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Mechanisms of Tumor Cell Necrosis
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56 Current Pharmaceutical Design, 2010, 16, 56-68
Mechanisms of Tumor Cell Necrosis
Sergey Y. Proskuryakov1 and Vladimir L. Gabai2*
1
Medical Radiology Research Center, Obninsk, Russia; 2Boston University Medical School, Boston, MA, USA
Abstract: Until recently, necrosis, unlike apoptosis, was considered as passive and unregulated form of cell death. However, during the
last decade a number of experimental data demonstrated that, except under extreme conditions, necrosis may be a well-regulated process
activated by rather specific physiological and pathological stimuli. In this review, we consider mechanisms and the role of necrosis in
tumor cells. It became recently clear that the major player in necrotic cascade is a protein kinase RIP1, which can be activated by number
of stumuli including TNF, TRAIL, and LPS, oxidative stress, or DNA damage (via poly-ADP-ribose polymerase). RIP1 kinase directly
(or indirectly via another kinase JNK) transduces signal to mitochondria and causes specific damage (mitochondrial permeability
transition). Mitochondrial collapse activates various proteases (e.g., calpains, cathepsin) and phospholipases, and eventually leads to
plasma membrane destruction, a hallmark of necrotic cell death. Necrosis, in contrast to apoptosis, usually evokes powerful inflammatory
response, which may participate in tumor regression during anticancer therapy. On the other hand, excessive spontaneous necrosis during
tumor development may lead to more aggressive tumors due to stimulatory role of necrosis-induced inflammation on their growth.
Keywords: Programmed cell death, RIP1 kinase, mitochondria, oxidative stress, inflammation, anti-cancer therapy, tumor progression.
INTRODUCTION death accompanied by a rapid efflux of cell constituents in extra-

A few decades ago the discovery of new patterns of cell death cellular space without activation of caspases or autophagy. We will
led to emergence of the concept of apoptosis. During apoptosis discuss mechanisms of tumor cell necrosis and its implication in
there were remarkably arranged morphological and biochemical anti-cancer therapy and tumor development.
events while necrosis was considered as apparently deranged (or
1. OCCURRENCE OF TUMOR CELL NECROSIS IN VITRO
accidental) form of cell death. Morphologically, necrosis is quite
AND IN VIVO
different from “classical” apoptosis. In the course of apoptosis cells
first shrink, their nuclei condense, and then they disintegrate into Instant killing of cells by extreme conditions such as severe
well-enclosed apoptotic bodies, while during necrosis cells first temperature or acids causes unregulated processes of destruction of
swell (“oncosis”), then the plasma membrane collapses, and cells cell membranes and cytosol. This led to the common assumption
are rapidly lysed. Biochemical hallmarks of apoptosis such as that when cell destruction is accompanied by a rapid disruption of
activation of specific proteases (caspases) and oligonucleosomal the plasma membrane, cytoplasmic structures, and the nucleus, it
DNA fragmentation are usually absent in necrotic cells. However, indicates that the death is passive and unregulated. However, such
improvement of methods of differentiation of apoptosis and conclusion disregarded many phenomena where necrotic cell death
necrosis revealed that there are many examples when some bio- is a regulated process activated by rather specific physiological and
chemical and morphological characteristics of both modes of cell pathological conditions. Of note, a variety of conditions that
death can be found in the same cell. This indicates that there is a activate necrosis may activate apoptosis and autophagic cell death
spectrum of suicidal programs in cells, and “classical” necrosis and as well, dependent on intensity of stimuli and cell type.
apoptosis are the extremes of the spectrum (see ref [1] for review). Among the most common conditions of activation of necrosis
Moreover, during last decade necrotic cell death has brought much are ischemia and hypoxia, which can be simulated in vitro by
more attention and now regarded by many researchers as specific deprivation of oxygen, glucose and other nutrients (Table 1). These
form of programmed cell death (PCD), type III PCD, along with hypoxic and ischemic conditions are often occurs during tumor
apoptosis (type I PCD), and autophagy (type II PCD) (see ref [1-5] growth in vivo due to inadequate vascularization. The main cause of
for review). necrosis under these conditions is obviously energy deprivation,
The most common assays for measuring necrosis in vitro is and to survive this stress cells need either increase energy
permeability of cell plasma membrane to vital dyes (like trypan production or decrease energy consumption. Since in the absence of
blue or propidium iodide), and efflux of cytosolic enzymes (lactate oxygen and/or nutrients cells have limited capability to boost ATP
dehydrogenase, creatine kinase). Reduction of tetrazolium salts generation, decreasing ATP consumption is often the only way to
(e.g., MTT assay) is also often used, but it rather measures mito- avoid necrosis. For instance, protective effect of AG1478, an
chondrial activity of cells which can be compromised not only due inhibitor of EGF receptor, from hypoxia-induced necrosis of glioma
to necrosis, but to apoptosis or autophagic cell death as well. Of cells is associated with preservation of ATP [6]. Furthermore,
note, however, is that in vitro apoptosis as well as autophagic death LKB1, serine-threonine kinase which is often mutated in tumors, is
finally also leads to plasma membrane permeabilization important for activity of AMP-activated protein kinase and survival
(“secondary necrosis”), but it does not usually occur in vivo during of glucose-deprived tumor cells [7]. Another interesting example of
apoptosis since apoptotic cells are digested by macrophages or adaptation to avoid hypoxia-induced necrosis has been recently
surrounding cells before their plasma membrane becomes described. Ginouves and co-workers found that, while acute
disrupted. Furthermore, secondary necrosis may also occur due to hypoxia (1% 02 ) caused stabilization of hypoxia-induced factors
mitotic catastrophe as a consequence irreversible DNA damage. In (HIF1 and HIF2), under chronic hypoxia (more than 3 days)
this review the term “necrosis” will be mostly attributed to cell these factors are degraded and this degradation is crucial for
protection of cells from hypoxia-induced necrosis [8].
*Address correspondence to this author at the Dept Biochemistry, Boston Several components of immune system are capable of inducing
University Medical School, 715 Albany St, Boston, MA 02118, USA; necrosis, among them TNF, FAS, and TRAIL. Activation of FAS
E-mail: gabai@bu.edu receptors with anti-FAS ligand is also able to provoke necrosis
1381-6128/10 $55.00+.00 © 2010 Bentham Science Publishers Ltd.

