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Oncogene (2011) 30, 4687–4696
& 2011 Macmillan Publishers Limited All rights reserved 0950-9232/11
The autophagy conundrum in cancer: influence of tumorigenic
metabolic reprogramming
CH Eng and RT Abraham
Pfizer Oncology Research Unit, Pearl River, NY, USA
Tumorigenesis is often accompanied by metabolic changes
that favor rapid energy production and increased biosynthetic capabilities. These metabolic adaptations promote the
survival and proliferation of tumor cells, and in conjunction
with the hypoxic and metabolically challenged tumor
microenvironment, influence autophagic activity. Autophagy is a catabolic process that allows cellular macromolecules to be broken down and re-utilized as metabolic
precursors. Stimulation of autophagy promotes the survival
of tumor cells under stressful metabolic and environmental
conditions, and counters the potentially deleterious effects
of mitochondrial dysfunction and the ROS that these
organelles generate. However, inhibition of autophagy has
also been reported to fuel tumorigenesis. In spite of the
advances in our understanding of the relationship between
autophagy and tumorigenesis, it remains unclear whether
the therapeutic approaches targeting autophagy should aim
to increase or decrease autophagic flux in tumor tissues in
human patients. Here, we review how metabolic reprogramming influences autophagic activity in tumors, and discuss
how inhibition of autophagy might be exploited to target
tumor cells that show altered metabolism.
Oncogene (2011) 30, 4687–4696; doi:10.1038/onc.2011.220;
published online 13 June 2011
Keywords: autophagy; cancer metabolism; glutaminolysis;
Autophagy, a term based on the Greek words for ‘self’
(auto) and ‘eating’ (phagy), describes a lysosomemediated degradative process where a cell recycles
intracellular macromolecules and organelles to regenerate metabolic building blocks. In unicellular organisms
such as yeast, autophagy is a mechanism that promotes
survival when extracellular nutrients are limiting.
Autophagy also supports cellular homeostasis in multicellular organisms during fasting. However, in metazoans, autophagy contributes to a diversity of other
Correspondence: Dr CH Eng, Pfizer Oncology Research Unit, 401 N.
Middletown Road, Building 200, Room 4603, Pearl River, NY 10965,
E-mail: [email protected]
Received 26 February 2011; revised and accepted 2 May 2011; published
online 13 June 2011
processes, such as developmental cell death (Berry and
Baehrecke, 2007; Denton et al., 2009), clearance of
harmful protein aggregates (Renna et al., 2010) and
mitochondrial clearance during erythrocyte maturation
(Kundu et al., 2008; Sandoval et al., 2008; Novak et al.,
2010). By targeting damaged macromolecules and
organelles for degradation, autophagy also protects
cells from both internal and external insults, including
metabolic, oxidative and chemotherapeutic stress.
These functions of autophagy are protective and have
a prominent role in numerous pathological conditions,
including neurodegeneration (Wong and Cuervo, 2010),
Crohn’s disease (Brest et al., 2010) and pathogenic
infections (Deretic, 2010). Its role in cancer, however, is
less defined. The first genetic link between autophagy
and cancer occurred with the discovery that Beclin-1, a
tumor suppressor, was an autophagy-related protein.
Beclin1 resides in a chromosomal region frequently
deleted in sporadic human breast, ovarian and prostate
cancers (Aita et al., 1999). In breast cancer cell lines with
reduced Beclin-1 levels, re-expression of the protein
restored autophagy and suppressed tumorigenesis
(Liang et al., 1999), whereas mice heterozygous for
beclin1 were more prone to tumorigenesis (Qu et al.,
2003). A core complex consisting of Beclin-1, Vps34
(a class-III phosphatidylinositol-3-kinase (PI3K)) and
Vps15 promotes autophagy by mediating autophagosome nucleation (Funderburk et al., 2010).
Since the discovery of Beclin-1, other studies have also
shown that autophagy-deficient cells are more tumorigenic than their autophagy-competent counterparts
(Liang et al., 1999; Marino et al., 2007; Mathew et al.,
2009), in part owing to an increase in the DNA damage
response and chromosomal instability (Karantza-Wadsworth et al., 2007; Mathew et al., 2007). In addition,
mice harboring deletions in autophagy-related (Atg)
genes are more prone to tumorigenesis (Marino et al.,
2007; Takamura et al., 2011). These data lead to the
hypothesis that autophagy has tumor-suppressive functions. However, accumulating evidence also suggests that
autophagy promotes rather than suppresses malignant
progression. Inhibition of autophagy in pancreatic
tumor cells grown as xenografts in nude mice prevents
tumor growth (Yang et al., 2011). In more advanced
tumors, autophagy protects cancer cells against chemotherapeutic or environmentally imposed metabolic
stress, which is consistent with observations that cancer
cells residing in poorly perfused tumor tissues often show
markers of increased autophagic activity (Degenhardt
Influence of metabolic reprogramming on tumor autophagic activity
CH Eng and RT Abraham
et al., 2006). Thus, autophagy might be suppressed
during the initial stages of transformation, and subsequently upregulated after the tumor is fully established
(Chen and Debnath, 2010; Galluzzi et al., 2010; Mathew
and White, 2011) (Amaravadi et al., 2011).
