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Extracellular acidity as favouring factor of tumor progression and metastatic dissemination



The bidirectional interactions between tumor cells and the so-called "host reactive stroma" play a critical role in most of the events characterizing tumor progression and distant organ colonization. This review discusses critical components of tumor environment involved in tumor cell dissemination. More specifically, it addresses some of the experimental evidences providing that acidity of tumor environment facilitates local invasiveness and metastasis formation, independently from hypoxia, with which acidity may be associated. Besides, acidity renders tumor cells resistant to radiation therapy and chemotherapeutic drugs. Therefore, this review examines the strategies for raising the low extracellular pH of tumors that might have considerable potential in cancer therapy.
Experimental Oncology 34, 79–84, 2012 (June) 79
L. Calorini*, S. Peppicelli, F. Bianchini
Dipartimento di Patologia e Oncologia Sperimentali, Universitа degli Studi di Firenze, e Istituto Toscano
Tumori, Firenze, Italy
The bidirectional interactions between tumor cells and the so-called “host reactive stroma” play a critical role in most of the events
characterizing tumor progression and distant organ colonization. This review discusses critical components of tumor environment
involved in tumor cell dissemination. More specifically, it addresses some of the experimental evidences providing that acidity of tumor
environment facilitates local invasiveness and metastasis formation, independently from hypoxia, with which acidity may be associ-
ated. Besides, acidity renders tumor cells resistant to radiation therapy and chemotherapeutic drugs. Therefore, this review examines
the strategies for raising the low extracellular pH of tumors that might have considerable potential in cancer therapy.
Key Words: extracellular acidity, invasiveness, metastatic dissemination, proton pump inhibitors.
Capacity of cancer to evolve and change has been
named “tumor progression” [1]. Biological character-
istics that define tumor progression have been exten-
sively described, although the underlying mechanisms
remain unknown. Malignant tumor cells accumulate
increasingly genetic alterations, generated by random
mutational events, leading them to assume all the
characteristics of invasive cells. In concert with this
“genetic instability”, a key role in favouring changes
in tumor cells is played by local host factors [2, 3].
Among local factors, particular attention has been
devoted to the interactions that tumor cells establish
with various host cells that reside in or are attracted into
tumor environment. The bidirectional interaction be-
tween tumor cells and host cells, is recognized as cru-
cial for the decision whether tumor cells progress
toward metastatic dissemination or remain dormant
[4–8]. Indeed, tumor growth and metastasis is sig-
nificantly reduced in fibroblast-deficient mice, while
injection of wild-type fibroblasts into these mice can
reverse this phenotype, providing a clear evidence for
the involvement of fibroblasts in the emergence of me-
tastasis [9–11]. This type of activated cells, commonly
identified by the expression of -smooth muscle actin
(-SMA) and referred as “myofibroblasts” [12], was
named cancer-associated fibroblasts (CAFs) and ac-
tively participates at all stages of metastatic cascade.
In addition cells of monocyte/macrophage lineage
enter into the tumor mass via blood vessels throughout
life span of tumors, from early-stage lesions to late-
stage tumors that are invasive and metastatic, and are
indicated as tumor-associated macrophages (TAMs)
[13, 14]. TAMs are remarkable for the diverse activi-
ties in which they can engage on different occasions.
Quiescent macrophages respond to immune or bac-
terial stimuli by expressing new functional activities,
resulting in their capacity to recognize and destroy
transformed cells. On the contrary, macrophages
isolated from experimental and spontaneous tumors
show a reduced level of cytotoxic activities and was
proved to be relevant to tumor progression and mes-
tastases [15, 16]. Plasticity of both CAFs and TAMs
may be exploited by tumor cells to elicit distinct func-
tions at different stages of tumor progression. It is also
possible that changes expressed by these host cells
during tumor development might be related to their
location inside the tumor mass. Most tumors develop
an environment characterized by low oxygen tension
(hypoxia), elevated interstitial fluid pressure, low glu-
cose concentration and high lactate concentration.
These changes are largely caused by a combination
of poor tissue perfusion due to abnormal tumor vas-
culature, uncontrolled proliferation and altered energy
metabolism [17].
Cells require oxygen and nutrients for their survival
and growth. Likewise, neoplastic cells depend on near-
by capillaries for growth and once aggregates of tumor
cells reach the diffusion limit for critical nutrients and
oxygen, tumor cells become dormant. Indeed, some
human tumors can remain dormant for a number
of years at a stage where tumor cell proliferation and
death are balanced. But once new blood vessel forma-
tion is initiated, the so-called angiogenic switch, tumor
progression and metastasis follow [18]. The new vessel
formation governed by a balance of pro- and anti-
angiogenenic factors is often disturbed in tumors lea-
ding to a vasculature characterized by dilated, tortuous
and incomplete vessels. The molecular mechanisms
causing this abnormal vascular architecture are still
debated, but the uncontrolled vascular endothelial
Received: May 23, 2012.