Mechanisms of Tumor Cell Necrosis Current Pharmaceutical Design, 2010, Vol. 16, No. 1 57
Table 1. Activators and Inhibitors of Necrosis
Activators Cells Inhibitors References
Ischemia Various cells Necrostatin-1 [77]

Hypoxia Glioma AG1478 [6]
+
MPP PC-12 - [61]
Rotenone PC-12 - [61]
Antimycin A Rat-1 - [63]
Glutamate HT-22 Necrostatin-1 [174]
TNF L929, MEF BHA [29]

TNF+zVAD U937, THP-1 GA [27]
FAS T-cells - [175]
TRAIL (pH6.5) HT-29 zVAD [26]
LPS Macrophages - [176]
Anti-CD47 B-lymphocytes, CLL TPCK [9]
IL1+IFN L929 SB203580; sc-514 [56]
ZVAD L929 Rapamycin [129]
Ceramide Jurkat, 3T3 - [28]
ROS MEF SP600125 [38]

RNS U937, MEF Rotenone [39,55]
Photodynamic therapy HeLa MitoQ [177]
As203 HeLa NAC, PARP inhibitors [81]
Tamoxifen MCF7, MDA-MB231 Estrogens, PD98059, BA [13,14]
MNNG MEF PARP inhibitors, SP600125 [10,30]
Imatinib K562 TPCK [18]
Honokiol HL-60, MCF7, HEK293 CsA [16]
Shikonin MCF7, HEK293 Necrostatin-1 [17]
Resveratrol MCF7 - [15]

Abbreviations and Notes: AG1478 – Inhibitor of EGF receptor; MPP+- 1-Methyl-4-Phenylpyridinium; BHA- butylated hydroxyanisole; GA – geldanamycin; zVAD – caspase
inhibitor; TPCK – serine protease inhibitor; SB203580 – p38 MAPK inhibitor; NAC- N-acetylcystein; sc514 – NFkB inhibitor; SP600125 – JNK inhibitor; MNNG – N-methyl- N’-
nitro- N’ –nitrosoguanidine; PD98059 – ERK1/2 inhibitor; BA – bonkrekic acid; CsA – cyclosporin A.
along with apoptosis, but ligation of CD47 receptors in lymphoid which is apparently associated with inability to stabilize tumor
cells induces almost exclusively necrosis [9](Table 1). suppressor p53 [19].
Various treatments used for anticancer therapy also may cause There are several key events during cancer initiation and
necrosis of tumor cells. Among them are DNA-alkylating drugs progression, and suppression of apoptosis is considered among the
such as nitrogen mustard or MNNG [10], arsenic trioxide [11], most important [20]. Indeed, many oncogenes can activate
photodynamic therapy [12], and tamoxifen [13,14] (Table 1). apoptotic program, and cancer cells often disable tumor suppressors
Resveratrol [15] and some components of traditional Chinese implicated in apoptosis such as p53 or Bax, or overexpress
herbal medicine such as honokiol [16] and shikonin [17] are also apoptosis inhibitors such as Bcl-2, Bcl-x, or survivin. Furthermore,
capable to induce necrosis of tumor cells (Table 1). Importantly, during in vivo growth, tumor cells may be subjected to ischemia
resistance to apoptosis does not prevent killing of tumor cells by and attack by cytotoxins from immune system, and disabling
these agents. Accordingly, caspase inhibitors failed to prevent apoptosis can protect tumor cells from these adverse conditions and
necrosis of BCR-ABL-positive human leukemic cells treated with a promote tumorigenesis. The above findings that apoptosis-resistant
novel anticancer drug (a protein kinase inhibitor) imatinib cells are still vulnerable to therapy-induced necrosis may have
(Gleevec) [18](Table 1). Interestingly, genotoxic stresses such as clinical implications. Indeed, Dinnen et al found that p53 C-
actinomycin D, UV radiation, or cisplatin caused apoptosis in terminal 22-aa peptide (aa 361-382) linked to truncated 17-aa
young fibroblasts, but necrosis in old (senescent) human fibroblasts, peptide from Drosophila antennapedia homeobox domain (to
58 Current Pharmaceutical Design, 2010, Vol. 16, No. 1 Gabai et al.
facilitate cellular uptake) can induce selective necrosis in p53-

mutant prostate tumor cells [21].
Recent studies from E. White’s lab also demonstrated important
role of autophagy in protection of tumor cells from ischemic
conditions in vitro and in vivo during tumor development. It was
found that apoptosis-resistant cells die via necrosis only when
autophagy is suppressed (e.g., by activation of AKT) [22] (see also
section 4 below).
2. SIGNALING PATHWAYS AND MODULATORS OF