Induction of autophagy by chemotherapeutics may
also facilitate cellular survival, and this hypothesis is
now being examined in human clinical trials. Hydroxychloroquine, a well-established inhibitor of lysosomal
function and therefore autophagy, is being tested in
patients with established tumors in combination with
conventional cytotoxic chemotherapy (reviewed by
Livesey et al., 2009). Upregulation of autophagy
prolongs the survival of tumor cells treated with
cytotoxic drugs; thus, concurrent inhibition of autophagy in patients receiving these drugs may augment
tumor cell death (Livesey et al., 2009; Solomon and Lee,
2009; Amaravadi et al., 2011). However, certain
cytotoxic drugs have been identified that induce cell
death as a result of enhanced autophagic activity
(Turcotte et al., 2008; Salazar et al., 2009). Thus, other
groups continue to promote the idea that overstimulation of autophagy leads to cell death resulting from
excessive self-cannibalization. Although autophagy as a
cell death mechanism can occur in vitro with experimental manipulation, there is limited available evidence
that physiological autophagic cell death occurs in
mammalian cells in vivo (Kroemer and Levine, 2008).
Confirmation of autophagic cell death is usually shown
by increased survival upon knockdown of Atg proteins,
but as these proteins may have roles in other cell death
mechanisms (Yousefi et al., 2006), the notion of cell
death resulting from autophagy remains controversial
(Kroemer and Levine, 2008; Marino et al., 2011).
Efforts to understand the roles of autophagy in
tumorigenesis will require understanding how the events
that drive tumorigenesis affect autophagy. One such
notable tumorigenic event is the reprogramming of
cellular metabolism. In order to sustain continuous cell
division, tumor cells persistently engage in protein,
nucleic acid and lipid biosynthesis. Although mutations
in specific metabolic enzymes are rare, tumor cells alter
their metabolic pathways to upregulate glycolytic,
glutaminolytic and lipogenic programs.
Physiological control of autophagy: mTOR
Before examining the effect of tumorigenic metabolic
perturbations on autophagy, we must first understand
how autophagy is regulated in normal cells. Autophagy
is initiated when a double-membrane structure termed
the autophagosome engulfs cytoplasmic constituents
and fuses with the lysosome. The resulting autolysosome
represents the end of the line for the autophagosomal
cargo; in the acidic microenvironment, lysosomal
enzymes digest the macromolecules and the resulting
degradation products are exported from the lysosomal
lumen into the cytoplasm, where they are recycled back
into metabolic pathways. Initially thought to be a nonselective process, recent studies show that recognition
and autophagic clearance of certain cargo, such as
organelles, and ubiquitinated or aggregated proteins, are
mediated by specific targeting proteins (Narendra et al.,
2008; Kirkin et al., 2009; Filimonenko et al., 2010).
Autophagy is activated by limitations in glucose,
amino acids and oxygen. Early studies in budding yeast
linked amino-acid starvation-induced autophagy to
inhibition of the yeast orthologs of the mammalian
target of rapamycin (mTOR), a central component of
nutrient responsiveness in all eukaryotic cells. The mTOR
complex-1 (mTORC1), a multi-protein complex defined
by the signatory subunit Raptor, integrates signals
relating to the availability of growth factors, amino
acids, glucose, ATP and oxygen in the host cell.
Inhibition of mTORC1 by lack of these factors, or by
pharmacological agents such as rapamycin, suppresses
mRNA translation (a major consumer of metabolic
precursors and energy) and upregulates catabolism
through stimulation of autophagic flux. The activity of
mTORC1 is stimulated by the small GTPase Rheb,
which itself is inhibited by the heterodimeric tuberous
sclerosis complex (TSC) composed of TSC1 and TSC2.
Although mTOR also resides in the mTOR complex-2
(mTORC2), defined by the Rictor subunit, the relationship between mTORC2 activity and autophagy appears
to be more cell type- and context-dependent (Mammucari
et al., 2007; Thoreen et al., 2009; Gurusamy et al., 2010).