*Correspondence: E-mail: lido.calorini
Abbreviations used: α-SMA — α-smooth muscle actin; CA — car-
bonic anhydrase; CAFs — cancer-associated fibroblasts; ECM —
extracellular matrix; HIF — hypoxia-inducible factor; MCTs — mono-
carboxylate transporters; MMPs — matrix metalloproteinases;
NHEs — Na+/H+ exchangers; PAI — plasminogen activator inhibitors;
TAMs — tumor-associated macrophages; uPA — urokinase-type
plasminogen activator; uPAR — uPA receptor; V-ATPase — vacuolar
H+-ATPase; VEGF-A — vascular endothelial growth factor A.
Exp Oncol 2012
34, 2, 79–84
80 Experimental Oncology 34, 79–84, 2012 (June)
growth factor A (VEGF-A) signalling may be a key
contributor. VEGF-A is a strong mitogen and survival
growth factor for vascular endothelial cells and induces
mobilization and recruitment of endothelial precursor
cells [19, 20]. Furthermore, VEGF-A contributes to the
angiogenic phenotype by increasing the permeability
of existing vessels, permitting extravasation of fibrino-
gen and clotting factors and resulting in a fibrin-rich
stroma that supports migration of endothelial cells
and formation of new vasculature. However, the
uncontrolled secretion of VEGF-A results in a lower
perfusion rates in tumors than in many normal tissues.
Blood flow in tumors is unevenly distributed and can
even reverse its direction in some vessels, therefore,
regions with poor perfusion are common. These
environmental features vary widely in different areas
of tumors, reflecting tumor cell heterogeneity. In addi-
tion, the uncontrolled growth of tumor cells compress
the intra-tumor lymphatic vessels. Consequently, there
are no functional lymphatic vessels inside solid tumors,
whereas functional lymphatic vessels are present only
in peri-tumoral tissues [21, 22]. Both, the high perme-
ability of tumor blood vessels and the lack of functional
lymphatics are keys contributors to the development
of an interstitial hypertension in neoplastic tissues
[23]. As a result, the hydrostatic and colloid osmotic
pressures become almost equal between intravascular
and extravascular spaces, compromising the delivery
of nutrients as well as therapeutic agents. Since tumor
interstitial hypertension is a reflection of global patho-
physiology of tumors, it may be used for diagnosis
and/or prognosis. The consequent metabolic hallmark
of tumor environment is hypoxia. Hypoxia character-
izes the microenvironment of many solid tumors and
it has been shown to affect many biological proper-
ties of tumor cells implicated in tumor progression,
response to therapy, including clinical outcome of pa-
tients [24–26]. The mechanism behind these effects
is related to the induction of hypoxia-inducible factor
(HIF) family of transcription factors. Under conditions
of acute or chronic hypoxia, HIF-1 is stabilized, form
a heterodimer with HIF-1, allowing this factor to bind
a core sequence and increase transcription of target
genes. This factor regulates many cellular processes
including apoptosis, cell proliferation, angiogenesis
and glucose metabolism [27, 28]. Thus, hypoxia in-
creases genetic instability, blood vessel formation and
a switch to anaerobic metabolism.
Hypoxia, elevated interstitial fluid pressure, low
glucose and high lactate concentration resulting from
a predominant anaerobic metabolism, are responsible
of low extracellular pH (pHe) in tumor tissues. As a con-
sequence, the second metabolic hallmark of tumor
environment is tumor acidosis.
In this review, we will discuss evidence that aci-
dity of tumor extracellular space represents a direct
contributor to the process of tumor progression and
that normalization of pHe could be considered a new
strategy for tumor therapy.
In contrast to normal cells, which rely on mitochon-
drial oxidative phosphorylation to generate the energy
needed for cellular processes, most cancer cells,
even in the presence of sufficient oxygen to support
mitochondrial respiration, use “aerobic glycolysis”,
a phenomenon termed “the Warburg effect” [29,
30]. This phenomenon was first reported by Warburg
in the 1920s, leading to hypothesis that cancer results
from impaired mitochondrial metabolism. Although
the “Warburg hypothesis” has proven incorrect,
an increased conversion of glucose to lactic acid
in tumor cells has been continuously demonstrated
(½ (glucose) = lactate- + H+). The clinical application
of the imaging technique positron-emission tomog-
raphy (PET) using the glucose analog 2-(18F)-fluoro-
2-deoxy-D-glucose (FDG) tracer, demonstrated that
most primary and metastatic human lesions express
a high glucose uptake [31]. FDG-PET combined with
computer tomography (PET/CT) has a high sensitivity
and specificity for the detection of metastases of most
epithelial cancers. A possible explanation for the
switch to aerobic glycolysis is that proliferating tumor
cells have important metabolic requirements beyond
ATP, and some glucose must be diverted to macromo-
lecular precursors such as acetyl-CoA for fatty acids,
glycolytic intermediates for nonessential amino acids
and ribose for nucleotides. Moreover, some tumors
possess a greater capacity to pump lactic acid and
protons out to the extracellular space through specific
transporters, to maintain an appropriate neutral/slight
alkaline intracellular pH essential for cell proliferation.