NECROSIS
2.1. Receptors and Protein Kinase Cascades

Among the best studied examples of necrotic cell death is that
activated by engagement of TNF, FAS, or TRAIL receptors.
Normally, activation of these receptors leads to apoptosis via
FADD/Caspase-8 signalling; however, when apoptosis is inhibited
(genetically or by caspase inhibitors), alternative necrotic pathway
is initiated. Critical role in this receptor-activated necrosis is played
by protein kinase RIP1, which was demonstrated in several studies,
including its knockdown [23] (Fig. 1). Along with activation of
necrosis, RIP1 is involved in anti-apoptotic signalling via activation
of NF-kB; however, caspase-8 can cleave RIP1 thus promoting
apoptosis and preventing necrosis [24].
Activation of RIP1, beside death receptors, can be caused by
LPS via toll-like receptors (TLR) TLR3 and TLR4 [25]. As in the
case of death receptor activation, LPS normally induce apoptosis in
macrophages when NF-kB dependent anti-apoptotic pathway is
inhibited, but when apoptosis is suppressed, RIP1-dependent
necrosis is initiated. Therefore, switching between apoptosis and
necrosis during activation of death and TLR receptors generally
depends on activity of caspase-8: when this caspase is disabled,
RIP1-dependent necrosis is activated. However, there is interesting Fig. (1). Key role of RIP1 kinase in necrotic signaling.
exception when at acidic pH 6.5 (which can often occur in tumors Diverse stimuli activate signaling pathways intercepting at the level of RIP1
due to high glycolysis), TRAIL induce RIP1-dependent necrosis of kinase.
tumor cells which was accompanied by caspase-8 and caspase-3 See text for further explanation.
activation and prevented, rather than accelerated, by caspase
inhibitors [26]. binding domain; upon activation by DNA breaks it attaches
What are downstream targets of RIP1 kinase? In human oligomers of ADP-ribose to itself and some other nuclear proteins.
myelomonocytic U937 and TNP-1 cells treated with TNF or FAS in Excessive activation of PARP, for example, as a result of profound
the presence of caspase inhibitor, necrosis was dependent on RIP1- induction of DNA breaks, was believed to be a cause of cell
induced inhibition of ATP/ADP translocator in mitochondria , necrosis due to ATP depletion [32,33] which is resulted from use of
which leads to decrease in ATP levels [27]. RIP1-deficient cells ATP for synthesis of the PARP substrate NAD+(see also section 2.3
also defective in ceramide accumulation, and prevention of its below). PARP inhibition (e.g., by 3-aminobenzamide and nicotina-
accumulation protected them from TNF-induced necrosis [28]. mide) suppressed cell necrosis [32] or switched it to apoptosis,
Recent study of Kim et al. [29] on L929 murine fibrosarcoma cells which was associated a marked increase in caspase activity [34,35].
demonstrated that RIP1 is also essential for recruitment of NADPH During apoptosis, PARP, similar to RIP1, is normally inactivated
oxidase Nox1 in complex with TRADD (Fig. 1). Nox1 is by caspase-specific cleavage, forming an 89-kDa fragment, a
responsible for generation of ROS (in particular, superoxide O2-), biochemical hallmark of apoptosis. If this mechanism of PARP
and ROS is essential mediators of TNF-induced necrotic cell death inactivation is not operational, for example, in a PARP mutant
since anti-oxidant BHA almost completely prevented cell death. resistant to caspase cleavage, cells become more sensitive to
Another important player in RIP1/Nox1 – dependent necrosis is necrosis induced by UV radiation or TNF [36,37]. In these cells
MAP kinase JNK. This kinase is also involved in apoptotic expressing mutant PARP, as well as in their wild-type counterpart,
signalling under many stressful conditions, and, in some cases, its inhibition of PARP activity reduced necrosis and increased
transient activation may play role in mitogenic response and cell apoptosis [37]. Hence, proteolytic or pharmacological inactivation
survival. However, in case of TNF-induced necrosis, sustained of PARP is one of the ways to prevent cell elimination through
activation of JNK appears to be essential for cell death, since JNK necrotic pathway. Xu et al found that MNNG-induced necrosis,
inhibition, similar to inhibition of ROS production, protected cell which depends on PARP activity, was significantly suppressed in
from necrosis [29] (Fig. 1). It is quite possible that besides ROS, RIP1 and JNK1 knockout cells [30]. Since poly-ADP ribosylation
activation of JNK in TNF-induced necrosis is also mediated by was not inhibited in these knockout cells, it places RIP1 and JNK
ceramide [28], a known JNK stimulator. downstream of PARP-1 (Fig. 1). Surprisingly, even necrosis
Unexpected link between genotoxic stress, poly (ADP-ribose) induced by hydrogen peroxide and reactive nitrogen species (RNS)
polymerase (PARP-1) and RIP1-JNK necrotic cascade has been also depends on activation of TRAF2-RIP1-JNK signaling cascade
found by Xu et al. [30]. DNA alkylating agents such as MNNG in [38,39]. Therefore, besides receptor-activated necrosis, RIP1-JNK
high concentrations is able to induce necrosis via PARP-1 signaling appears to be important for necrosis induced by some
stimulation [10,31]. PARP-1 is a nuclear enzyme containing a Zn- genotoxic agents, ROS and RNS (Fig. 1).
JNK and another related stress kinase, p38 are also involved in Peroxinitrite is a highly active nitrogen compound that is formed in
necrotic death of cells following transient energy deprivation in organisms and it is often used as a model simulating action of
vitro, or ischemia/reperfusion of different organs in vivo [40]. cytokines on effector cells. Peroxinitrite formation is caused by
Ischemia/reperfusion-induced necrosis was also inhibited by expression of inducible NO synthase (iNOS) and ROS generation
expression of dominant-negative form of Rac, an upstream com- as a result of reaction between NO and superoxide anione. Pro-
ponent of stress-signaling cascade, although this protective effect necrotic effect of peroxinitrite in U937 is mediated by hydrogen
may be also related to inhibition of ROS production [41], similar to perioxide apparently generated by mitochondria, since mito-
effect of dominant-negative RAC on TNF-induced necrosis [29]. chondrial inhibitor rotenone prevented necrosis [55]. Recently,
On the other hand, activation of MAP kinase ERK and AKT, which necrosis of L929 tumor cells caused by prolonged incubation with
protect cells from stress-induced apoptosis, can protect against IL-1 and IFN was shown to depend on expression of iNOS and
necrotic death as well. For example, AKT overexpression reduced NO production, which, in turn, was dependent on p38 MAP kinase
necrotic zone formation in ischemic myocardium [42]. Accor- and NF-kB (IKK) activities [56].
dingly, ischemia/reperfusion-induced ERK activation seems to be In most cases, antioxidants suppress both apoptotic and necrotic
protective against necrosis, since its inhibition aggravated necrosis cell destruction. It seems that oxidative stress induces an apoptotic
of myogenic cells in vitro [43] and myocardial infarction in vivo response when cells can maintain their reducing capacity against
[44]. Therefore, it seems that proapoptotic (JNK, p38) and anti- ROS, whereas necrosis is triggered when this reducing homeostasis
apoptotic kinases (AKT, ERK) play a similar role in necrosis. Of is disturbed (e.g., by excess of ROS or damage of natural anti-
note, tumor cells often have higher activity of AKT and ERK oxidative systems).
signaling cascades which can make them potentially more resistant