The recognition of nutrient availability by mTOR is
modulated through multiple upstream regulators (Abraham and Eng, 2008). Glucose deprivation results in an
elevated AMP/ATP ratio, leading to the activation of
AMP-activated protein kinase (AMPK), which phosphorylates and activates TSC2, causing mTOR inactivation. Similarly, hypoxia induces the expression of
REDD1 (for regulated in development and DNA
damage responses), which stimulates TSC activity and,
in turn, suppression of mTORC1. Finally, the presence
of amino acids allows the activation of mTOR by Rheb,
in part through the Rag GTPase-mediated translocation
of mTOR to the lysosomal surface (Sancak et al., 2010).
Amino-acid starvation, which inhibits mTOR, also
facilitates autophagosome–lysosome fusion through
lysosome clustering (Korolchuk et al., 2011).
Inhibition of mTOR stimulates autophagy through an
evolutionarily conserved pathway involving the protein
kinase ULK1, a homolog of the yeast Atg1 kinase.
ULK1 is found in complex with the Atg proteins FIP200
(which is thought to be the functional homolog of yeast
Atg17) and Atg13. In response to nutrients, mTOR
binds to the ULK1–Atg13–FIP200 complex and phosphorylates both ULK1 and Atg13. The binding of
mTOR to the complex and phosphorylation of ULK1
by mTOR inhibits the kinase activity of ULK1
(Hosokawa et al., 2009; Jung et al., 2009). Nutrient
limitation or treatment with rapamycin results in the
inhibition of mTOR, dissociation of mTOR from the
complex, and a reduction in ULK1 and Atg13
phosphorylation. Recent studies identified AMPK as a
positive regulator of the ULK1 complex activity, which
is necessary for autophagy (Lee et al., 2010; Egan et al.,
2011; Kim et al., 2011a).
Influence of metabolic reprogramming on tumor autophagic activity
CH Eng and RT Abraham
Autophagy is also stimulated through mTOR-independent pathways, although the exact mechanisms are
less well-understood. Numerous pharmacological agents
have been reported to stimulate autophagy in an
mTOR-independent manner (Sarkar et al., 2007; Zhang
et al., 2007), with some involvement of Ca2 þ and inositol
1,4,5-trisphosphate signaling (Williams et al., 2008).
Amino-acid regulation of autophagy also occurs in an
mTOR-independent manner in muscle cells (Mordier
et al., 2000) through FoxO3-mediated transcription of
the autophagy genes MAP1LC3B (microtubule-associated protein 1 light chain 3 b) and BNIP3 (BCL2/
adenovirus E1B 19 kDa protein-interacting protein 3)
(Mammucari et al., 2007), and in liver cells (Kanazawa
et al., 2004). A small interfering RNA screen for
autophagy modulators also identified a number of
growth factors and cytokines that suppress autophagy
independently of mTOR through inhibition of class-III
PI3K (Lipinski et al., 2010).
Effects of metabolic reprogramming on autophagy:
The first connection between tumorigenesis and metabolic reprogramming was made in the 1920s, when the
legendary German biochemist Otto Warburg first noted
that tumor slices showed an unusually high level of
glycolytic activity, even under normoxic conditions, in
contrast to normal differentiated cells, which primarily
metabolize glucose through oxidative phosphorylation.
This metabolic aberration is the basis of a clinically
validated tumor-imaging modality termed 18F-deoxyglucose positron emission tomography (FDG–PET).
Tumors are visualized by FDG–PET based on their
elevated rates of glucose uptake compared with most
normal tissues within the body (Vander Heiden et al.,
Warburg hypothesized that this alteration in glucose
metabolism resulted from mitochondrial dysfunction. It
is now appreciated that cancer cell mitochondria are not
globally dysfunctional, but rather reprogrammed to
support their increased biosynthetic needs. As mitochondria in tumor cells are re-purposed to produce the
precursors necessary for protein, lipid and nucleic acid
biosynthesis, the activity of the ATP-generating electron
transport chain is reduced and glycolytic metabolism is
increased to meet the bioenergetic and biosynthetic
needs of cancer cells. Although less efficient, the
increased pace of energy production by aerobic glycolysis compared with oxidative phosphorylation is also
beneficial to rapidly proliferating cells (DeBerardinis
et al., 2008).
The switch from oxidative phosphorylation to glycolysis is induced by the activation of several oncogenes.
