Unraveling the Sweet Secrets of HCC: Glucometabolic Rewiring in Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) is the primary form of liver cancer. It causes ∼ 800 000 deaths per year, which is expected to increase due to increasing rates of obesity and metabolic dysfunction associated steatotic liver disease (MASLD). Current therapies include immune checkpoint inhibitors, tyrosine kinase inhibitors, and monoclonal antibodies, but these therapies are not satisfactorily effective and often come with multiple side effects and recurrences. Metabolic reprogramming plays a significant role in HCC progression and is often conserved between tumor types. Thus, targeting rewired metabolic pathways could provide an attractive option for targeting tumor cells alone or in conjunction with existing treatments. Therefore, there is an urgent need to identify novel targets involved in cancer-mediated metabolic reprogramming in HCC. In this review, we provide an overview of molecular rewiring and metabolic reprogramming of glucose metabolism in HCC to understand better the concepts that might widen the therapeutic window against this deadly cancer.

Liver-specific overexpression of HKDC1 increases hepatocyte size and proliferative capacity.

A primary role of the liver is to regulate whole body glucose homeostasis. Glucokinase (GCK) is the main hexokinase (HK) expressed in hepatocytes and functions to phosphorylate the glucose that enters via GLUT transporters to become glucose-6-phosphate (G6P), which subsequently commits glucose to enter downstream anabolic and catabolic pathways. In the recent years, hexokinase domain-containing-1 (HKDC1), a novel 5th HK, has been characterized by our group and others. Its expression profile varies but has been identified to have low basal expression in normal liver but increases during states of stress including pregnancy, nonalcoholic fatty liver disease (NAFLD), and liver cancer. Here, we have developed a stable overexpression model of hepatic HKDC1 in mice to examine its effect on metabolic regulation. We found that HKDC1 overexpression, over time, causes impaired glucose homeostasis in male mice and shifts glucose metabolism towards anabolic pathways with an increase in nucleotide synthesis. Furthermore, we observed these mice to have larger liver sizes due to greater hepatocyte proliferative potential and cell size, which in part, is mediated via yes-associated protein (YAP) signaling.

Aiding Cancer's "Sweet Tooth": Role of Hexokinases in Metabolic Reprogramming.

Hexokinases (HKs) convert hexose sugars to hexose-6-phosphate, thus trapping them inside cells to meet the synthetic and energetic demands. HKs participate in various standard and altered physiological processes, including cancer, primarily through the reprogramming of cellular metabolism. Four canonical HKs have been identified with different expression patterns across tissues. HKs 1-3 play a role in glucose utilization, whereas HK 4 (glucokinase, GCK) also acts as a glucose sensor. Recently, a novel fifth HK, hexokinase domain containing 1 (HKDC1), has been identified, which plays a role in whole-body glucose utilization and insulin sensitivity. Beyond the metabolic functions, HKDC1 is differentially expressed in many forms of human cancer. This review focuses on the role of HKs, particularly HKDC1, in metabolic reprogramming and cancer progression.

The hexokinase "HKDC1" interaction with the mitochondria is essential for liver cancer progression.

Liver cancer (LC) is the fourth leading cause of death from cancer malignancies. Recently, a putative fifth hexokinase, hexokinase domain containing 1 (HKDC1), was shown to have significant overexpression in LC compared to healthy liver tissue. Using a combination of in vitro and in vivo tools, we examined the role of HKDC1 in LC development and progression. Importantly, HKDC1 ablation stops LC development and progression via its action at the mitochondria by promoting metabolic reprogramming and a shift of glucose flux away from the TCA cycle. HKDC1 ablation leads to mitochondrial dysfunction resulting in less cellular energy, which cannot be compensated by enhanced glucose uptake. Moreover, we show that the interaction of HKDC1 with the mitochondria is essential for its role in LC progression, and without this interaction, mitochondrial dysfunction occurs. As HKDC1 is highly expressed in LC cells, but only to a minimal degree in hepatocytes under normal conditions, targeting HKDC1, specifically its interaction with the mitochondria, may represent a highly selective approach to target cancer cells in LC.

Gestational Insulin Resistance Is Mediated by the Gut Microbiome-Indoleamine 2,3-Dioxygenase Axis.