not only to apoptosis, but necrosis as well. On the other hand, Akt 2.3. ATP and Mitochondria
activity blocks autophagy, an important tumor cell survival mecha- Now it seems obvious that mitochondria play a crucial role in
nism under chronic ischemic conditions. As a result, apoptosis- determination of cell fate under stresses. First, as a source of ATP,
resistant cells with overexpression of active Akt are less resistant to mitochondria chose between ATP-dependent or -independent
ischemia than cells without such overexpression [22] (see also programs. Second, they generate ROS that control form of cell
section 4). suicide (see above), and finally, as a source of tanathogenic (death-
2.2. Reactive Oxygen and Nitrogene Species promoting) factors, mitochondria initiate or amplify the caspase-
dependent apoptotic program (mainly through efflux cytochrome c)
Necrotic cell death is almost always accompanied by generation or activate directly the execution phase (through efflux of apoptosis
of ROS, and various anti-oxidants can prevent necrosis or switch it induction factor, AIF, and other factors).
to apoptosis. Increased production of ROS and RNS can be caused
by macrophages during immunological response, by mitochondria, Apparently, maintenance of certain levels of ATP is required
and by some other mechanisms. Hydrogen peroxide, a component for execution of apoptotic programs. ATP or its derivate, dATP, is a
of ROS, is often used as a model reagent since it is produced as a cofactor of apoptosome [57], a high-molecular-weight complex
factor of immune defense and during various stresses. It can cause consisting of APAF-1 and caspase-9 [58], which activates a major
both apoptosis and necrosis of cells [34, 45] which can be execution caspase, caspase-3. Besides apoptosome, ATP also seems
prevented by the antioxidants glutathione , N-acetylcystein (NAC), necessary at other stages of the apoptotic program [59]. Generally,
BHA and others. When applied together with some antitumor drugs if the amount of ATP drops below some critical levels, this either
(VP-16, doxorubicin, cisplatin, and AraC) subtoxic doses of H2O2 can switch apoptotic cell death to necrotic (e.g., when cells exposed
can switch cell suicide to necrosis [46,47]. This effect was probably to genotoxic stress causing profound PARP activation, see section
associated with specific signaling role of H2O2 rather than with 2.1) or may cause necrosis by itself. For instance, in HeLa cells,
inhibition of caspases or PARP activation (see above) since much inhibition of glycolysis and respiration leads to more than 97%
higher concentration of H2O2 was necessary for the latter effect [34, decrease in ATP and, if ATP levels restored after 3 hr, cells die via
46]. A decrease in cellular content of glutathione can also switch a apoptosis, but more prolonged energy deprivation evoked necrotic
form of cell death induced by ROS. For example, apoptosis of cell death [60].
U937 tumor cells induced by Cd2+, cisplatin, and melphalan was Mitochondrial inhibitors alone can cause necrosis in some cells.
switched to necrosis when glutathione synthesis was inhibited [48, Inhibitors of complex I of respiratory chain, such as rotenone, 1-
49]. Interestingly, ROS-induced necrosis can also be modulated by methyl-4-phenylpyridium, or 6-hydroxytriptamine, which simulate
cell transformation. Transformation of 3T3 cells by SV40-T antigen cell loss during Parkinson’s disease, caused necrosis of PC12
promoted menadione-induced necrosis, which can be inhibited by neuroblastoma cells [61]. Furthermore, inhibitors of complex II, 3-
blocking FAS receptors, although inhibitors of caspases were nitropropionic acid, or complex III, antimycin A, also induced
ineffective [50]. necrosis [62,63]. Mitochondrial inhibitors, however, usually do not
Along with ROS, another mediator of various pathophysiolo- affect viability of tumor cells with high level of glycolysis that are
gical processes is nitrogen oxide (NO). Because nitrosylation/ capable of maintaining ATP levels without any respiration (see,
denitrosylation reaction is involved in regulation of caspase-3, a e.g., [64, 65]). Of note, drastic ATP depletion (below 3% to 5% of
key apoptotic caspase, NO may inhibit apoptosis directly through initial) for many hours resulted from hypoxia or starvation is not
caspase-3 nitrosylation [51], although other mechanisms of toxic for some cells (e.g., fibroblasts), whereas other cells (e.g.,
inhibition of apoptosis upstream caspase-3 may also exist [52,53]. neuronal and cardiac cells) rapidly die via necrosis (see Ref. [65]
The inhibition of apoptotic pathway may be the reason why NO can for review). The reason for such different sensitivity of cells to ATP
switch apoptosis to necrosis upon treatment with staurosporine, depletion is not clear, but may be associated with much more severe
ceramide, FAS, and retinoids [52]. On the other hand, FAS-induced ionic imbalance (in particular, Ca2+ imbalance) in sensitive cells.
denitrosylation of caspase-3 by thioredoxin-2 is required for On the other hand, necrosis can be induced in the cells with a
caspase-3 activation and promotion of apoptosis [54]. normal amount of ATP (e.g., TNF-induced necrosis of L929 fibro-
sarcoma cells, see section 2.1), or peroxynitrite-induced necrosis of
NO can also bind to iron of heme-containing complexes of U937 cells [66]. This indicates that, although the ATP levels may
respiratory chain and inactivate them, potentially leading to mito- control mode of cell death, there are other factors that contribute in
chondrial damage (see below). A deleterious effect of exogenous final outcome.
NO can be increased by Fe2+ ions and by blocking GSH synthesis
while NAC and SH-group donors can protect against NO. One such factor may be ROS produced by the mitochondrial
respiratory chain, and this ROS generation may trigger a necrotic
program, as discussed above. It was hypothesized that when cellular apoptotic, but also necrotic death. They delay or prevent necrosis
anti-oxidative defense is limited, ROS caused oxidation of the key evoked, for instance, by chemical anoxia [84], myocardial ischemia
molecules and release of executor proteases, lipases, and nucleases [85], hypoxia [86], TNF [28], or arsenic trioxide [11]. However, not
from mitochondria [67]. The emergence of such dangerous all necrotic programs are suppressed by proteins of the Bcl-2
mitochondria triggers the cell’s protective response in the form of family, for example, necrosis caused by peroxinitrite [87], or the
autophagy with participation of caspases [67,68]. This hypothesis mitochondrial uncoupler 3-acetylpyridine [88]. A balance between
may explain why in some cells inhibition of caspases, while the necrotic and apoptotic cell responses may also depend on a
inhibiting TNF-induced apoptosis, may trigger necrotic cell death. balance between pro- and antiapoptotic members of the Bcl-2
Indeed, TNF may activate mitochondrial ROS generation, and such family. For instance, the anti-necrotic effect of chronic hypergly-
dangerous ROS-producing mitochondria are normally eliminated cemia consists in activation of Bcl-2 expression and phos-
by caspase-dependent autophagy [67]. However, when caspases are phorylation of the proapoptotic protein Bad [89]. Increased expres-
inhibited, these mitochondria may trigger necrotic death of a whole sion of Bax in glioblastoma stimulated apoptosis, but coexpression
cell. Interestingly, because some viruses encode caspase inhibitors of Bcl-XL, surprisingly, switched cell death to necrosis [90]. Phos-