Transformation induced by Ras or Src increases glucose
uptake (Flier et al., 1987). Overexpression of myc, which
occurs in over 40% of human cancers, activates the
transcription of numerous enzymes involved in glucose
metabolism, including lactate dehydrogenase, hexokinase, glucose transporter-1 (GLUT-1) and pyruvate
kinase M2 (Dang et al., 2009). Finally, activation of the
PI3K pathway, which frequently occurs in malignant
cells through deregulated growth factor receptor signaling, mutations in the p110a catalytic subunit, loss of
phosphatase and tensin homolog (PTEN) or amplification of Akt, also results in increased glucose uptake and
glycolysis (Elstrom et al., 2004).
In order to maximize cell mass accumulation during
tumorigenesis, one might expect catabolic processes
such as autophagy to be suppressed. This hypothesis is
supported by data showing that transformation driven
by oncogenes such as Ras (Furuta et al., 2004), PI3K
(Petiot et al., 2000) and myc (Balakumaran et al., 2009),
or loss of the tumor suppressor PTEN (Arico et al.,
2001), suppresses autophagy. Activation of mTOR by
these transforming events (Shaw and Cantley, 2006;
Ravitz et al., 2007) is one mechanism by which
autophagy might be downregulated in response to
oncogenic signaling. Cytokine receptor engagement also
triggers signaling events that suppress autophagy
(Lipinski et al., 2010). These studies support the
hypothesis that certain early drivers of transformation
suppress autophagy, perhaps to accommodate the need
for a rapid increase in cell mass to fuel cell proliferation.
One notable exception to oncogene-induced suppression of autophagy is mutation of K-Ras. Expression of
mutant K-Ras enhances aerobic glycolysis (Racker
et al., 1985; Vizan et al., 2005), owing, in part, to
increased expression of GLUT-1 (Yun et al., 2009)
and PFKFB3 (6-phosphofructo-2-kinase/fructose-2,6bisphosphatase) (Telang et al., 2006). The dependence
of K-Ras-mutated cells on glucose partially results from
the need for increased carbon flux through the pentose
phosphate pathway (Weinberg et al., 2010). Although
previous work showed suppression of autophagy with
expression of mutant K-Ras (Furuta et al., 2004), a
number of recent studies have illustrated the ability of
mutant K-Ras to promote basal autophagy (Guo et al.,
2011; Kim et al., 2011b; Lock et al., 2011). In these
studies, suppression of autophagy limited the ability of
mutant K-Ras to promote anchorage-independent
growth in vitro and tumorigenesis in vivo. Autophagy
may sustain transformation induced by mutant K-Ras
through the maintenance of bioenergetics. Cells harboring K-Ras mutations that were autophagy-defective had
decreased glycolytic capacity (Lock et al., 2011) and
abnormal mitochondria owing to suppressed mitophagy, resulting in reduced tricarboxylic acid (TCA)
intermediates and reduced oxygen consumption (Guo
et al., 2011). Thus, not only is autophagy stimulated
when glycolysis is upregulated owing to mutations in
K-Ras, but autophagy may also be necessary to
maintain metabolic homeostasis after a shift to glycolysis. This notion is also supported by studies showing
that enhancements to the glycolytic pathway can
stimulate autophagy (Figure 1), as discussed below.
Loss of the tumor suppressor p53 promotes glycolysis
through enhanced glucose transport (Kawauchi et al.,
2008), enhanced phosphoglycerate mutase expression
(Kondoh et al., 2005) and suppression of the mitochondrial enzyme SCO2 (synthesis of cytochrome c oxidase 2)
Influence of metabolic reprogramming on tumor autophagic activity
CH Eng and RT Abraham
(2) Glyceraldehyde-3-phosphate
(2) 1,3-bisphosphoglycerate
Figure 1 Regulation of autophagy by glycolysis. Glycolysis
positively regulates autophagy through various mechanisms.
Expression of the glycolytic enzyme GAPDH promotes the
transcription of Atg12, which stimulates autophagy. GAPDH, by
binding to Rheb, also prevents the activation of mTOR.
Conversely, inhibition of glycolysis by TIGAR re-routes glycolytic
intermediates toward the pentose phosphate pathway, resulting in
the suppression of autophagy by glutathione-dependent reduction
of ROS. Atg, autophagy-related genes; GAPDH, glyceraldehyde-3phosphate dehydrogenase; mTOR, mammalian target of rapamycin; ROS, reactive oxygen species; TIGAR, TP53-induced glycolysis and apoptosis regulator.
