Heterogeneity of hepatocellular carcinoma: from mechanisms to clinical implications.

Hepatocellular Carcinoma (HCC) is one of the most common types of primary liver cancer. Current treatment options have limited efficacy against this malignancy, primarily owing to difficulties in early detection and the inherent resistance to existing drugs. Tumor heterogeneity is a pivotal factor contributing significantly to treatment resistance and recurrent manifestations of HCC. Intratumoral heterogeneity is an important aspect of the spectrum of complex tumor heterogeneity and contributes to late diagnosis and treatment failure. Therefore, it is crucial to thoroughly understand the molecular mechanisms of how tumor heterogeneity develops. This review aims to summarize the possible molecular dimensions of tumor heterogeneity with an emphasis on intratumoral heterogeneity, evaluate its profound impact on the diagnosis and therapeutic strategies for HCC, and explore the suitability of appropriate pre-clinical models that can be used to best study tumor heterogeneity; thus, opening new avenues for cancer treatment.

Aptamer-mediated doxorubicin delivery reduces HCC burden in 3D organoids model.

Epithelial cell adhesion molecule (EpCAM) is a surface marker which is frequently overexpressed in hepatocellular carcinoma (HCC) but minimally expressed on mature hepatocytes. We developed a specific aptamer against EpCAM (EpCAM-apt) and tested its potential as a drug delivery agent for HCC. The targeting ability of EpCAM-apt was confirmed in vitro and in vivo after which the complex was conjugated with doxorubicin (Dox) to form EpCAM-apt-Dox. The targeting efficacy of the drug-loaded complex against liver cancer stem-like cells (LCSCs) and therapeutic effects in HCC were evaluated. EpCAM-expressing (EpCAM+) HCC cells showed characteristics of stem like cells including greater proliferative capacity and tumour sphere formation. EpCAM-apt-Dox selectively delivered Dox to EpCAM+ HCC cells with high drug retention and accumulation versus control. EpCAM-apt-Dox reduced the self-renewal capacity and stem-like cell frequency in vitro. Elimination of cancer stem-like cells (CSCs) with EpCAM-apt-Dox significantly inhibited the growth of HCC cells and patient-derived HCC organoids but exerted minimal cytotoxicity to normal liver organoids. Moreover, EpCAM-apt-Dox suppressed the growth of xenograft tumours derived from HCC organoids in vivo and prolonged mouse survival without inducing adverse effects to major organs. Thus, aptamer-based drug delivery to the stem-like cell population is a promising strategy for HCC treatment.

The Application of Induced Pluripotent Stem Cells Against Liver Diseases: An Update and a Review.

Liver diseases are a major health concern globally, and are associated with poor survival and prognosis of patients. This creates the need for patients to accept the main alternative treatment of liver transplantation to prevent progression to end-stage liver disease. Investigation of the molecular mechanisms underpinning complex liver diseases and their pathology is an emerging goal of stem cell scope. Human induced pluripotent stem cells (hiPSCs) derived from somatic cells are a promising alternative approach to the treatment of liver disease, and a prospective model for studying complex liver diseases. Here, we review hiPSC technology of cell reprogramming and differentiation, and discuss the potential application of hiPSC-derived liver cells, such as hepatocytes and cholangiocytes, in refractory liver-disease modeling and treatment, and drug screening and toxicity testing. We also consider hiPSC safety in clinical applications, based on genomic and epigenetic alterations, tumorigenicity, and immunogenicity.

Developing liver organoids from induced pluripotent stem cells (iPSCs): An alternative source of organoid generation for liver cancer research.