BACKGROUND & AIMS: Normal gestation involves a reprogramming of the maternal gut microbiome (GM) that contributes to maternal metabolic changes by unclear mechanisms. This study aimed to understand the mechanistic underpinnings of the GM-maternal metabolism interaction. METHODS: The GM and plasma metabolome of CD1, NIH-Swiss, and C57 mice were analyzed with the use of 16S rRNA sequencing and untargeted liquid chromatography-mass spectrometry throughout gestation. Pharmacologic and genetic knockout mouse models were used to identify the role of indoleamine 2,3-dioxygenase (IDO1) in pregnancy-associated insulin resistance (IR). Involvement of gestational GM was studied with the use of fecal microbial transplants (FMTs). RESULTS: Significant variation in GM alpha diversity occurred throughout pregnancy. Enrichment in gut bacterial taxa was mouse strain and pregnancy time point specific, with the species enriched at gestation day 15/19 (G15/19), a point of heightened IR, being distinct from those enriched before or after pregnancy. Metabolomics revealed elevated plasma kynurenine at G15/19 in all 3 mouse strains. IDO1, the rate-limiting enzyme for kynurenine production, had increased intestinal expression at G15, which was associated with mild systemic and gut inflammation. Pharmacologic and genetic inhibition of IDO1 inhibited kynurenine levels and reversed pregnancy-associated IR. FMT revealed that IDO1 induction and local kynurenine level effects on IR derive from the GM in both mouse and human pregnancy. CONCLUSIONS: GM changes accompanying pregnancy shift IDO1-dependent tryptophan metabolism toward kynurenine production, intestinal inflammation, and gestational IR, a phenotype reversed by genetic deletion or inhibition of IDO1. (Gestational Gut Microbiome-IDO1 Axis Mediates Pregnancy Insulin Resistance; EMBL-ENA ID: PRJEB45047. MetaboLights ID: MTBLS3598).

Hexokinase domain-containing protein-1 in metabolic diseases and beyond.

Glucose phosphorylation by hexokinases (HKs) traps glucose in cells and facilitates its usage in metabolic processes dependent on cellular needs. HK domain-containing protein-1 (HKDC1) is a recently discovered protein with wide expression containing HK activity, first noted through a genome-wide association study (GWAS) to be linked with gestational glucose homeostasis during pregnancy. Since then, HKDC1 has been observed to be expressed in many human tissues. Moreover, studies have shown that HKDC1 plays a role in glucose homeostasis by which it may affect the progression of many pathophysiological conditions such as gestational diabetes mellitus (GDM), nonalcoholic steatohepatitis (NASH), and cancer. Here, we review the key studies contributing to our current understanding of the roles of HKDC1 in human pathophysiological conditions and potential therapeutic interventions.

Mid-term clinical and health-related quality of life outcomes for the Trifecta bioprosthesis.

BACKGROUND: The Trifecta valve has been reported to have excellent hemodynamics. Controversy exists on occurrence of patient-prosthesis mismatch (PPM) and data on mid-term outcome is sparse. Health-related quality of life (HRQoL) assessment for the Trifecta valve has not been reported before. The aim of this study was to report the mid-term clinical and HRQoL outcomes in patients undergoing Trifecta valve implantation at our institution. METHODS: In this prospective, observational study, patients undergoing an aortic valve replacement (AVR) using the Trifecta valve were included. Data collection was retrospective from prospectively collected institutional database. Clinical and echocardiographic data were collected prospectively during follow-up. Quality of life was assessed using the Short Form-36 (SF-36) questionnaire. RESULTS: Forty-seven patients were included in the study of which 9 (19%) were women. Isolated AVR was carried out in 33 (70%) patients. In-hospital mortality and 30-day mortality were 1 (2.1%) and 2 (4.2%), respectively. With a mean indexed effective orifice area (iEOA) 0.96 ± 0.1, none of the patients had severe PPM. Moderate PPM was seen in 19%. The mean follow-up was 3 ± 1.7 years. The 5-year survival estimate was 83.2% in the overall cohort, 81.4% in the isolated and 87.5% in the concomitant procedure group. Freedom from re-operation and structural valve degeneration at 5 years was 95.7% and 97.8%. The mean physical health composite was 69.24 ± 2 and the mean mental health composite was 69.7 ± 25, indicating excellent mental and physical well-being among patients. CONCLUSION: The Trifecta valve provides satisfactory hemodynamics, survival and freedom from re-operation and excellent HRQoL at mid-term follow-up.