The inefficient removal of protons and lactic acid from
the extracellular spaces, due to the poorly perfused
tumor tissue and absence of functional lymphatic
vessels, creates a reversed pH gradient characte-
rized by an acidic pHe and an alkaline intracellular pH
(pHi) [32].
In vitro and in vivo studies revealed that tumor cells
have pHi ranging from 7.12 to 7.56 (pHi of normal cells:
6.99-7.20), and pHe of 6.2-6.9 (pHe of normal extracel-
lular space: 7.3-7.4). Degree of acidity in tumors tends
to be associated with a poorer prognosis [33]. Indeed,
tumor acidity contributes to aggressiveness of tumor
cells, stimulating increased mutation rate [34]. Acute
and chronic acidosis, hypoxia and reoxygenation injury
all together promote DNA instability even in very small
tumors leading to the selection of cells with additional
genetic defects. Moreover, a minimum in pHe has been
observed near tumor periphery, where tumor cells are
invading normal tissues [35]. Hypoxia, also, stimulates
invasiveness in tumor cells [27]. Could be expected
that low extracellular pH and hypoxia always co-
localize within tumor regions, instead, there is often
a lack of spatial correlation among these parameters.
Potential explanations of this lack of correlation could
be due to the enhanced glucose uptake for glycolytic
ATP generation in conditions of high oxygen tension,
or to the possibility that some tumor vessels carrying
hypoxic blood, are unable to deliver adequate quan-
Experimental Oncology 34, 79–84, 2012 (June) 81
tity of oxygen to the cells, but are able to carry away
the waste products (e.g., lactic acid). Low pHe has
shown to affect several steps of metastatic cascade.
In some tumor cells, low pH promotes angiogenesis
through VEGF [36] and IL-8 [37], however in other
models of tumor cells acidosis inhibits VEGF [38].
Role of acidic pH in angiogenesis is still not completely
understood. On the other hand, influence of acidity
in invasiveness of tumor cells into host tissues is well
demonstrated. Invasiveness is a multistep process
based on extracellular matrix-degrading proteinases,
such as serine and metallo-proteinases, reorganiza-
tion of cytoskeleton and an integrin-mediated for-
mation and release of focal adhesion contacts [39].
It has been reported that an acidic pHe may enhance
invasion of tumor cells facilitating the redistribution
of active cathepsin B, a lysosomal aspartic proteinase
with acidic pH optima, to the surface of malignant cells
[40, 41]. Acid-activated catepsins L also participate
to amplify proteinase cascade through activation of
urokinase-type plasminogen activator (uPA) [42]. The
uPA system, made by uPA, two main plasminogen
activator inhibitors (PAI-1, PAI-2) and uPA receptor
(uPAR), is critical for tumor cell-driven degradation
of extracellular matrix (ECM) in many steps of meta-
static cascade [43, 44]. Activation of cathepsins D and
L in an acidic tumor environment reduce perfusion
of tumor regions, generating angiogenesis inhibitors
such as angiostatin [45] and endostatin [46] from pro-
teolysis of plasminogen and collagen, enhancing the
chaotic vascular organization of tumors. Furthermore,
acidic pH can promote the conversion of matrix me-
talloproteinases (MMPs) in their active forms. MMPs
have long been associated with invasiveness and dis-
semination of tumor cells, due to their capacity to help
tumor cells to cross structural barriers, inclu ding base-
ment membranes and structural components of the
ECM, such as collagen fibers [47–49]. Degradation
of structural components of ECM is considered es-
sential in tumor-induced angiogenesis. MMPs also
participate in the release of cell-membrane-precursor-
forms of many growth factors. The expression of MMPs
in tumors is regulated in a paracrine manner by growth
factors and inflammatory cytokines secreted by tumor
infiltrating inflammatory cells as well as tumor cells
themselves, and a continuous cross talk between tu-
mor cells and inflammatory cells during the invasion
process was demonstrated. Incubation of human and
mouse melanoma cells in a low pH medium stimulate
MMP expression and an increase in vitro invasiveness
and in vivo metastasis formation in immunodeficient
mice [50–53].
Another important component of basement mem-
brane to be degraded by tumor cells to disseminate
are the heparan sulphate chains. Toyoshima and
Nakajima report that heparanase has an optimal
pH of 4.2, but a significant heparanase activity persists
at pH 6.0–6.5, suggesting that the acidic environment
of tumors may activate the degrading properties of tu-
mor heparanases [54].