(Matoba et al., 2006). Another target of p53, the TP53induced glycolysis and apoptosis regulator (TIGAR),
a fructose 1,6-bisphosphatase, directly inhibits both
glycolysis and autophagy. TIGAR inhibits glycolysis by
reducing the levels of fructose 1,6-bisphosphate and
redirecting glucose metabolism toward the pentose
phosphate pathway (Bensaad et al., 2006). The pentose
phosphate pathway produces NADPH, which is required for the regeneration of reduced glutathione;
thus expression of TIGAR enhances glutathione levels
and increases the antioxidant capacity of the cell
(Figure 1). Production of glutathione decreases the
levels of reactive oxygen species (ROS), resulting in the
suppression of autophagy (Bensaad et al., 2009). Thus,
loss of the p53 tumor suppressor would decrease
TIGAR levels, resulting in the upregulation of both
glycolysis and autophagy.
Another key glycolytic enzyme is glyceraldehyde-3phosphate dehydrogenase (GAPDH), which catalyzes
the conversion of glyceraldehyde-3-phosphate to 1,3bisphosphoglycerate (Figure 1). Expression of GAPDH
stimulates the transcription of Atg12, and both increased glycolysis and Atg12 expression by GAPDH are
required to protect cells from caspase-independent cell
death (Colell et al., 2007). GAPDH expression induces a
transient decrease in mitochondrial mass and maintenance of ATP levels after mitochondrial insults,
suggesting that autophagy may protect cells by eliminating damaged mitochondria, and enhanced glycolysis
provides enough ATP necessary for mitochondrial
regeneration and cell survival. Interestingly, GAPDH
may also promote autophagy by negatively regulating
mTOR activity. GAPDH competes with mTOR for
binding to the mTOR activator Rheb, and under lowglucose conditions, expression of GAPDH, through
binding to Rheb, prevents the activation of mTOR (Lee
et al., 2009), which would promote autophagy.
Enhanced glycolysis directly correlates with stimulation of autophagy (Figure 1). This correlation occurs not
only in tumor cells, but also in activated T cells, another
highly proliferative cell type. Similar to tumor cells,
activation of T cells results in a metabolic switch from
oxidative phosphorylation to aerobic glycolysis (Fox
et al., 2005). Autophagy is required for T-cell activation
and functions to meet their increased bioenergetic needs
(Hubbard et al., 2010). Thus, induction of autophagy
may be necessary to maintain cellular bioenergetics after
a shift to glycolytic metabolism, not only in tumor cells
but in proliferative normal cells as well.
Effects of metabolic reprogramming on autophagy:
hypoxia and ROS
Although upregulation of aerobic glycolysis supports the
increased biosynthetic demands of tumor cells under
normoxic conditions, glycolysis is also enhanced if access
to oxygen is limited owing to inadequate tissue perfusion.
Hypoxic tumor tissues show markers of increased
autophagic activity (Rouschop et al., 2010). Hypoxia
also generates ROS (Semenza, 2010), which directly
induce autophagy (Djavaheri-Mergny et al., 2006; Chen
et al., 2009; Huang et al., 2009). Generation of ROS also
occurs with nutrient starvation (Scherz-Shouval et al.,
2007) and mitochondrial electron transport.
The master transcriptional regulators of hypoxic
responses are the hypoxia-inducible factors (HIFs).
Active HIF is a heterodimer comprising HIF-1a or
HIF-2a partnered with the constitutively expressed
HIF-1b subunit. The expression of the HIF-1/2a
subunits is tightly controlled through oxygen-dependent
hydroxylation by prolyl hydroxylases. In the presence of
oxygen, HIF-1/2a is targeted for degradation. Under
low-oxygen conditions, or in the presence of ROS, HIF1/2a becomes stabilized, partners with HIF-1b and
enters the nucleus, where the HIF complexes orchestrate
hypoxia-induced gene expression (Semenza, 2010).
Stabilization of HIF-1a promotes mitochondrial
autophagy (mitophagy) through transcription of the
Bcl-2 homology-domain-3-containing proteins BNIP3
and BNIP3L. BNIP3 stimulates autophagy by preventing the inhibitory binding of Bcl-2 to Beclin-1, which
frees Beclin-1 to participate in autophagosome formation (Figure 2). Mitophagy induced by BNIP3 results in
the clearance of damaged mitochondria, which are
major sources of cell-damaging ROS (Zhang et al.,
2008). Inhibition of autophagy sensitizes cells to hypoxic
cell death (Bellot et al., 2009; Rouschop et al., 2009) and
ROS-induced cell death (Huang et al., 2009). Collectively, these results suggest that ROS-induced autophagic activity limits cellular damage by removing damaged
macromolecules and dysfunctional mitochondria.
