Hexokinase is an enzyme found in nearly all living cells that converts glucose into energy. It performs the first step in glucose phosphorylation, creating glucose-6-phosphate. This process is important for cellular energy production and is regulated to maintain cellular energy balance. Hexokinase inhibitors are compounds designed to block this enzyme’s activity.
Hexokinase and Its Variants
Hexokinase initiates glucose metabolism by phosphorylating glucose to glucose-6-phosphate, trapping glucose inside the cell and directing it into various metabolic pathways. This phosphorylation is the first committed step in glucose utilization. Glucose-6-phosphate can then be used for energy production through glycolysis, converted to glucose-1-phosphate for glycogen synthesis, or enter the pentose phosphate pathway.
Mammalian cells contain four main isoforms of hexokinase: HK1, HK2, HK3, and HK4, also known as glucokinase (GCK). These isoforms have different kinetic properties, expression patterns across tissues, and subcellular locations.
HK1 is widely distributed in most tissues, including the brain and red blood cells, where it regulates glucose metabolism. HK1 is activated by high inorganic phosphate levels and inhibited by its product, glucose-6-phosphate.
HK2 is found predominantly in insulin-sensitive tissues like muscle, heart, and adipose tissues, and is also often found in tumor cells. Like HK1, HK2 can localize to mitochondria.
HK3 is a less understood isoform, primarily expressed in hematopoietic cells and tissues, including granulocytes, lung, kidney, and liver, though at lower levels compared to HK1 and HK2. Unlike HK1 and HK2, HK3 lacks the N-terminal mitochondrial binding domain.
HK4, or glucokinase, is mainly expressed in the liver and pancreas, and also in enteroendocrine cells and the brain. Glucokinase has a lower affinity for glucose compared to HK1-3, meaning it only becomes highly active when glucose levels are elevated, acting as a glucose sensor. Unlike HK1, HK2, and HK3, glucokinase is not inhibited by glucose-6-phosphate, but its activity is influenced by insulin and glucagon.
How Hexokinase Inhibitors Work
Hexokinase inhibitors interfere with the enzyme’s ability to phosphorylate glucose. One method is competitive inhibition, where the inhibitor structurally resembles glucose and binds to the enzyme’s active site. This prevents glucose from binding and undergoing phosphorylation. For example, glucose-6-phosphate, the product of the hexokinase reaction, can competitively inhibit hexokinase by binding to the ATP site.
Another mechanism is allosteric inhibition, where the inhibitor binds to a site on the enzyme different from the active site. This binding causes a change in the enzyme’s shape, which then reduces the active site’s ability to bind glucose or catalyze the reaction. For instance, glucose-6-phosphate can also allosterically inhibit hexokinase by binding to its N-terminal half, indirectly affecting the active site located in the C-terminal half. Some inhibitors are designed to be isoform-specific, targeting certain hexokinase variants more effectively due to their unique structural or regulatory features.
Therapeutic Applications of Hexokinase Inhibitors
Hexokinase inhibitors show promise in therapeutic applications, particularly in cancer treatment. Cancer cells often exhibit the “Warburg effect,” relying heavily on glycolysis for energy even when oxygen is available. This metabolic shift helps cancer cells meet increased energy demands and provides building blocks for rapid proliferation. Many tumors overexpress specific hexokinase isoforms, especially HK2, which is often bound to the outer mitochondrial membrane.
Inhibiting hexokinase in cancer cells can disrupt their energy supply and metabolic processes, potentially leading to cell death or reduced growth. By blocking the first step of glucose metabolism, these inhibitors starve cancer cells of the glucose-6-phosphate needed for glycolysis and other biosynthetic pathways. For example, 2-deoxy-D-glucose (2-DG) is a glucose analog phosphorylated by hexokinase but cannot be further metabolized, effectively trapping glucose and blocking glycolysis.
Lonidamine is another hexokinase inhibitor investigated for its anti-cancer effects. 3-bromopyruvate (3-BP) is an alkylating agent that inhibits hexokinase and affects mitochondria, leading to rapid ATP depletion and tumor destruction in animal models. 3-BP has been shown to sensitize leukemic cells to anti-leukemic drugs by dissociating HK2 from the mitochondrial complex. Research continues to explore these and other hexokinase inhibitors as potential treatments, sometimes in combination with other therapies, to enhance effectiveness and overcome drug resistance in various cancers.
Beyond cancer, hexokinase inhibition is explored for other conditions. Increased HK2 levels have been observed in microglia from Alzheimer’s disease (AD) mouse models and patients. Inhibiting HK2 in microglia promotes the clearance of β-amyloid, a hallmark of AD, by increasing ATP generation through lipid metabolism. This suggests a potential therapeutic strategy for neurodegenerative diseases by modulating hexokinase activity.