to avoid apoptosis of infected cells, the ability to trigger necrosis phorylation status of Bcl-2 also may play a role in cell necrosis. For
when caspases are inhibited may be an important part of the cellular instance, suppression of inhibitory phosphorylation of Bcl-2 by
antiviral defence [69]. Twist-1 protected from necrosis caused by anti-cancer drugs cis-
Being the source of apoptogenic factors (cytochrome c, platinum, VP-16, and daunorubicin [91].
Smac/Diablo, AIF), in addition to ROS, mitochondria can be the A recent work from Susin’s lab unraveled a role of mito-
source of pro-necrotic factors as well. Under some conditions (e.g., chondria in MNNG-induced, PARP-1 dependent necrosis [31] (see
high Ca2+, oxidative stress) mitochondria undergo drastic changes section 2.1). It was found that Bax, but not Bak knockout
accompanied by deenergization of the inner membrane, swelling, completely prevented necrosis; furthermore, Bcl-2 overexpression
and permeabilization, a process called mitochondrial permeability also suppressed necrosis. Activation of BAX was mediated by
transition (MPT). This is usually an irreversible process leading to cytosolic protease calpain (see section 2.6 below) which leads to
mitochondrial “death” (mitochondrial apoptosis or “mitoptosis” AIF release and necrosis [31].
[60]). It was suggested that MPT may be inductor of necrotic cell A protein from the Bcl-2 family, BNIP3, which causes mainly
death through release of some mitochondrial factors (e.g., Ca2+, necrotic cell death, has been discovered [92]. Pro-necrotic functions
proteases, lipases) [70,71]. Indeed, inhibitors of MPT such as CsA, of this protein in transfected cells are manifested by earlier plasma
or bonkrekic acid may protect from necrosis caused by oxidative membrane permeabilization, cytoplasm vacuolization, and
stress, hypoxia–reoxygenation in vitro [72], or ischemia– autophagy of mitochondria. These morphological changes were
reperfusion in vivo [73], or some other stimuli (Table 1). accompanied by mitochondrial depolarization and ROS generation
Cyclophilin D (CypD) is a mitochondrial matrix protein and and were blocked by inhibitors of MPT CsA and bonkrekic acid
indispensable component of MPT; recent studies showed that [92]. Interestingly, integration of BNIP3 into the mitochondrial
knockout of CypD gene induce resistance to necrosis induced by membrane upon necrosis of neurons was prevented by necrostatin-1
ROS and Ca2+ overload in vitro, as well as ischemia-reperfusion of [82], which also may explain inhibitory effect of necrostatin on
heart and brain in vivo [74, 75]. Accordingly, knockout of CypD MPT and ischemic necrosis in cardiac cells (see section 2.3 above).
and specific inhibition of cyclophilin D with compound Debio-025 Indeed, expression of dominant-negative form of BNIP3 in
also protected mice from necrosis caused by muscular dystrophy myocardium reduced ischemic injury (e.g., release of creatine
[76]. Interestingly, necrostatin-1, a small molecule inhibitor of kinase) [93]. BNIP3 accumulation is activated by hypoxia via HIF1
necrosis (Table 1), but not apoptosis which was recently found by and it is highly expressed in some tumors, including those of breast,
Yaun’s lab [77], also prevented MPT in mitochondria [78,79]. lung, and cervix, although in colorectal and pancreatic cancer it is
Specific cellular target of necrostatin-1, however, appears to be often epigenetically silenced (see [94] for review). Apparently, at
RIP1 kinase (see section 2.1) rather than mitochondria [80], and least in some tumors, suppression of BNIP3 provide them with
protection of mitochondria is obviously a secondary effect of RIP1 growth advantage under conditions of hypoxia/ischemia that occurs
inhibition (see section 2.1). during tumor development [95].
Despite its name, apoptosis-inducing factor (AIF), when Thus, the main anti-apoptotic and anti-necrotic effect of Bcl-
released from mitochondria, may also play a crucial role in 2/Bcl-xL proteins is believed to consist in preservation of mito-
necrosis, which was shown for arsenic trioxide-induced necrosis of chondrial integrity (i.e., prevention of MPT, efflux of cytochrome c,
human cervical cancer cells [81], and MNNG-induced necrosis of and other proapoptotic/pronecrotic factors).
fibroblasts [3,31]. Release of AIF was also blocked by necrostatin-1
[82]. 2.5. Heat Shock Proteins
Recent study from D. Green’s lab showed that mitochondria Heat shock proteins (Hsps) are other important regulators of
can preserve their integrity and sustain cell survival despite cell death, and, along with other prosurvival proteins such as Bcl-2,
complete loss of cytochrome c provided caspases are blocked and they are often overexpressed in tumor cells [96-98]. The most
autophagy is activated. Important role in such survival is played by studied of them are Hsp70 and Hsp27, and they were first described
GAPDH, which both generate ATP via glycolysis and stimulates as inhibitors of apoptosis caused by diverse stimuli (see [97-99] for
transcription of Atg12, a protein involved in autophagy [83]. review). However, later it was established that Hsps can protect
cells not only from apoptosis, but also from autophagic cell death
Therefore, mitochondria may be the source of three relatively
[100], senescence [101, 102] and, apparently, mitotic catastrophe
independent signals that trigger or switch cell death pathways:
[103]. Their overexpression also protect cells from necrosis caused
ATP, ROS, and apoptogenic/necrogenic factors such as cytochrome
by heat shock [104], oxidative stress [105], nitric oxide [106], and
c and AIF. The final outcome of cell suicide is apparently depen-
ischemia/reperfusion [65]. For instance, after myocardial ischemia/
dent on interplay between these factors.
reperfusion, transgenic mice overexpressing Hsp70 in the heart had
2.4. Proteins of the Bcl-2 Family a smaller infarct zone, a lower level of creatine kinase (indicator of
necrosis) in blood plasma, and better recovery of mechanical
Proteins of the Bcl-2 family play a very significant role in the function [65,107]. Small Hsps, Hsp27 and its homolog B-
determination of cell sensitivity to lethal signals. Antiapoptotic crystallin, can also protect cardiomyocytes from ischemia-induced
members of this family (Bcl-2, Bcl-X L, etc.) can inhibit not only necrosis in vitro and in vivo [108,109]. The protective effect of
Hsp70 in myocardial ischemia was not associated with preservation depletion in CD95-stimulated Jurkat cells was suppressed by the
of ATP level during ischemia, but ATP recovery in the myocardium pan-caspase inhibitor zVAD.fmk [127]. This inhibitor (but not z-
of Hsp70-expressing animals was faster and higher than in control DEVD.fmk, an inhibitor of caspase-3) also reduced TNF-induced
[110]. These data may indicate that Hsp70 preserve mitochondrial necrosis of fibrosarcoma cells [128], as well as TRAIL-induced
functions during ischemia/reperfusion and/or accelerate the necrosis of human colon cancer cells at low pH [26]. Of note,
recovery of these functions. Furthermore, the protective effect of however, that at least in some cells (e.g., L929), zVAD.fmk is able
Hsp70 against NO-induced necrosis in human -cells was not to suppress cathepsine B activity [129].
associated with suppression of lipid peroxidation, but also with During the past years, a number of data emerged demonstrating
rescue of mitochondrial functions (tetrazolium reduction) [111]. wide occurrence of caspase-independent programmed cell death,
However, since neither Hsp70 nor Hsp27 are localized to both apoptotic and necrotic [130, 131]. For instance, TNF-induced