More recently, two of the most important H+ trans-
porters, the ubiquitously expressed Na+/H+ exchanger
isoform (NHE1) [55] and the plasma membrane type
of vacuolar H+-ATPases (V-ATPases) [56] were found
to be implicated in migration of tumor cells. NHE1 in-
fluences the formation of invadopodia, structures that
regulate cell motility [57]. Cell motility is driven by cy-
cles of actin polymerization, integrin-mediated cell
adhesion and acto-myosin contraction. Thus the
moving tumor cells, in the absence of proteinases,
make contact with collagen fibers and proceed along
fibers [39]. V-ATPases are a family of ATP-dependent
proton pumps particularly expressed by invasive pan-
creatic [58] and breast carcinomas [59], and inhibition
of V-ATPases expression in hepatocarcinoma using
siRNA abrogates invasion and metastatic diffusion
of these tumor cells [60]. On the whole, an acidic
condition may potentiate several proteinases critical
for tumor cells when they detach from the primary tu-
mor, migrate into the blood, extravasate and colonize
in distant host tissues.
Moreover, extracellular lactic acid can suppress
tumoricidal activity of cytotoxic lymphocytes and
natural killer cells, an effect mediated by lactate/H+
co-transporter that under neutral conditions remove
lactic acid from leukocytes [61]. Acidification of the ex-
tracellular space may also influence radiation therapy
and chemotherapy. Indeed, acidity of tumors reduce
sensitivity of tumor cells to radiation therapy [62–64].
This protective effect is considered to be due to the
decreased fraction of proliferating tumor cells [65]
and the reduced fixation of radiation-induced DNA
damage [66].
Extracellular acidity also confers a special re-
sistance against weakly basic drugs to tumor cells.
It has been reported that chronic and acute sodium
bicarbonate-induced alkalosis is able to circumvent
this drug resistance and enhance the anti-tumor ac-
tivities of two weakly basic drugs, such as doxorubicin
[67] and mitoxantrone [68]. These results suggest
that induction of metabolic alkalosis using sodium
bicarbonate can produce a net gain in the therapeutic
index of the several chemotherapeutic agents, and
open up the possibility that normalization of pHe may
have a therapeutic utility.
Since acidity of tumor environment appears
to contribute to cancer aggressiveness, chemo- and
radiation resistance and, even, evasion of immune
reactions, measures to normalize pHe of tumors may
be used in tumor therapy.
A number of researches have explored the pos-
sibility to correct the extracellular acidity of tumors.
Studies revealed that in tumors levels of CO2 are
higher and concentration of bicarbonate, the princi-
pal physiologic buffer used to control pH, are lower
than in blood or in healthy tissues [69, 70]. Therefore,
it is possible that an increased concentration of sodium
bicarbonate can reduce aggressiveness of tumor
82 Experimental Oncology 34, 79–84, 2012 (June)
cells. Indeed, the alkalization of melanoma-bearing
animals by sodium bicarbonate was found to inhibit
the development of spontaneous metastases [71]. In-
terestingly, a similar dose of bicarbonate used in these
latter experiments has been administered chronically
(>1 year) in patients with renal tubular acidosis [72]
and sickle cell anemia without adverse effects [73].
Computer simulation used to verify the ability of so-
dium bicarbonate to increase pHe of tumors in vivo
also indicates that the normalization of tumor acidity
reduces invasiveness of tumor cells without altering
the pH of blood or normal tissues [74].
As an alternative strategy for correcting low pHe,
several authors explored the inhibition in tumor cells
of key pH regulators that maintain a neutral/alkaline
intracellular pH by extruding lactate or protons [75].
pH regulators in tumor cells include extracellular
forms of carbonic anhydrase (CA), Na+/HCO3- co-
transporters, Na+/H+ exchangers (NHEs), mono-
carboxylate transporters (MCTs) and the vacuolar
H+-ATPase (V-ATPases). The raise of pHe promoted
by these inhibitors is constantly associated with a de-
crease of intracellular pH. Acidity of pHi tends to sup-
press the efficiency of glycolysis, sustaining the raise
of pHe [76], and may exert anti-proliferative and pro-
apoptotic effects on tumor cells themselves [77–79].
Consequently, pH regulators might be considered true
anticancer drugs.
Two CA isozymes, CA9 and CA12, are overex-
pressed in tumor cells and their activity is associated
with malignancy and resistance to therapy [80]. Sul-
phonamide CA inhibitors that target CA9 were found
effective to block the growth of primary tumor and
metastases in a mouse model of breast cancer [81].
Some of these compounds are in advanced preclini-
cal evaluation. V-ATPases, while originally identified
in intracellular compartments, they have increasingly
been shown to play essential roles in proton transport
across the plasma membrane of a variety of cell types,
including tumor cells [59, 76, 82]. The likely similarity
between V-ATPase and the H+/K+ ATPase, the enzyme
involved in proton secretion in gastric parietal cells,
prompted the investigation of proton pump inhibitors
(PPIs), such as omeprazole and esomeprazole, for
inhibiting V-ATPase. When activated by acidic pHe
of tumors, these drugs can inhibit V-ATPase by a co-
valent interaction. Both in vitro and in vivo experi-
ences indicate that non-toxic doses of PPIs, analo-
gous to those used for treatment of Zollinger-Ellison
syndrome, exert anti-proliferative and pro-apoptotic
effects on melanoma cells [83]. PPIs were also dem-
onstrated to inhibit growth of B-cell lymphoma cells
transplanted into severe combined immunodeficient
mice [77]. Knockdown of V-ATPase expression by siR-
NA in cells isolated from a human hepatocarcinoma
markedly reduced metastatic dissemination of these
cells [84]. Importantly, Hashioka et al report that PPIs
have anti-inflammatory effects and decrease mono-
cytic neurotoxicity [85]. Recently, Lee et al. found that
omeprazole exerts a cancer-preventive role against
colitis-induced carcinogenesis, a chemopreventive
action independent of gastric acid suppression [86].
