Biliary atresia, Primary Sclerosing Cholangitis, Primary Biliary Cirrhosis, etc.), environmental stressors/toxin exposures (polychlorinated biphenyls, arsenic, androgenic steroids, etc.), and other etiopathogenic agents (aflatoxins, oral contraceptives, vinyl chloride, etc.) dictate the evolution of hepatocellular carcinoma (HCC). These triggers activate the innate immune complement cascade via classical (involving C1q complex), alternative (C3b-dependent activation), or lectin (triggered by carbohydrates) pathways. Abnormal activation of these complement pathways modulates functional effects of intrahepatic immune and epithelial cell compartments and disseminates significant perturbation of effector innate and adaptive cells, cytokine and chemokine expressions, and sustained cancer stem cell (CSC) activities. The collective net result of these processes defines the progression of HCC tumorigenesis.

Figure 2

Aberrant complement activation: a driver for disease progression in hepatocellular carcinoma. A schematic of the liver microenvironment depicting the transition of a healthy immunologically quiescent intrahepatic microenvironment to dysregulated immune status following activation cues to the complement system. In a healthy liver, immune and epithelial cells function in synergy to preserve normal architecture of bile ducts, quiescence of complement molecules, and homeostasis of immune cells. Complement activating disease-triggering signals orchestrate the evolution of dysregulated complement molecules (increased C3, C5, etc) and altered complement regulatory factors (CFH, etc). These acute and/or chronic sustenance of dysregulated complement molecules and their complex interaction with the immune and epithelial cell compartments drive the progression of hepatocellular carcinoma (HCC). Loss of complement regulatory factors and divergent activation of the complement system leads to abnormal hepatocyte architecture, deranged cellular and effector functions, and reactive bile duct profiles. Cumulatively, these events lead to epithelial to mesenchymal transition (EMT) and tumor angiogenesis which worsen the disease, resulting in poor clinical outcomes and death.

Figure 3

Dysregulated expressions of complement components orchestrate the pathobiology of hepatocellular carcinoma. Breakdown of the tightly controlled activation and regulatory component signals of the complement system results in dysregulation of the normal homeostatic cellular processes within the liver microenvironment. Increased levels of C1q, C3, C5, C4BP, CD46, CD59, and MASP2 and decreased levels of C2, C4, C8A, CR1, MBL2, CFH, CFHR3, and CFHL1 with potential immunoregulatory functions contribute to increased cell proliferation, metastasis, EMT, altered immune cell functions, etc. resulting in recurrent episodes and/or poor overall or disease-free survival in patients with HCC. Images from Motifolio drawing toolkit software (http://motifolio.com) were used for rendering the figure.

Figure 4

Bioinformatics analysis identifies dominant signatures of immune–mediated biological processes. Supervised gene ontology (GO) annotation analysis of complement components in hepatocellular carcinoma (HCC) was performed via ToppGene Suite portal (http://toppgene.cchmc.org) for in-silico enrichment of biological processes with a threshold False Discovery Rate (FDR) corrected P value of <0.05. ToppGene Suite is a freely available online tool used for functional enrichment, prioritization of candidate genes using transcriptome, ontology, phenotype, proteome, and functional annotations. GO: biological processes identified by ToppGene were further subjected to functional enrichment using CIMminer (https://discover.nci.nih.gov/cimminer/home.do). Red areas depicted in the heatmap show closely related biological processes linked to immunity that are shared by a major group of complement molecules shown on the horizontal axis.

Figure 5

Protein-protein interaction network analysis identifies biological relatedness of complement components to regulators of hepatocellular carcinoma (HCC) pathogenesis. Complement molecules of relevance to HCC were subjected to protein-protein interaction (PPI) network analysis using the network-based gene prioritization algorithm, ToppGenet of the ToppGene Suite (http://toppgene.cchmc.org). ToppGenet identifies and prioritizes candidate genes based on functional annotations, similar expressions, and network and topographical features. A Step Size of 6 and the Prioritization method of k-Step Markov were used as default analytical parameters. The Cytoscape-compatible ToppGenet output file was used to generate the graphical network. The first shell of 41 interacting proteins (grey color) associated directly with the input complement proteins (blue) in the PPI were generated by Cytoscape.