Metabolomic analysis of a selective ABCA1 inducer in obesogenic challenge provides a rationale for therapeutic development.

BACKGROUND: Therapeutic agents with novel mechanisms of action are needed to combat the growing epidemic of type 2 diabetes (T2D) and related metabolic syndromes. Liver X receptor (LXR) agonists possess preclinical efficacy yet produce side effects due to excessive lipogenesis. Anticipating that many beneficial and detrimental effects of LXR agonists are mediated by ABCA1 and SREPB1c expression, respectively, we hypothesized that a phenotypic optimization strategy prioritizing selective ABCA1 induction would identify an efficacious lead compound with an improved side effect profile over existing LXRß agonists. METHODS: We synthesized and characterized a novel small molecule for selective induction of ABCA1 vs. SREBP1c in vitro. This compound was evaluated in both wild-type mice and a high-fat diet (HFD) mouse model of obesity-driven diabetes through functional, biochemical, and metabolomic analysis. FINDINGS: Six weeks of oral administration of our lead compound attenuated weight gain, glucose intolerance, insulin signaling deficits, and adiposity. Global metabolomics revealed suppression of gluconeogenesis, free fatty acids, and pro-inflammatory metabolites. Target identification linked these beneficial effects to selective LXRß agonism and PPAR/RXR antagonism. INTERPRETATION: Our observations in the HFD model, combined with the absence of lipogenesis and neutropenia in WT mice, support this novel approach to therapeutic development for T2D and related conditions.

Hepatic HKDC1 Expression Contributes to Liver Metabolism.

Glucokinase (GCK) is the principal hexokinase (HK) in the liver, operating as a glucose sensor to regulate glucose metabolism and lipid homeostasis. Recently, we proposed HK domain-containing 1 (HKDC1) to be a fifth HK with expression in the liver. Here, we reveal HKDC1 to have low glucose-phosphorylating ability and demonstrate its association with the mitochondria in hepatocytes. As we have shown previously that genetic deletion of HKDC1 leads to altered hepatic triglyceride levels, we also explored the influence of overexpression of HKDC1 in hepatocytes on cellular metabolism, observing reduced glycolytic capacity and maximal mitochondrial respiration with concurrent reductions in glucose oxidation and mitochondrial membrane potential. Furthermore, we found that acute in vivo overexpression of HKDC1 in the liver induced substantial changes in mitochondrial dynamics. Altogether, these findings suggest that overexpression of HKDC1 causes mitochondrial dysfunction in hepatocytes. However, its overexpression was not enough to alter energy storage in the liver but led to mild improvement in glucose tolerance. We next investigated the conditions necessary to induce HKDC1 expression, observing HKDC1 expression to be elevated in human patients whose livers were at more advanced stages of nonalcoholic fatty liver disease (NAFLD) and similarly, found high liver expression in mice on diets causing high levels of liver inflammation and fibrosis. Overall, our data suggest that HKDC1 expression in hepatocytes results in defective mitochondrial function and altered hepatocellular metabolism and speculate that its expression in the liver may play a role in the development of NAFLD.

Hepatic hexokinase domain containing 1 (HKDC1) improves whole body glucose tolerance and insulin sensitivity in pregnant mice.

Hexokinase domain containing 1, a recently discovered putative fifth hexokinase, is hypothesized to play key roles in glucose metabolism. Specifically, during pregnancy in a recent genome wide association study (GWAS), a strong correlation between HKDC1 and 2-h plasma glucose in pregnant women from different ethnic backgrounds was shown. Our earlier work also reported diminished glucose tolerance during pregnancy in our whole body HKDC1 heterozygous mice. Therefore, we hypothesized that HKDC1 plays important roles in gestational metabolism, and designed this study to assess the role of hepatic HKDC1 in whole body glucose utilization and insulin action during pregnancy. We overexpressed human HKDC1 in mouse liver by injecting a human HKDC1 adenoviral construct; whereas, for the liver-specific HKDC1 knockout model, we used AAV-Cre constructs in our HKDC1fl/fl mice. Both groups of mice were subjected to metabolic testing before and during pregnancy on gestation day 17-18. Our results indicate that hepatic HKDC1 overexpression during pregnancy leads to improved whole-body glucose tolerance and enhanced hepatic and peripheral insulin sensitivity while hepatic HKDC1 knockout results in diminished glucose tolerance. Further, we observed reduced gluconeogenesis with hepatic HKDC1 overexpression while HKDC1 knockout led to increased gluconeogenesis. These changes were associated with significantly enhanced ketone body production in HKDC1 overexpressing mice, indicating that these mice shift their metabolic needs from glucose reliance to greater fat oxidation and ketone utilization during fasting. Taken together, our results indicate that hepatic HKDC1 contributes to whole body glucose disposal, insulin sensitivity, and aspects of nutrient balance during pregnancy.