mitochondria, it seems unlikely that their protective action is cell death of hepatocytes and some tumor cells apparently requires
associated with direct effect on mitochondrial structure. Probably, lysosomal cysteine protease cathepsine B [132]. However, in L929
these chaperones suppress signal transduction pathways leading to cells, zVAD.fmk-induced necrosis was aggravated by cathepsin
mitochondrial damage and cell death. These pathways may include inhibitors [129]. Interestingly, cathepsine B expression appears to
the stress kinases JNK and p38 (see section 2.1.). Indeed, activity of be higher in immortalized and transformed cells making them more
these kinases was markedly elevated during ischemia/reperfusion, sensitive to TNF-induced cell death [133].
and their inhibition suppressed necrosis in vitro and in vivo [112].
Because activation of JNK and p38 under in vitro “ischemia” of Another cysteine protease, Ca2+-dependent calpain, may
myogenic cells was reduced in Hsp70-expressing cells [40], these participate in ischemia-induced cell death of hepatocytes and
kinases may be the targets of anti-necrotic effect of Hsp70 in the neurons after ischemia/reperfusion [134,135]. Mitochondrial
myocardium. Interestingly, protection of the kidney from ischemia/ calpain I was shown to be able to cleave AIF [136], which may be
reperfusion by ischemic preconditioning was also associated with critical for release of AIF from mitochondria and neuronal cell
stress kinase suppression, although in this case it was Hsp27 rather death [135]. Calpain also mediates activation of BAX and release of
than Hsp70 that was accumulated in the preconditioned kidney AIF from mitochondria in MNNG-induced necrosis [31], whereas
[113]. Although role of Hsps in protection of tumor cells from in arsenic trioxide-induced necrosis calpain mediates cleavage of
ischemia-induced necrosis occurring during tumor development has nuclear Bax [137].
not been assessed directly, it seems quite probable since higher Finally, some as yet unidentified serine proteases can
levels of Hsp27 or Hsp70 promote growth of tumors, while participate in TNF-induced necrosis of L929 cells [138], necrosis of
downregulation of these proteins suppresses it [114-118]. kidney cells induced by “chemical hypoxia” [139], CD47-induced
Therefore, the molecular chaperones Hsp70 and Hsp27, along necrosis of lymphoid cells [9], or Gleevec-induced necrosis of
with proteins of the Bcl-2 family, are powerful inhibitors of BCR-ABL-positive leukemic cells [18]. One of such serine
necrosis. It seems that, although they have quite different mecha- proteases may be mitochondrial Omi/HtrA2, which is released from
nism of action, the main targets of their protective effect are mitochondria upon Gleevec treatment [18].
mitochondria, either directly (in case of Bcl-2/Bcl-xL) or indirectly Therefore, the role of caspases, key executor caspases in
(in case of Hsp70/Hsp27). apoptosis, is more diverse in necrosis. Their inhibition may either
suppress or activate necrosis depending on cell line and stimuli. It is
2.6. Proteases, Nucleases, and Phospholipases probable that switching from apoptosis to necrosis in the presence
Cysteine proteases of the caspase family perform crucial of caspase inhibitors, at least in some cases, may be associated with
functions in suicide elimination of cells: transduction of lethal ATP depletion due to PARP1 activation (see section 2.1). If caspase
signal via cascade of caspases and a final destruction of various inhibition prevents caspase-dependent PARP inactivation, it may
protein targets (PARP, lamins, cytoskeletal proteins) [119]. In many cause ATP depletion, blockade of ATP-dependent apoptosis, and
models it is the only way of execution of apoptosis, because in the triggering of necrosis. At present, however, little is known how
presence of endogenous or exogenous caspase inhibitors, or in the caspases participates in necrosis. In most cases, however, caspases
absence of caspase expression, suicidal programs either completely are dispensable for necrosis, and other proteases such as cathepsine,
blocked or, more often, switched to a necrotic pathway. As we calpain, and serine proteases are involved in execution of this form
discussed in section 2.1, inhibition of caspases does not prevent of cell death.
TNF-induced death but rather switches it from apoptosis to Along with proteolysis, necrosis is also accompanied by
necrosis. Accordingly, caspase inhibitors switched to necrosis cell degradation of DNA. Degradation of DNA during necrosis usually
death caused by many anticancer treatments such as irradiation, occurs randomly, forming a “smear” pattern on agarose gels, while
camphotechine, etoposide, or dexametasone, or inducers of MPT apoptotic DNA fragmentation occurs to oligonucleosome fragments
[120-122]. LCC human carcinoma cells deficient in caspases died forming a remarkable “ladder” pattern on the gels. The main
via necrosis in the presence of a zinc chelator, while caspase- apoptotic nuclease is CAD (caspase-activated DNase), whereas
expressing cells died via apoptosis [123]. Furthermore, in mice caspase-independent DNase I and II are probably implicated in
without Apaf-1 or caspase-3/caspase-9, apoptotic cell death during necrosis. For instance, an increase in DNase I-like endonuclease
development switched to necrotic [124]. There are some models activity was observed in the kidney cortex after ischemia/
where caspase inhibition not only prevented apoptosis but also reperfusion [140], and activation of DNase II was found in the
severely aggravated necrosis. In L929 fibrosarcoma cells, inhibition necrotic hippocampus after global ischemia [141]. However, the
of caspases increased the cell’s sensitivity to TNF-induced necrosis mechanisms of activation of these nucleases are presently not
by a factor of 1000 [125]. These data indicate that caspases may known. Obviously, DNA damage during necrosis is also mediated
also play an antinecrotic role consisting of elimination of “harmful” by AIF.
mitochondria that produce high level of ROS, and if such Activation of some phospholipases during necrosis, especially
mitochondrial killing fails, necrosis is triggered (see section 2.3). cytosolic Ca2+-dependent phospholipase A2 (cPLA2), has been also
Additionally, caspase inactivation promote autophagy-mediated demonstrated (see [142] for review). Activity of cPLA2 was
degradation of catalase which causes ROS generation and cell death increased in hyppocampal slices immediately following exposure to
[126]. ischemic conditions, and this enhancement lasted for at least 24 h;
However, in some circumstances the execution of the necrotic furthermore, pharmacological blockade of cPLA2 (by bromo-
program requires caspase activation. Necrotic death caused by ATP phenacyl bromide or AACOCF3) prevented neuronal death [143].
Likewise, TNF-induced necrosis of MCF7 cells was suppressed by necessary. Indeed, there are indications that choice of program of
cPLA2 inhibitors [144]. Knockdown of one of isoforms of cPLA2, autodestruction occurs before initiation of the irreversible phase of
Ca2+- independent cPLA2 also delayed hypoxia-induced necrosis cell response to a lethal signal. The hallmark of apoptosis, exter-
of neuronal cells [145]. However, in peroxynitrite-induced necrosis nalization of phosphatidylserine (PS), that designates a cell with an
of U937 cells, cPLA2 apparently plays a protective role [55]. In “eat me” message, is the earliest feature of apoptosis triggering
contrast to necrosis, cPLA2 activity was dispensable for TNF- [147]. However, in necrotic cells this feature is usually registered