Influence of metabolic reprogramming on tumor autophagic activity
CH Eng and RT Abraham
Figure 2 Stimulation of autophagy by ROS and hypoxia. ROS,
which are produced during nutrient starvation or hypoxia,
stimulate autophagy through multiple signaling pathways. Oxidation and inactivation of Atg4A by ROS allows the buildup of LC3
conjugated to phosphatidylethanolamine and subsequent promotion of autophagy. ROS also inhibit mTOR through activation of
TSC2 by ATM. Stimulation of autophagy by hypoxia occurs
through suppression of mTOR through HIF-1a-dependent activation of REDD1. HIF-1a also induces the transcription of BNIP3/
BNIP3L, which prevents the binding of Bcl-2 to Beclin, allowing
Beclin to stimulate autophagy. Finally, hypoxia promotes autophagy through an HIF-1-independent pathway involving transcriptional induction of Atg5 and LC3 through the PERK-responsive
transcription factors ATF4 and CHOP. The activation of
autophagy by hypoxia and ROS results in a quality-control
feedback loop by elimination of damaged mitochondria through
mitophagy, thereby reducing ROS. Atg, autophagy-related genes;
HIF, hypoxia-inducible factor; mTOR, mammalian target of
rapamycin; ROS, reactive oxygen species; TSC2, tuberous sclerosis
Hypoxia-induced autophagy also occurs in cells
deficient in BNIP3. The unfolded protein response in
the endoplasmic reticulum is activated by hypoxia, and
part of this response includes activation of PERK
(protein kinase regulated by RNA (PKR)-like endoplasmic reticulum kinase). Through the PERK-responsive transcription factors CHOP (CCAAT-enhancerbinding proteins (C/EBP) homologous protein) and
ATF4 (activating transcription factor 4), the autophagy
genes Atg5 and MAP1LC3B are transcriptionally
induced. The induction of MAP1LC3B transcription is
necessary to maintain autophagy as LC3 is consumed by
the autophagic processes in hypoxic cells (Rouschop
et al., 2010).
As mentioned previously, mTOR is a critical regulator of autophagy and also functions as a signal
transducer for decreased oxygen availability. Hypoxia
signals to mTOR through the transcriptional target
REDD1, a protein, which is activated by HIF-1a and
represses mTORC1, resulting in increased autophagy
(Brugarolas et al., 2004). Inhibition of mTOR also
occurs with increased ROS generated during hypoxia.
ROS activates ataxia telangiectasia mutated (ATM),
leading to the subsequent phosphorylations of LKB1,
AMPK and finally TSC2, resulting in the stimulation of
TSC2 activity and subsequent inhibition of mTOR
(Alexander et al., 2010). Thus, inhibition of mTOR is
another mechanism by which hypoxia and ROS
promote autophagy (Figure 2).
Although high levels of ROS are deleterious to cells,
lower concentrations of ROS may be beneficial owing to
their enhancement of autophagy. During nutrient starvation, accumulation of ROS results in the maintenance of
autophagy through oxidation of Atg4A (Scherz-Shouval
et al., 2007). The Atg4 isoforms A–D are cysteine proteases
that drive autophagosomal maturation by cleaving LC3
prior to conjugation with phosphatidyl-ethanolamine (PE),
and subsequently de-lipidate LC3 after autophagosome–
lysosome fusion. Oxidation of the catalytic cysteine in
Atg4A inhibits its activity, thereby suppressing the delipidation of LC3, promoting the accumulation of LC3-PE.
Expression of a non-oxidizable Atg4A or Atg4B mutant
prevents autophagosome formation. Thus, ROS-mediated
inhibition of Atg4 is required for the maintenance of
autophagy during nutrient starvation.
In at least one instance, induction of autophagy
increases ROS and results in cell death. Upon caspase
inhibition, stimulation of autophagy results in the
degradation of the ROS scavenger, catalase. In this
setting, autophagy enhances ROS accumulation by
removing a major ROS-detoxifying enzyme (Yu et al.,
2006). However, this finding may be specific for the
context of caspase inhibition, as the majority of the
studies detailed above show that autophagy induced by
hypoxia or ROS enables cellular survival through
clearance of dysfunctional mitochondria and reduction
of ROS (Figure 2).
Effects of metabolic reprogramming on autophagy:
Although Warburg initially hypothesized that tumor
cells switch from oxidative phosphorylation to aerobic
glycolysis because of defective mitochondria, it has
become clear that mitochondrial function is vital to
support anabolic metabolism in rapidly proliferating
tumor cells. One such mitochondrial process, the TCA
cycle, is essential for generating both NADPH and
precursors of fatty acid synthesis. The consumption of
glucose through glycolysis at the expense of oxidative
phosphorylation necessitates the use of an alternative
fuel for mitochondrial processes. Glucose and glutamine
are the two most highly metabolized nutrients in tumor
cells, and glutamine has emerged as a dominant carbon
source for the TCA cycle in highly proliferative cells.
