ReviewHexokinase-2 bound to mitochondria: Cancer's stygian link to the “Warburg effect” and a pivotal target for effective therapy☆
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Isoforms of hexokinase
Hexokinase catalyzes the essentially irreversible first step of the glycolytic pathway where glucose is phosphorylated to glucose-6-phosphate (G-6-P) via phosphate transfer from ATP.
The basis for this reaction is the entrapment of G-6-P inside the cell for commitment to either the glycolytic pathway, primarily for energy (ATP) generation via glycolysis and oxidative phosphorylation, or the shunting of this metabolite to the pentose–phosphate pathway to be
Discovery of the exceptional importance of HK-2 to cancer metabolism, i.e., to the “Warburg effect”
Glucose is an essential metabolite, both as a key source of cellular energy currency and a precursor carbon source for biosynthesis (anabolism) in mammalian tissues. Most normal tissues metabolize 6-carbon glucose to 3-carbon pyruvate (“glycolysis”) and then harness the energy within this molecule in the form of ATP via “oxidative phosphorylation” in mitochondria. That is, they oxidize the pyruvate to CO2 and H2O using the tricarboxylic acid cycle and the mitochondrial electron transport chain,
Key events that led to the discovery that the HK-2 metabolic step can be used for monitoring clinical cancers via PET analysis
Subsequent to the discovery in 1977 [12] that a mitochondrial-bound form of hexokinase is the key player in the “Warburg effect” in cancer, a newly developed diagnostic tracer technology capitalized on this discovery to detect cancers non-invasively in humans. Thus, in 1982 and 1983 a “deoxy” analog of glucose (2-deoxy-d-glucose) that can be phosphorylated by HK-2 but not metabolized further, and that had been labeled with the positron emitter 18F (18FDG), was used successfully to image cancers
The metabolic rationale for the propensity of tumors to selectively express HK-2
There are three likely reasons: (1) based on the binding affinities described in the “Background”, it is obvious that the selection of hexokinases over glucokinase (HK-4) will be quite favorable from a metabolic standpoint, as isoforms of the former can harness glucose with over 100-fold higher affinity than the latter enzyme; (2) the selection HK-2 rather than HK-3 and HK-4 is likely due to the fact that HK-2 in contrast to these isoforms has a N-terminal hydrophobic domain that allows it to
Discoveries that revealed that the tumor HK-2 gene promoter is highly “promiscuous” in facilitating transcriptional up-regulation under both adverse and favorable metabolic states of the host
Several systemic and cellular stimuli promote the specific expression and transcriptional up-regulation of HK-2 (and to a lesser extent HK-1) in highly glycolytic malignant tumors. The first indication for enhanced transcription came via northern-blot based mRNA expression studies [21], [22], [23], [24]. These revealed an approximately 100-fold increase in the mRNA levels for HK-2, strongly suggesting activation and up-regulation of HK-2 gene transcription. Based on these initial findings it
Epigenetic and genetic factors involved in the marked over-expression of HK-2 in tumors—findings that revealed that in liver cancer cells exhibiting a pronounced Warburg effect the HK-2 gene is subject to both epigenetic regulation and regulation by amplification
Sequence analysis and comparison of the HK-2 promoters from normal tissue (hepatocytes) and a malignant tumor (AS-30D hepatoma) that exhibits a robust Warburg effect failed to identify any significant nucleotide differences. Less than 1% of the nucleotide positions were altered and were not in critical cis-element harboring regions [30]. In addition, based on available data (www.ncbi.nlm.nih.gov), it is known that each of the four hexokinase isozymes is encoded by different chromosomal loci. In
HK-2 as a possible metabolic and bio-energetic flux regulator of normal tissues that is dysfunctional in tumors
With its intimate metabolic coupling to mitochondrial ATP output (and the ADP input into mitochondria via “porins”, e.g., VDACs), HK can be considered a metabolic regulator that closely balances fluxes between glycolysis and mitochondrial respiration as discussed in a recent review by Wilson [3]. In essence, in normal tissues where HK-2 is either silent or expressed at low levels, the phosphorylation rate of incoming glucose via HK-1 can be coordinated with the rate of mitochondrial oxidative
Discovery that HK-2 in addition to its growth related roles in cancer also helps immortalize cancer cells
As noted above, HK-2 is localized predominantly on the outer mitochondrial membrane where it is bound to one or more VDAC proteins. Various metabolic and signal-transduction related stimuli have been implicated in regulating hexokinase-VDAC binding, including intracellular lactate, pH, ATP/ADP, glucose/glucose-6-phosphate metabolite couples, and protein kinase-B (PKB/Akt) [35], [36], [37], among others. In addition to being critical for the unique metabolism of many cancers,
Targeting tumors for destruction by silencing the HK-2 transcript or via small-molecule-mediated inhibition of the enzyme
Based on the factors discussed above it is quite evident that “knock-down” or silencing of HK-2 expression should have a deleterious effect on tumor proliferation. This was evaluated first via anti-sense RNA approaches against HK-2, where anti-sense messages against HK-2 were expressed via retroviral-mediated transduction in malignant hepatoma cells (Mathupala and Pedersen, Proc Am Assoc Can Res 1999:22 (abstract # 145)). In this study, although a dramatic reduction in tumor proliferation was
Releasing HK-2 from the VDAC anchor to disrupt tumor glycolysis
Based on current inferences on the HK-2/VDAC interaction in preventing tumor apoptosis, disruption of the same should facilitate tumor apoptosis. This in fact has been tested with several compounds that reportedly disrupt the VDAC-HK-2 interaction. Among the compounds tested are the antifungal compounds clotrimazole and bifanazole [52], methyl jasmonate [53], and peptide sequences that correspond to the HK-2 N-terminal [45], [46]. In each case, induction of apoptosis was observed in the
The penultimate step in glycolysis—mitochondrial pyruvate metabolism and the Warburg effect
In contrast to the above approaches, others have examined the feasibility of targeting alternate steps of the glycolytic pathway as a mode of disrupting energy metabolism in malignant tumors [34], [54], [55], [56] (Fig. 2). These have included (1) inhibition of lactic acid efflux from tumors by silencing or inhibiting lactate transporters via interfering RNA or cinnamic acid derivatives (ACCA) [34], [55], [56], (2) up-regulation of the influx of pyruvate into mitochondria by inhibiting pyruvate
Concluding remarks and prospects for the future
The work of a handful of dedicated, if not stubborn, tumor metabolism research groups over the past seven decades have systematically unraveled the biochemical choreography that exists between signal transduction cascades and metabolic pathways in tumors to promote malignancy (i.e., proliferation). A first benefit to cancer patients has been the utilization of the high glucose influx of malignant tumors via mitochondrial-bound hexokinase (HK-2 and to some extent HK-1) as a tool to develop
Conflict of interest
None.