Inhibition of mTOR complexes protects cancer cells from glutamine starvation induced cell death by restoring Akt stability.

Glutamine, a well-established oncometabolite, anaplerotically fuels mitochondrial energy metabolism and modulates activity of mammalian/mechanistic target of rapamycin complexes (mTOR). Currently, mTOR inhibitors are in clinical use for certain types of cancer but with limited success. Since glutamine is essential for growth of many cancers, we reasoned that glutamine deprivation under conditions of mTOR inhibition should be more detrimental to cancer cell survival. However, our results show that when cells are deprived of glutamine concomitant with mTOR inhibition, hepatocarcinoma cells elicit an adaptive response which aids in their survival due to enhanced autophagic flux. Moreover, inhibition of mTOR promotes Akt ubiquitination and its proteasomal degradation however we show that Akt degradation is abrogated by increased autophagy following glutamine withdrawal. Under conditions of glutamine deficiency and mTOR inhibition, the enhanced stability of Akt protein may provide survival cues to cancer cells. Thus, our data uncovers a novel molecular link between glutamine metabolism, autophagy and stability of Akt with cancer cell survival.

Studies on the Tissue Localization of HKDC1, a Putative Novel Fifth Hexokinase, in Humans.

Hexokinase domain component 1 (HKDC1) is a recently discovered novel protein, which is being promoted as a putative fifth hexokinase. Although the exact role HKDC1 plays in physiology is still unclear, it has been shown to be important during pregnancy in the regulation of glucose homeostasis. In this study, we have comprehensively studied the expression pattern of HKDC1 in the human body. Using human tissue sample, immunohistochemistry imaging was performed. Our studies indicate that the tissues with highest HKDC1 expression were the brush border epithelium of the intestines, parts of the pancreas, and lung alveolar macrophages. Future directions will be to understand the role of this fifth hexokinase in these tissues, with a focus on its relative function as compared with other endogenously expressed hexokinases.

Dual histone reader ZMYND8 inhibits cancer cell invasion by positively regulating epithelial genes.

Enhanced migratory potential and invasiveness of cancer cells contribute crucially to cancer progression. These phenotypes are achieved by precise alteration of invasion-associated genes through local epigenetic modifications which are recognized by a class of proteins termed a chromatin reader. ZMYND8 [zinc finger MYND (myeloid, Nervy and DEAF-1)-type containing 8], a key component of the transcription regulatory network, has recently been shown to be a novel reader of H3.1K36Me2/H4K16Ac marks. Through differential gene expression analysis upon silencing this chromatin reader, we identified a subset of genes involved in cell proliferation and invasion/migration regulated by ZMYND8. Detailed analysis uncovered its antiproliferative activity through BrdU incorporation, alteration in the expression of proliferation markers, and cell cycle regulating genes and cell viability assays. In addition, performing wound healing and invasion/migration assays, its anti-invasive nature is evident. Interestingly, epithelial-mesenchymal transition (EMT), a key mechanism of cellular invasion, is regulated by ZMYND8 where we identified its selective enrichment on promoters of CLDN1/CDH1 genes, rich in H3K36Me2/H4K16Ac marks, leading to their up-regulation. Thus, the presence of ZMYND8 could be implicated in maintaining the epithelial phenotype of cells. Furthermore, syngeneic mice, injected with ZMYND8-overexpressed invasive breast cancer cells, showed reduction in tumor volume and weight. In concert with this, we observed a significant down-regulation of ZMYND8 in invasive ductal and lobular breast cancer tissues compared with normal tissue. Taken together, our study elucidates a novel function of ZMYND8 in regulating EMT and invasion of cancer cells, possibly through its chromatin reader function.