Many proteins involved in glutamine metabolism are
upregulated in tumor cells, including glutamine transporters and enzymes involved in glutamine breakdown.
Overexpression of Myc promotes the expression of
the glutamine transporters ASCT2 and SN2, and the
mitochondrial enzyme glutaminase, which converts
glutamine to glutamate (Wise et al., 2008; Gao et al.,
2009). These events cause cells transformed with Myc to
be dependent on glutamine for survival (Yuneva et al.,
2007). Oncogenic induction of glycolysis also creates a
dependence on glutamine, as shown by the requirement
of K-Ras-transformed cells for glutamine metabolism in
anchorage-independent growth (Weinberg et al., 2010)
and re-entry into the cell cycle (Gaglio et al., 2009).
Influence of metabolic reprogramming on tumor autophagic activity
CH Eng and RT Abraham
After transport into the cell, glutamine is metabolized
to glutamate by mitochondrial glutaminase. This reaction liberates one molecule of ammonia. The further
conversion of glutamate to a-ketoglutarate leads to the
production of a second molecule of ammonia if the
reaction is mediated by glutamate dehydrogenase.
Ammonia is also produced through the de-amidation
of other amino acids, although in tumor cells it is
thought that glutamine is the main source of free
ammonia. Although ammonia has nutritive properties
in plants and fungi, a large body of evidence suggests
that it is primarily a toxic waste product in mammalian
cells. Indeed, organisms have developed efficient means
of ammonia detoxification, through concentration in the
liver where it is converted to urea, which is excreted by
the kidneys. However, in localized regions, such as the
interstitial space of tumors that may not be readily
accessible to the bloodstream, ammonia can accumulate
to low millimolar levels (Eng et al., 2010).
Recent findings suggest that the ammonia present in
tumor interstitial space might be beneficial to tumor cell
survival, through stimulation of autophagy. Although
higher concentrations of ammonia block autophagy
by raising the lysosomal pH, lower concentrations
(2–4 mM) of ammonia paradoxically promote autophagy
in cancer cells (Eng et al., 2010). Basal autophagy, which
maintains cellular viability, is decreased in cells cultured
in glutamine-deficient medium, whereas increasing
glutamine concentrations upregulates basal autophagy
(Sakiyama et al., 2009). Ammonia-induced autophagy
protects cells from apoptosis induced by tumor necrosis
factor-a and limits cell proliferation under metabolic
stress (Eng et al., 2010). Cell death induced by long-term
culture in glutamine-free medium is also rescued by low
concentrations of ammonia (Meng et al., 2010). As these
autophagy-promoting concentrations of ammonia are
also found in tumor interstitial fluids, ammonia may act
as a diffusible autophagy-inducing signal that protects
tumor cells located in poorly vascularized regions from
metabolic stress (Eng and Abraham, 2010). Thus,
glutamine enhances tumor cell proliferation and survival
not only through the anabolic effects of replenishment
of TCA cycle intermediates, but also through the
catabolic protection from metabolic stress offered
through autophagy.
Effects of metabolic deregulations on autophagy:
Upregulation of de novo lipogenesis is another metabolic
alteration associated with tumor aggressiveness. In
contrast to normal cells, where the primary sources of
fatty acids are exogenous dietary lipids, tumor cells
synthesize the majority of their fatty acids. This
lipogenic phenotype may be related to the Warburg
effect, as a significant portion of glucose taken up by
tumor cells is used for the production of fatty acids
(DeBerardinis et al., 2007; DeBerardinis et al., 2008).
Three enzymes (ATP citrate lyase, acetyl-CoA carboxylase-A and fatty acid synthase) involved in the
conversion of the TCA cycle intermediate, citrate, to
fatty acids are upregulated in tumor tissues (Menendez
and Lupu, 2007), and expression of fatty acid synthase is
correlated with poorer prognosis in breast cancer
(Kuhajda et al., 1994). Upregulation of fatty acid
synthesis during tumorigenesis might be necessary for
increased membrane synthesis and production of lipid
messengers such as phosphatidylinositol-3,4,5-triphosphate and lysophosphatidic acid, and indeed, inhibition
of fatty acid synthase decreases tumor cell proliferation
(Lupu and Menendez, 2006). In addition, transcriptional profiling of an isogenic cell line panel manifesting
increasing levels of malignant behavior showed correlations with lipid metabolism (Hirsch et al., 2010).