Primary liver cancer (PLC) represents a significant proportion of all human cancers and constitutes a substantial health and economic burden to society. Traditional therapeutic approaches such as surgical resection and chemotherapy often fail due to tumour relapse or innate tumour chemoresistance. There is a dearth of efficient treatments for PLC in part due to the poor capacity of current laboratory models to reflect critical features of the native tumour in vivo. The increasing incorporation of organoid systems has led to a resurgence of interest in liver cancer research. Organoid systems show promise as the gold standard for recapitulating tumours in vitro. Further, developments in culturing techniques will improve the various shortcomings of the current systems. Induced pluripotent stem cell (iPSC)-derived liver organoids are a promising alternative to the conventional liver organoid model as it circumvents the need to rely on primary resections which are often scarce. In this concise review, we will discuss novel techniques for organoid culture with a focus on organoid co-cultures and their advantages over traditional organoid systems. A detailed technical protocol for the generation of iPSC-derived liver organoids is provided as an appendix.

Role of the constitutive androstane receptor (CAR) in human liver cancer.

BACKGROUND: The constitutive androstane receptor (CAR) is a member of the nuclear receptor superfamily (subfamily 1, group I, member 3, also known as NR1I3) that is almost exclusively expressed in the liver. CAR interacts with key signalling pathways such as those involved in drug, energy and bilirubin metabolism. In mouse models, activation of CAR leads to tumorigenesis by inducing pro-proliferative and anti-apoptotic signalling. However, many previous reports have shown species differences between CAR activity in animal models and humans. Recent studies have demonstrated that the mode of action of CAR in rodent liver tumorigenesis is not applicable to humans. Despite this, many studies still continue to study the role of CAR in animal models, hence, there is a need to further explore the role of CAR in human diseases particularly cancers. While there is limited evidence for a role of CAR in human cancers, some studies have proposed a tumour-suppressive role of CAR in liver cancer. In addition, recent studies exploring CAR in human livers demonstrated a hepato-protective role for CAR in and more specifically, its ability to drive differentiation and liver regeneration. This review will discuss the role of CAR in liver cancer, with a focus on species differences and its emerging, tumour-suppressive role in liver cancer and its role in the regulation of liver cancer stem cells.

An aptamer-based drug delivery agent (CD133-apt-Dox) selectively and effectively kills liver cancer stem-like cells.

Liver cancer has no effective therapies, hence a poor survival. Cancer stem-like cells not only contribute to cancer initiation and progression, but also to drug resistance, cancer metastasis, and eventually treatment failure. Hence, any approaches that can effectively kill cancer stem-like cells hold a great potential for cancer treatment. CD133 is a robust marker for liver cancer stem-like cells. We developed a specific aptamer against CD133 (CD133-apt), and then loaded this aptamer with an anticancer drug doxorubicin (CD133-apt-Dox). The efficacy of CD133-apt-Dox in targeting liver cancer stem-like cells and its overall effect in treating liver cancer were investigated using multiple in vitro and in vivo studies including in patients-derived liver cancer organoids. We have observed that CD133-apt could preferably delivered doxorubicin to CD133-expressing cells with efficient drug accumulation and retention. CD133-apt-Dox impaired the self-renewal capacity of liver cancer stem-like cells and attenuated their stem-ness phenotypes in vitro or in vivo. CD133-apt-Dox significantly inhibited the growth of liver cancer cells and patients-derived organoids and reduced the growth of xenograft tumours in nude mice inhibited the growth of DEN-induced liver cancer in immunocompetent mice. Hence, aptamer-mediated targeting of CD133 is a highly promising approach for liver cancer therapy.

Application of organoids in translational research of human diseases with a particular focus on gastrointestinal cancers.

Gastrointestinal (GI) cancers constitute the largest portion of all human cancers and represent a significant health burden on modern society. Conventional therapeutic approaches such as chemotherapy and surgical resections often fail due to poor treatment response or tumour relapse. Unfortunately, drug discovery for GI cancers has stalled as current cancer models fail to recapitulate critical features of the parent tumour, leading to poor translation from bench to bedside. Recent advances in three-dimensional (3D) cell culturing techniques have driven the surge of interest in stem cell-derived organoid models, a promising platform with a plethora of potential applications due to its ability to retain crucial architectural, genomic and transcriptional properties of the native tissue. In this review article, we discuss current applications and advantages of organoid models in the translational research of GI cancers with a particular focus on primary liver cancer that currently lack effective curative treatments.