Evidence that PPIs play a role in normalization of low
pH and abrogate inflammation, renders these drugs
suitable to target critical mechanisms involved in tumor
progression. Moreover, clinical data provide that PPIs
have a very low level of systemic toxicity as compared
with standard chemotherapeutic agents. NHE is crucial
in pH regulation and is expressed in eve ry cell type.
There are several NHE inhibitors, structu rally related
to amiloride and cariporide, however the diffused
presence of NHE in many tissues and its role in crucial
physiological processes, confers to this class of agents
potential risk of side effects. Inhibitors of V-ATPase
and NHE have been shown to have an additive im-
pact on intracellular pH and on thermosensitization
[87]. Therefore, it is crucial to develop agents that
selectively target NHE in tumor. At the same time,
potent, non-toxic selective MCT inhibitors are needed.
MCTs, are overexpressed in many tumors and the
isoform MCT1 regulates the entry and exit of lactate
from tumor cells. The inhibition of MCT1 was found
to induce a switch from lactate-fuelled respiration
to glycolisis, which was accompanied by a retardation
of tumor growth in a mouse model of lung carcinoma
and in transplanted human colorectal carcinoma [88].
Tumor stroma manifest some degree of plasti-
city, a property controlled by tumor cells themselves.
Indeed, tumor cells influence host stromal elements
to produce relevant effectors that act as tumor pro-
moters. The metabolic hallmarks of this space are
hypoxia and acidosis. We have elucidated how the
extracellular acidity per se, may promote an aggres-
sive and metastatic phenotype in tumor cells and how
these findings suggest the possibility of a novel and
effective therapeutic strategy based on the control
of tumor acidity.
This study was funded by grants from Istituto
Toscano Tumori, Ente Cassa di Risparmio di Firenze.
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Copyright © Experimental Oncology, 2012
... The acidic tumor microenvironment is the result of tumor metabolism, which conversely affects the metabolism and biological behavior of cancer cells. Studies have shown that the acidic tumor microenvironment promotes local tumor infiltration and metastatic tumor growth independent of hypoxic conditions (Calorini et al. 2012;Peppicelli et al. 2014), and the pH around the tumor site is approximately 6.7-7.1 (Abadjian et al. 2017;Gallagher et al. 2008;Stuwe et al. 2007). Thus, human CRC cells were cultured in alkaline (pH 7.4) and acidic (pH 6.8) culture media to simulate the acidic tumor microenvironment, and the results showed that cell migration was promoted in acidic (pH 6.8) culture medium compared with alkaline (pH 7.4) culture medium, which was consistent with previous studies (Calorini et al. 2012;Peppicelli et al. 2014). ...
... Studies have shown that the acidic tumor microenvironment promotes local tumor infiltration and metastatic tumor growth independent of hypoxic conditions (Calorini et al. 2012;Peppicelli et al. 2014), and the pH around the tumor site is approximately 6.7-7.1 (Abadjian et al. 2017;Gallagher et al. 2008;Stuwe et al. 2007). Thus, human CRC cells were cultured in alkaline (pH 7.4) and acidic (pH 6.8) culture media to simulate the acidic tumor microenvironment, and the results showed that cell migration was promoted in acidic (pH 6.8) culture medium compared with alkaline (pH 7.4) culture medium, which was consistent with previous studies (Calorini et al. 2012;Peppicelli et al. 2014). We further assessed the enzyme changes during the glucose metabolism pathway through metabolic PCR that contained 384 genes related to glucose metabolism. ...