Although little is known about the relationship
between cancer-mediated abnormalities in lipogenesis
and autophagy, recent work suggests that autophagy
regulates lipid metabolism in normal tissues. Triglycerides and other lipids are substrates for autophagic
degradation, and inhibition of autophagy leads to
increased triglyceride lipid stores and decreased triglyceride breakdown in hepatocytes (Singh et al., 2009).
Fatty acids are stored in lipid droplets in the form of
triglycerides. As inhibition of autophagy promotes
triglyceride storage, it may counteract the effects of
increased fatty acid synthesis owing to overexpression of
lipogenic enzymes in tumor cells.
In order to maintain cellular homeostasis in the setting
of rapid cellular proliferation, transformed cells alter
numerous metabolic pathways. Increased glucose uptake through glycolysis hastens ATP production and
this altered utilization of glucose necessitates an
upregulation of glutamine metabolism in order to fuel
mitochondrial pathways. Anerobic glycolysis is also
necessary for cells to adapt to a hypoxic microenvironment, which is associated with both nutrient deprivation
and increased levels of ROS. All of these metabolic
perturbations result in a stimulation of autophagy.
Autophagy appears to have dual roles in tumorigenesis. Suppression of autophagy may be a requirement
for the early stages of transformation, as evidenced by
the observations that autophagy-defective cells are more
tumorigenic than their wild-type counterparts. In addition, autophagy might be diminished during oncogeneinduced transformation (such as by activation of PI3K),
although there are clear exceptions (such as mutation of
K-Ras). If one could intervene early on in the
tumorigenic process, stimulation of autophagy might
promote growth suppression and counter ROS generation, organelle dysfunction and genomic instability,
thereby interfering with tumor formation.
However, at the time of diagnosis, tumors are
generally well-established. Metabolic alterations such
as increased glycolysis, glutaminolysis and hypoxia/
ROS accumulation that accompany the establishment of
tumors results in the upregulation of autophagic flux.
Thus, therapeutic inhibition of autophagy at this stage
Influence of metabolic reprogramming on tumor autophagic activity
CH Eng and RT Abraham
Tumor Cell Survival
Proliferative and
survival signals
Proliferative and
survival signals
tumor stromal cells
challenged tumor cells
nutrients, oxygen
replete tumor cells
nutrients, oxygen
Tumor Cell Death
Figure 3 Effect of autophagy inhibition within the tumor landscape. The tumor microenvironment contains a gradient of oxygen and
nutrients (increasing from green to purple), within which reside tumor cells themselves and supporting stromal cells. Autophagy is
stimulated in tumor cells owing to metabolic deregulations and glutaminolytic production of ammonia, which acts as a diffusible
stimulator of autophagy in the neighboring, metabolically challenged cells (green). Tumor cells also stimulate autophagy in adjacent
stromal cells (brown), which allows the stromal cells to impart proliferative and survival signals back to the tumor cells. Inhibition of
autophagy would block both autocrine- and paracrine-mediated survival pathways, resulting in tumor cell death.
would result in a bioenergetic imbalance, and render
cancer cells unable to cope with their metabolically
stressed microenvironment and promote their demise.
The altered metabolic program and persistent proliferation of tumor cells also results in misfolded proteins,
damaged mitochondria, ROS and excess lipids, and
without autophagy to alleviate the effects of these
harmful agents, tumor cells might be coaxed into
poisoning themselves with the byproducts of their own
metabolic alterations. Additionally, inhibition of autophagy might block pathways used by tumor cells to
survive chemotherapeutic stress, and thus sensitize cells
to cytotoxic chemotherapy.
Finally, communication between tumor cells and
tumor stroma also contributes to the metabolic and
autophagic landscape of the tumor microenvironment
(Figure 3). The high rate of glutaminolysis in tumor cells
results in the production of ammonia, which could
diffuse to the nutrient- and oxygen-limited regions of the
tumor to combat metabolic stress by stimulating
autophagy (Eng and Abraham, 2010). Tumor-associated
stromal cells also reside in the same metabolically
stressed environment as tumor cells, and contribute to
the survival and proliferation of the tumor cells
themselves (McAllister and Weinberg, 2010). Part of
this survival mechanism is because of autophagy
induced in adjacent cancer-associated fibroblasts (Martinez-Outschoorn et al., 2010). Thus, inhibition of
autophagy would not only promote tumor cell death
in a cell-autonomous manner, but also interfere with
cellular survival that results from heterogeneous cell–cell
interactions within the diverse tumor landscape.
A deeper understanding of the metabolic drivers within
specific tumor types and the contributions of distinct
tumor compartments will guide the use of autophagic
inhibitors as efficacious therapeutic agents.
Conflict of interest
The authors declare no conflict of interest.
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