induced apoptosis of HeLa cells; moreover, during apoptosis after plasma membrane destruction; therefore, necrotizing cells are
cPLA2 underwent caspase-dependent cleavage and inactivation not recognized by phagocytes and cannot be digested until their
[146]. Such inactivation of cPLA2 during apoptosis may represent a intracellular contents are spilled into the extracellular space [148].
mechanism to avoid the inflammatory response against apoptotic In some cases, however, appearance of PS on necrotic cells is
cells that may be evoked by products of phospholipid hydrolysis. an early and sufficient event for phagocytosis, which occurs even
before perforation of plasma membrane. Effective absorption of
2.7. Molecular Scenario of Necrotic Cell Death
necrotic cells with inhibited caspases and not exposing PS suggest
The data described in the previous sections suggest a possible that, in addition to PS, other ligands of the “eat me” type should
molecular scenario of tumor cell necrosis. There are several exist, among them are vitronectin receptors [149, 150]. Interes-
receptors implicated in triggering necrosis; among them are TNF tingly, in artificially mixed population of apoptotic and necrotic
receptors and other receptors of this family (FAS, TRAIL), and cells, macrophages preferred the necrotic cells [150]. One of the
TLR. Another important sensor is DNA: its damage may be reasons for this preference may be abundant histidine-rich plasma
induced either directly (e.g., by radiation or anticancer drugs) or protein (HRG) which selectively recognizes necrotic, but not
indirectly through oxidative stress (e.g., upon ischemia/reperfusion apoptotic cells, and enhance their phagocytosis [151]. Additionally,
or other ROS generating treatments). Massive DNA breaks may apoptotic and necrotic cells induce different cell signaling events in
cause activation of PARP, depleting its substrate NAD+ and, bone marrow macrophages: apoptotic cells suppress ERK1/2, but
subsequently, ATP, which may lead to necrosis due to energy activate JNK and p38 kinases, while necrotic cells activate ERK1/2,
deficiency. Stimulation of the receptors, oxidative stress, and DNA but have no effects on JNK and p38 [152]. Recent scanning
damage are powerful activators of RIP1 and JNK, which leads to electron microscopy study by Krysko et al. [153] also revealed
mitochondrial damage. Indeed, mitochondria, besides their role in difference in internalization mechanisms of apoptotic and necrotic
ATP generation along with glycolysis, obviously play the key role cells by macrophages: apoptotic bodies were taken up by macro-
in determination of a pathway of cell suicide. Mitochondria are phages with formation of tight fitting phagosomes, whereas necrotic
powerful sources of tanathogenic factors such as cytochrome c, cells were internalized by macro-pinocytotic mechanism involving
AIF, and ROS, and they are the main targets of cell survival system formation of multiple ruffles directed toward necrotic debris.
(e.g., proteins of the Bcl-2 family) The amount of ATP may be the
Depending on molecular signals from necrotic cells (which are
essential factor that determines the choice of the cell suicide
alien to surrounding cells), diverse type of these cells (neutrophils,
pathway, but there are obviously other important (but yet unknown)
macrophages, and others) become involved in the immune
factors.
response. It was found that necrotic cells are more efficient than
Finally, the last stage of necrotic destruction is the activation of apoptotic cells in their capacity to stimulate the antigen-presenting
proteases. In several models of necrosis, this destruction is executed cells (APC) and T-cell response [154]. On the other hand, apoptotic
by caspases, but in most cases inhibition of caspases during stresses cells induced in APC the secretion of cytokines that inhibit Th1
may trigger necrosis rather than suppress it. This indicates that response [155]. Necrotizing tumor cells also potentiate maturation
caspase activity is sometimes necessary, paradoxically, for protec- of dendritic cells and optimal presentation of tumor antigens [154].
tion of cells from stresses, possibly through caspase-mediated These data indicate that a much more robust immune response is
elimination of ROS-generating mitochondria. Among proteases evoked during necrosis than from apoptosis. This may be
probably involved in necrotic digestion are calpains, cathepsins, physiologically important upon some dangerous situations such as
and serine proteases, but their cellular targets in necrotic cell viral or bacterial infection, trauma, or abnormal (transformed) cells,
destruction are yet to be elucidated. when strong stimuli produced by necrosis are required for
There are many questions, however, remain to be answered. mobilization of all cell defense forces (dendritic cells, monocytes,
Among them: are there any physiological stimuli activating exclu- and neutrophils). Accordingly, activation of necrosis of colon tumor
sively necrosis in mammalian cells; what determines choice of in vivo by depletion of cytochrome c, a critical component of apop-
suicidal pathway by mitochondria; how mitochondrial collapse is tosis, markedly decreased their tumorigenic ability [156].
translated to collapse of plasma membrane. Importantly, this decreased tumorigenicity was dependent on host
immune system, since in immunodeficient animals the cytochrome
3. NECROTIC DEATH AS A MODULATOR OF IMMUNE c – depleted cells grew as fast as control cells.
RESPONSE What signals does the immune system receive from necrotic
Histological data show necrosis as a phenomenon that involves cells? Some of the signals are already known; among them are high
large population of cells, contrary to apoptosis which involves mobility group box 1 (HMGB1), Hsp70, calreticuline, and uric acid
individual cells. This type of cell destruction may be determined by (see [157] for review). When delivered in the extracellular space,
combination of several causes: toxicity of some factors released by these substances activate APC, including dendritic cells [157].
necrotic cells (e.g., ROS, NOS) and destroying adjacent cells HMGB1 protein is a powerful pro-inflammatory factor; it is
(“bystander effect”), suppression of phagocytosis by ROS, and poor passively released by necrotic cells whereas in apoptotic cells it
mobilization of macrophages. In contrast to apoptotic cells with tightly bound to chromatin [158]. HMGB1 can either bind to TLR4
their content being isolated before phagocytosis, necrotic cells or RAGE (receptor for advanced glycation end products) (Fig. 2).
present a powerful inflammatory and immunogenic stimuli. Furthermore, HMGB1 along with Hsps and uric acid released by
Cellular thanatogenic mechanisms usually trigger the apoptotic necrotic cells is a potent adjuvant in vivo. Important role in
form of cell destruction to avoid inflammatory and autoimmune inflammation is also attributed to Hsp70 [159]. Its elevated levels in
reactions that are potentially dangerous for an organism. However, necrotic neoplastic cells markedly increased their immunogenecity
in some cases the necrotic pathway is activated, and triggering by promoting the Th1 response and APC maturation [154]. Thus,
necrosis instead of apoptosis is not just a cell’s failure, but may Hsp70 is not only a marker of necrosis, but also a specific signal for
have a positive effect when a strong inflammatory response is the immune system. Hsp70 has high immunogenicity by itself and
Fig. (2). Necrosis as a modulator of immune response.