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Purpose This study was to investigate the biological effect of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase 2 (PFKFB2) in colorectal cancer (CRC). Methods PFKFB2 was selected by metabolism polymerase chain reaction (PCR) array from CRC cells under alkaline culture medium (pH 7.4) and acidic culture medium (pH 6.8). The expression of PFKFB2 mRNA and protein was detected by quantitative real-time PCR and immunohistochemistry in 70 paired fresh and 268 paired paraffin-embedded human CRC tissues, respectively, and then the prognostic value of PFKFB2 was investigated. The effects of PFKFB2 on CRC cells were also verified in vitro, which were through detecting the change of migration, invasion, sphere formation, proliferation, colony formation, and extracellular acidification rate of CRC cells after PFKFB2 knockdown in alkaline culture medium (pH 7.4) and overexpression in acidic culture medium (pH 6.8). Results PFKFB2 expression was downregulated in acidic culture medium (pH 6.8). In addition, we found PFKFB2 expression decreased in human CRC tissues compared with the adjacent normal tissues. Furthermore, the OS and DFS rate of CRC patients with low PFKFB2 expression was significantly shorter than those of patients with high PFKFB2 expression. Multivariate analysis indicated that low PFKFB2 expression was an independent prognostic factor for both OS and DFS in CRC patients. Moreover, the abilities of migration, invasion, spheroidizing ability, proliferation, and colony formation of CRC cells were significantly increased after depletion of PFKFB2 in alkaline culture medium (pH 7.4) and decreased after overexpression of PFKFB2 in acidic culture medium (pH 6.8) in vitro. Epithelial–mesenchymal transition (EMT) pathway was found and verified involved in the PFKFB2-mediated regulation of metastatic function in CRC cells. Further, glycolysis of CRC cells was significantly elevated after knockdown of PFKFB2 in alkaline culture medium (pH 7.4) and decreased after overexpression of PFKFB2 in acidic culture medium (pH 6.8). Conclusion PFKFB2 expression is downregulated in CRC tissues and associated with worse survival for CRC patients. PFKFB2 could inhibit metastasis and the malignant progression of CRC cells by suppressing EMT and glycolysis.
... Whereas hypoxia and nutrient deprivation have been welldocumented, TMA in PDAC has not been intensively studied. However, recent studies have confirmed that TAM is involved in initiating the early events of malignant transformation [17][18][19][20]; more crucially, promoting tumor progression and metastasis. In the context of TMA-mediated cancer progression, it is well known that invasiveness and metastasis of cancer cells are accelerated by a variety of extracellular proteases such as metalloproteinases (MMPs), thiol proteases, serine proteases and acid proteases, which are responsible for degrading the tumor barriers, creating ideal condition to favor tumor metastasis. ...
Pancreatic ductal adenocarcinoma (PDAC) is an aggressive and devastating disease, which is characterized by invasiveness, rapid progression and profound resistance to treatment. It has been best characterized that tumor microenvironment such as hypoxia and nutrient deprivation contributes to cancer progression; however, the role of tumor microenvironment acidification (TMA), a major feature of tumor tissue, has not been intensively studied. Interestingly, clinicopathological clues have recently unraveled that TMA is involved in promoting cancer progression although the exact signaling pathways is poorly understood. In PDAC, the TAM is tightly regulated by proton (H+) transporters and pumps. This review dissects and summarizes the roles of these H+-extruding regulators in facilitating PDAC progression.
... Multiple sovereign studies have established that lactate produced by hypoxic tumors reaches oxidative tumor cells by the monocarboxylate transporter 1, and it is exploited there as a significant mitochondrial energy substrate [101][102][103]. Mechanistically, lactate imitates as a quencher to free radicals to keep an eye on the oxidative stress in tumors and induce radioresistance in tumor cells [104][105][106]. Lactate also overpowers and modifies the immune cell function to repress host immunosurveillance and endorse tumor cell metastasis [107][108][109][110]. ...
Pyruvate is irreversibly decarboxylated to acetyl coenzyme A by mitochondrial pyruvate dehydrogenase complex (PDC). Decarboxylation of pyruvate is considered a crucial step in cell metabolism and energetics. The cancer cells prefer aerobic glycolysis rather than mitochondrial oxidation of pyruvate. This attribute of cancer cells allows them to sustain under indefinite proliferation and growth. Pyruvate dehydrogenase kinases (PDKs) play critical roles in many diseases because they regulate PDC activity. Recent findings suggest an altered metabolism of cancer cells is associated with impaired mitochondrial function due to PDC inhibition. PDKs inhibit the PDC activity via phosphorylation of the E1a subunit and subsequently cause a glycolytic shift. Thus, inhibition of PDK is an attractive strategy in anticancer therapy. This review highlights that PDC/PDK axis could be implicated in cancer's therapeutic management by developing potential small-molecule PDK inhibitors. In recent years, a dramatic increase in the targeting of the PDC/PDK axis for cancer treatment gained an attention from the scientific community. We further discuss breakthrough findings in the PDC-PDK axis. In addition, structural features, functional significance, mechanism of activation, involvement in various human pathologies, and expression of different forms of PDKs (PDK1-4) in different types of cancers are discussed in detail. We further emphasized the gene expression profiling of PDKs in cancer patients to prognosis and therapeutic manifestations. Additionally, the pharmaceutical inhibition of the PDK/PDC axis by small molecule inhibitors and natural compounds at different clinical evaluation stages has also been discussed comprehensively.