Chromatin reader ZMYND8 is a key target of all trans retinoic acid-mediated inhibition of cancer cell proliferation.

All trans retinoic acid (ATRA), an active vitamin-A derivative, has been shown to regulate gene expression program and thus imparts anti-proliferative activity to cancer cells. Previously, we identified a dual histone reader ZMYND8 (zinc finger MYND (Myeloid, Nervy and DEAF-1)-type containing 8), to be a novel target of ATRA. In the present study, we attempted to decipher the detail mechanism of its transcription regulation. ATRA can reprogram the epigenetic landscape in the upstream regulatory region of ZMYND8 thereby promoting its expression. Interestingly, there is a unique H3K27Me3 to H3K27Ac switch upon ATRA-treatment. We show here that ATRA causes dynamic changes in recruitment of transcription factor YY1 in concert with HDAC1 at ZMYND8 promoter. Further, we show that ATRA treatment triggers an anti-proliferative activity in cancer cells through regulation of ZMYND8 expression. Subsequently, in 4T1-induced syngenic tumor mouse model, ATRA injection caused significant upregulation of ZMYND8. Overall our findings highlight a novel mechanism underlying ATRA-mediated changes in ZMYND8 expression which, in turn, activates the anti-proliferative program in a cancer cell. Thus, histone reader mediated modulation of epigenetic language could play a significant role in retinoid based therapeutic strategy which is well exploited to combat tumor growth.

Studies on the effect of sodium arsenate on the enzymes of carbohydrate metabolism, brush border membrane, and oxidative stress in the rat kidney.

Arsenic is an environmental pollutant and its contamination in drinking water poses serious world wide environmental health threats. It produces multiple adverse effects in various tissues, including the kidney. However, biochemical mechanism and renal response to its toxic insult are not completely elucidated. We hypothesized that sodium arsenate (ARS) induces oxidative stress and alters the structure and metabolic functions of kidney. Male Wistar rats were administered ARS (10 mg/kg body weight/day), intraperitoneally daily for 10 days. ARS administration increased blood urea nitrogen, serum creatinine, cholesterol, glucose, and phospholipids but decreased inorganic phosphate, indicating kidney toxicity. The activity of brush border membrane (BBM) enzymes significantly lowered in both cortex and medulla. Activity of hexokinase, lactate dehydrogenase, glucose-6-phosphate dehydrogenases, and NADP-malic enzyme significantly increased whereas malate dehydrogenase, glucose-6-phosphatase, and fructose 1,6 bis phosphatase decreased by ARS exposure. The activity of superoxide dismutase, GSH-peroxidase, and catalase were selectively altered in renal tissues along with an increase in lipid peroxidation. The present results indicated that ARS induced oxidative stress caused severe renal damage that resulted in altered levels of carbohydrate metabolism and BBM enzymes.

Studies on the protective effect of dietary fish oil on cisplatin induced nephrotoxicity in rats.

Cisplatin (CP) is a major antineoplastic drug for the treatment of solid tumors, however, dose dependent nephrotoxicity remains the major concern for its long term use. Several agents/strategies were attempted to prevent CP nephrotoxicity but were not found suitable for clinical practice. Dietary fish oil (FO) enriched in ω-3 fatty acids has been shown to prevent/reduce the progression of certain types of cancers, cardiovascular and renal disorders. The present study was undertaken to see whether FO can prevent CP-induced nephrotoxic and other deleterious effects. Rats were prefed experimental diets for 10days and then received a single dose of CP (6mg/kg body weight) intraperitoneally while still on diet. Serum/urine parameters, enzymes of carbohydrate metabolism, brush border membrane (BBM) and oxidative stress in rat kidney were analyzed. CP nephrotoxicity was recorded by increased serum creatinine and blood urea nitrogen. CP decreased the activities of metabolic enzymes, antioxidant defense system and BBM enzymes. In contrast, FO alone increased enzyme activities of carbohydrate metabolism and brush border membrane (BBM). FO feeding to CP treated rats markedly enhanced resistance to CP-elicited deleterious effects. Dietary FO supplementation ameliorated CP induced specific metabolic alterations and oxidative damage due to its intrinsic biochemical antioxidant properties.