References (60)
- et al.
Functional organization of mammalian hexokinases: characterization of chimeric hexokinases constructed from the N- and C-terminal domains of the rat type I and type II isozymes
Arch Biochem Biophys
(1995) - et al.
Functional organization of mammalian hexokinases: both N- and C-terminal halves of the rat type II isozyme possess catalytic sites
Arch Biochem Biophys
(1996) - et al.
Functional organization of mammalian hexokinases: characterization of the rat type III isozyme and its chimeric forms, constructed with the N- and C-terminal halves of the type I and type II isozymes
Arch Biochem Biophys
(1997) - et al.
Functional organization of mammalian hexokinase II. Retention of catalytic and regulatory functions in both the NH2- and COOH-terminal halves
J Biol Chem
(1996) - et al.
Mitochondrial bound type II hexokinase: a key player in the growth and survival of many cancers and an ideal prospect for therapeutic intervention
Biochim Biophys Acta
(2002) - et al.
Energy metabolism of tumor cells. Requirement for a form of hexokinase with a propensity for mitochondrial binding
J Biol Chem
(1981) - et al.
Functional significance of mitochondrial bound hexokinase in tumor cell metabolism. Evidence for preferential phosphorylation of glucose by intramitochondrially generated ATP
J Biol Chem
(1988) - et al.
Differences in expression and intracellular distribution of hexokinase isoenzymes in rat liver cells of different transformation stages
Biochim Biophys Acta
(1994) - et al.
Glucose catabolism in cancer cells. Isolation, sequence, and activity of the promoter for type II hexokinase
J Biol Chem
(1995) - et al.
Evidence that transcription of the hexokinase gene is increased in a rapidly growing rat hepatoma
Biochem Biophys Res Commun
(1985)
Glucose catabolism in cancer cells. The type II hexokinase promoter contains functionally active response elements for the tumor suppressor p53
J Biol Chem
Glucose catabolism in cancer cells: identification and characterization of a marked activation response of the type II hexokinase gene to hypoxic conditions
J Biol Chem
Glucose catabolism in cancer cells: regulation of the Type II hexokinase promoter by glucose and cyclic AMP
FEBS Lett
Glucose metabolism in cancer: importance of transcription factor-DNA interactions within a short segment of the proximal region of the type II hexokinase promoter
J Biol Chem
Glucose metabolism in cancer. Evidence that demethylation events play a role in activating type II hexokinase gene expression
J Biol Chem
Mitochondrial hexokinase from differentiated and undifferentiated HT29 colon cancer cells: effect of some metabolites on the bound/soluble equilibrium
Int J Biochem
Distinct domains of Bcl-XL are involved in Bax and Bad antagonism and in apoptosis inhibition
Exp Cell Res
Hexokinase-mitochondria interaction mediated by Akt is required to inhibit apoptosis in the presence or absence of Bax and Bak
Mol Cell
Glucose catabolism in the rabbit VX2 tumor model for liver cancer: characterization and targeting hexokinase
Cancer Lett
Advanced cancers: eradication in all cases using 3-bromopyruvate therapy to deplete ATP
Biochem Biophys Res Commun
Clotrimazole and bifonazole detach hexokinase from mitochondria of melanoma cells
Eur J Pharmacol
A mitochondria-K+ channel axis is suppressed in cancer and its normalization promotes apoptosis and inhibits cancer growth
Cancer cell
Attenuation of LDH-A expression uncovers a link between glycolysis, mitochondrial physiology, and tumor maintenance
Cancer Cell
Hexokinases
Rev Physiol Biochem Pharmacol
An introduction to the isoenzymes of mammalian hexokinase types I-III
Biochem Soc Trans
Isozymes of mammalian hexokinase: structure, subcellular localization and metabolic function
J Exp Biol
Tumor mitochondria and the bioenergetics of cancer cells
Prog Exp Tumor Res
Warburg, me and hexokinase 2: multiple discoveries of key molecular events underlying one of cancers’ most common phenotypes, the “Warburg effect”, i.e., elevated glycolysis in the presence of oxygen
J Bioenerg Biomembr
High aerobic glycolysis of rat hepatoma cells in culture: role of mitochondrial hexokinase
Proc Natl Acad Sci USA
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S.P.M. is supported by NIH grant R01CA116257, and P.L.P. by NIH grants R01CA08018 and R01CA010951. Also, Y.H.K. and P.L.P. were supported in part by Grant BCTR0402523 from the Susan Komen Breast Cancer Foundation while some of the work discussed herein was conducted.