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Conventional chemotherapy, one of the most widely used cancer treatment methods, has serious side effects, and usually results in cancer treatment failure. Drug resistance is one of the primary reasons for this failure. The most significant drawbacks of systemic chemotherapy are rapid clearance from the circulation, the drug's low concentration in the tumor site, and considerable adverse effects outside the tumor. Several ways have been developed to boost neoplasm treatment efficacy and overcome medication resistance. In recent years, targeted drug delivery has become an essential therapeutic application. As more mechanisms of tumor treatment resistance are discovered, nanoparticles are designed to target these pathways. Therefore, understanding the limitations and challenges of this technology is critical for nanocarrier evaluation. Nano-drugs have been increasingly employed in medicine, incorporating therapeutic applications for more precise and effective tumor diagnosis, therapy, and targeting. Many benefits of nanoparticle (NP)-based drug delivery systems in cancer treatment have been proven, including good pharmacokinetics, tumor cell-specific targeting, decreased side effects, and lessened drug resistance. As more mechanisms of tumor treatment resistance are discovered, NPs are designed to target these pathways. At the moment, this innovative technology has the potential to bring fresh insights into cancer therapy. Therefore, understanding the limitations and challenges of this technology is critical for nanocarrier evaluation.
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Dynamic and heterogeneous interaction between tumor cells and the surrounding microenvironment fuels the occurrence, progression, invasion, and metastasis of solid tumors. In this process, the tumor microenvironment (TME) fractures cellular and matrix architecture normality through biochemical and mechanical means, abetting tumorigenesis and treatment resistance. Tumor cells sense and respond to the strength, direction, and duration of mechanical cues in the TME by various mechanotransduction pathways. However, far less understood is the comprehensive perspective of the functions and mechanisms of mechanotransduction. Due to the great therapeutic difficulties brought by the mechanical changes in the TME, emerging studies have focused on targeting the adverse mechanical factors in the TME to attenuate disease rather than conventionally targeting tumor cells themselves, which has been proven to be a potential therapeutic approach. In this review, we discussed the origins and roles of mechanical factors in the TME, cell sensing, mechano‐biological coupling and signal transduction, in vitro construction of the tumor mechanical microenvironment, applications and clinical significance in the TME.
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Extracellular vesicles (EVs) are heterogeneous lipid containers with a complex molecular cargo comprising several populations with unique roles in biological processes. These vesicles are closely associated with specific physiological features, which makes them invaluable in the detection and monitoring of various diseases. EVs play a key role in pathophysiological processes by actively triggering genetic or metabolic responses. However, the heterogeneity of their structure and composition hinders their application in medical diagnosis and therapies. This diversity makes it difficult to establish their exact physiological roles, and the functions and composition of different EV (sub)populations. Ensemble averaging approaches currently employed for EV characterization, such as western blotting or ‘omics’ technologies, tend to obscure rather than reveal these heterogeneities. Recent developments in single-vesicle analysis have made it possible to overcome these limitations and have facilitated the development of practical clinical applications. In this review, we discuss the benefits and challenges inherent to the current methods for the analysis of single vesicles and review the contribution of these approaches to the understanding of EV biology. We describe the contributions of these recent technological advances to the characterization and phenotyping of EVs, examination of the role of EVs in cell-to-cell communication pathways and the identification and validation of EVs as disease biomarkers. Finally, we discuss the potential of innovative single-vesicle imaging and analysis methodologies using microfluidic devices, which promise to deliver rapid and effective basic and practical applications for minimally invasive prognosis systems. Understanding the heterogeneity of extracellular vesicles is crucial for unraveling their functions. This review describes the benefits, challenges and contributions of the state-of-the art methods used in single-vesicle analysis.
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Uptake of weak acid and weak base chemotherapeutic drugs by tumors is greatly influenced by the tumor extracellular/interstitial pH (pHe), the intracellular pH (pHi) maintained by the tumor cells, and by the ionization properties of the drug itself. The acid-outside plasmalemmal pH gradient in tumors acts to exclude weak base drugs like the anthracyclines, anthraquinones, and vinca alkaloids from the cells, leading to a substantial degree of “physiological drug resistance” in tumors. We have induced acute metabolic alkalosis in C3H tumor-bearing C3H/hen mice, by gavage and by intraperitoneal (i.p.) administration of NaHCO3. 31P magnetic resonance spectroscopic measurements of 3-aminopropylphosphonate show increases of up to 0.6 pH units in tumor pHe, and 0.2 to 0.3 pH units in hind leg tissue pHe, within 2 hours of i.p. administration of NaHCO3. Theoretical calculations of mitoxantrone uptake into tumor and normal (hind leg) tissue at the measured pH, and pHI values indicate that a gain in therapeutic index of up to 3.3-fold is possible with NaHCO3 pretreatment. Treatment of C3H tumor-bearing mice with 12 mg/kg mitoxantrone resulted in a tumor growth delay of 9 days, whereas combined NaHCO3mitoxantrone therapy resulted in an enhancement of the TGD to 16 days.
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Metastasis is the primary cause of death from cancer due to the spread of disease throughout the body. Increasing evidence suggests that the hypoxic microenvironment serves as a driving force for the metastatic process. Fifty to sixty percent of solid tumors contain hypoxic areas, where the gene expression is reprogrammed by low oxygen microenvironment leading to aggressive invasive cancer cell behavior. Hypoxia upregulates multiple genes involved in different steps of metastatic process, including angiogenesis, proliferation, migration, invasion, motility, adhesion, ECM remodeling, and survival. Moreover, hypoxia confers tumor cells with chemo- and radio-resistance. At the end of this chapter, we discuss the facts linking hypoxia and cancer stem cells (CSC) mainly through the ability of hypoxic microenvironment to shift cells toward the undifferentiated phenotype.