Studies on the protective effect of dietary fish oil on uranyl-nitrate-induced nephrotoxicity and oxidative damage in rat kidney.

Human and animal exposure demonstrates that uranium is nephrotoxic. However, attempts to reduce it were not found suitable for clinical use. Dietary fish oil (FO) enriched in omega-3 fatty acids reduces the severity of cardiovascular and renal diseases. Present study investigates the protective effect of FO on uranyl nitrate (UN)-induced renal damage. Rats prefed with experimental diets for 15 days, given single nephrotoxic dose of UN (0.5mg/kg body weight) intraperitoneally. After 5d of UN treatment, serum/urine parameters, enzymes of carbohydrate metabolism, brush border membrane (BBM), oxidative stress and phosphate transport were analyzed in rat kidney. UN nephrotoxicity was characterized by increased serum creatinine and blood urea nitrogen. UN increased the activity of lactate dehydrogenase and NADP-malic enzyme whereas decreased malate, isocitrate and glucose-6-phophate dehydrogenases; glucose-6-phophatase, fructose-1, 6-bisphosphatase and BBM enzyme activities. UN caused oxidant/antioxidant imbalances as reflected by increased lipid peroxidation, activities of superoxide dismutase, glutathione peroxidase and decreased catalase activity. Feeding FO alone increased activities of enzymes of glucose metabolism, BBM, oxidative stress and Pi transport. UN-elicited alterations were prevented by FO feeding. However, corn oil had no such effects and was not similarly effective. In conclusion, FO appears to protect against UN-induced nephrotoxicity by improving energy metabolism and antioxidant defense mechanism.

Protective effect of green tea extract on gentamicin-induced nephrotoxicity and oxidative damage in rat kidney.

Gentamicin (GM) is an effective aminoglycoside antibiotic against severe infections but nephrotoxicity and oxidative damage limits its long term clinical use. Various strategies were attempted to ameliorate GM nephropathy but were not found suitable for clinical practice. Green tea (GT) polyphenols have shown strong chemopreventive and chemotherapeutic effects against various pathologies. We hypothesized that GT prevents GM nephrotoxicity by virtue of its antioxidative properties. A nephrotoxic dose of GM was co-administered to control and GT-fed male Wistar rats. Serum parameters and enzymes of oxidative stress, brush border membrane (BBM), and carbohydrate metabolism were analyzed. GM increased serum creatinine, cholesterol, blood urea nitrogen (BUN), lipid peroxidation (LPO) and suppressed superoxide dismutase (SOD) and catalase activities in renal tissues. Activity of hexokinase, lactate dehydrogenase increased whereas malate dehydrogenase decreased. Gluconeogenic enzymes and glucose-6-phosphate dehydrogenase were differentially altered in the cortex and medulla. However, GT given to GM rats reduced nephrotoxicity parameters, enhanced antioxidant defense and energy metabolism. The activity of BBM enzymes and transport of Pi declined by GM whereas GT enhanced BBM enzymes and Pi transport. In conclusion, green tea ameliorates GM elicited nephrotoxicity and oxidative damage by improving antioxidant defense, tissue integrity and energy metabolism.

Food as medicine: potential therapeutic tendencies of plant derived polyphenolic compounds.

The last two decades have witnessed a major drift in the interests of the scientific community towards explaining better means to containing the health risks of the human race. The century old chemotherapies against various disorders have never been a success, albeit not a total failure. Such therapies have a major drawback of side effects that give rise to unseen disorders that emerge as a new challenge. In this regard, the concept of foodstuffs as natural medicines is very attractive. Epidemiological studies suggest that the vegeteranian food habit is associated with reduced risk of cancer, cardiovascular and neurodegenerative disorders. Consistent with this hypothesis is the fact that the incidence of these disorders is least in Asian populations where fruits, vegetables and spices are the major elements in the human diet. Recent research has shown that plant-derived polyphenolic compounds are promising nutraceuticals for control of various disorders such as cardiovascular,neurological and neoplastic disease. The richness of the polyphenolic contents of green tea and red wine has made them popular choice for associated anticancer and cardiovascular health benefits. The present article is a brief review of the promises plant polyphenols, bioactive components of our food, hold for the future.