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Studies over the last few decades have demonstrated that the intracellular pH of solid tumors is maintained within a range of 7.0-7.2, whereas the extracellular pH is acidic. A low extracellular pH may be an important factor inducing more aggressive cancer phenotypes. Research into the causes and consequences of this acidic pH of tumors is highly dependent on accurate, precise, and reproducible measurements, and these have undergone great changes in the last decade. This review focuses on the most recent advances in the in vivo measurement of tumor pH by pH-sensitive PET radiotracers, MR spectroscopy, MRI, and optical imaging.
Increased levels of both the cysteine protease, cathepsin L, and the serine protease, uPA (urokinase-type plasminogen activator), are present in solid tumors and are correlated with malignancy. uPA is released by tumor cells as an inactive single-chain proenzyme (pro-uPA) which has to be activated by proteolytic cleavage. We analyzed in detail the action of the cysteine protease, cathepsin L, on recombinant human pro-uPA. Enzymatic assays, SDS-PAGE and Western blot analysis revealed that cathepsin L is a potent activator of pro-uPA. As determined by N-terminal amino acid sequence analysis, activation of pro-uPA by cathepsin L is achieved by cleavage or the Lys158-lle159 peptide bond, a common activation site of serine proteases such as plasmin and kallikrein. Similar to cathepsin B (Kobayashi et al., J. Biol. Chem. (1991) 266, 5147-5152) cleavage of pro-uPA by cathepsin L was most effective at acidic pH (molar ratio of cathepsin L to pro-uPA of 1:2,000). Nevertheless, even at pH 7.0, pro-uPA was activated by cathepsin L, although a 10-fold higher concentration of cathepsin L was required. As tumor cells may produce both pro-uPA and cathepsin L, implications for the activation of tumor cell-derived pro-uPA by cathepsin L may be considered. Different pathways activation of pro-uPA in tumor tissues may coexist: (i) autocatalytic intrinsic activation of pro-uPA; (ii) activation by serine proteases (plasmin, kallikrein. Factor XIIa); and (iii) activation by cysteine proteases (cathepsin B and L).
The high metabolic rate of tumours often leads to acidosis and hypoxia in poorly perfused regions. Tumour cells have thus evolved the ability to function in a more acidic environment than normal cells. Key pH regulators in tumour cells include: isoforms 2, 9 and 12 of carbonic anhydrase, isoforms of anion exchangers, Na+/HCO3- co-transporters, Na+/H+ exchangers, monocarboxylate transporters and the vacuolar ATPase. Both small molecules and antibodies targeting these pH regulators are currently at various stages of clinical development. These antitumour mechanisms are not exploited by the classical cancer drugs and therefore represent a new anticancer drug discovery strategy.
Metastasis is a multistep process that culminates in the spread of cells from a primary tumor to a distant site or organs. For tumor cells to be able to metastasize, they have to locally invade through basement membrane into the lymphatic and the blood vasculatures. Eventually they extravasate from the blood and colonize in the secondary organ. This process involves multiple interactions between the tumor cells and their microenvironments. The microenvironment surrounding tumors has a significant impact on tumor development and progression. A key factor in the microenvironment is an acidic pH. The extracellular pH of solid tumors is more acidic in comparison to normal tissue as a consequence of high glycolysis and poor perfusion. It plays an important role in almost all steps of metastasis. The past decades have seen development of technologies to non-invasively measure intra- and/or extracellular pH. Most successful measurements are MR-based, and sensitivity and accuracy have dramatically improved. Quantitatively imaging the distribution of acidity helps us understand the role of the tumor microenvironment in cancer progression. The present review discusses different MR methods in measuring tumor pH along with emphasizing the importance of extracelluar tumor low pH on different steps of metastasis; more specifically focusing on epithelial-to-mesenchymal transition (EMT), and anti cancer immunity.
The hallmarks of cancer comprise six biological capabilities acquired during the multistep development of human tumors. The hallmarks constitute an organizing principle for rationalizing the complexities of neoplastic disease. They include sustaining proliferative signaling, evading growth suppressors, resisting cell death, enabling replicative immortality, inducing angiogenesis, and activating invasion and metastasis. Underlying these hallmarks are genome instability, which generates the genetic diversity that expedites their acquisition, and inflammation, which fosters multiple hallmark functions. Conceptual progress in the last decade has added two emerging hallmarks of potential generality to this list-reprogramming of energy metabolism and evading immune destruction. In addition to cancer cells, tumors exhibit another dimension of complexity: they contain a repertoire of recruited, ostensibly normal cells that contribute to the acquisition of hallmark traits by creating the "tumor microenvironment." Recognition of the widespread applicability of these concepts will increasingly affect the development of new means to treat human cancer.