Necrotic cells release a variety of factors stimulating macrophages (M) via their specific receptors, which leads to generation of inflammatory cytokines by
macrophages
increases immunogenicity of some other macromolecular antigens which, apparently, coincides with high apoptotic sensitivity of
[160]. There are several receptors activated by Hsp70, among them corresponding normal cells such as thymocytes, lymphocytes etc.
CD91, CD40, CD14, and TLR receptors [98, 161] (Fig. 2). Radiation and anti-neoplastic drugs at clinically relevant doses kills
This activation of macrophages and dendritic cells by necrotic these cells mostly via apoptosis, and necrosis of these cells can be
cells is accompanied by inhibition of secretion of anti-inflammatory observed only when apoptosis is blocked, e.g., by caspase inhibitors
cytokines (IL-10, TGF-), and by release of pro-inflammatory or overexpression of anti-apoptotic proteins of Bcl-2 family. For
mediators (TNF-, L-1, IL-6, MIP-2, IL-8) and chemokines (Fig. this type of neoplasia, inhibitors of proteins of Bcl-2 family may be
2). Exposed to the factors of necrotic cells, dendritic cells enter the a promising way to increase their apoptotic sensitivity and effi-
mature state that is characterized by appearance of specific markers ciency of anti-cancer therapy.
(CD40, CD80, CD86), costimulating molecules (B7.1, B7.2), For most widespread tumors of epithelial origin such as breast
stimulation of T-cell proliferation etc. Immunization of animals and prostate cancers, apoptosis can be rarely found in vivo during
with dendritic cells loaded with necrotic tumor cells markedly tumor development or after anti-cancer therapy, and in vitro it can
suppressed the growth of appropriate tumors (see ref [162] for be caused only by relatively high (unattainable in clinic) doses of
review). On other hand, most cytokines that promote inflammation radiation and drug. Accordingly, most normal epithelial cells are
(e.g., TNF- or IL-6) can activate NF-kB signaling pathway in also relatively resistant to apoptosis. Moreover, there is no
tumor cells that protects them from apoptosis and promote tumor dependence between apoptotic sensitivity of epithelial tumors and
cell proliferation (see [163] for review). their response to anti-cancer therapy. For instance, high expression
of anti-apoptotic Bcl-2 protein in solid tumors paradoxically
4. NECROTIC DEATH IN ANTI-CANCER THERAPY AND correlate with better prognosis [164, 165] and there was no apparent
TUMOR DEVELOPMENT correlation between sensitivity of tumor to therapy and expression
Extensive studies of tumor cell death during the last decade of other pro- and anti-apoptotic members of BH3-family.
allow to come to several conclusions regarding connections bet- At the same time, necrosis of solid tumors in vivo can be
ween three main modes of death: apoptosis, necrosis, and auto- observed quite often after anti-cancer treatment or during tumor
phagic cell death. For neoplastic cells of hematopoetic origin, development. Since conventional anti-cancer drugs or radiation can
apoptosis is a prevalent form of cell death in vitro and in vivo,
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Received: June 18, 2009 Accepted: June 30, 2009
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Mechanisms of Tumor Cell Necrosis

Article in Current Pharmaceutical Design · January 2010

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Mechanisms of Tumor Cell Necrosis

Sergey Y. Proskuryakov1 and Vladimir L. Gabai2*

INTRODUCTION death accompanied by a rapid efflux of cell constituents in extra-

1381-6128/10 $55.00+.00 © 2010 Bentham Science Publishers Ltd.

Table 1. Activators and Inhibitors of Necrosis

Activators Cells Inhibitors References

Ischemia Various cells Necrostatin-1 [77]

Glutamate HT-22 Necrostatin-1 [174]

TNF L929, MEF BHA [29]

FAS T-cells - [175]

TRAIL (pH6.5) HT-29 zVAD [26]

LPS Macrophages - [176]

Anti-CD47 B-lymphocytes, CLL TPCK [9]

IL1+IFN L929 SB203580; sc-514 [56]

ZVAD L929 Rapamycin [129]

Ceramide Jurkat, 3T3 - [28]

ROS MEF SP600125 [38]

Photodynamic therapy HeLa MitoQ [177]

As203 HeLa NAC, PARP inhibitors [81]

Tamoxifen MCF7, MDA-MB231 Estrogens, PD98059, BA [13,14]

MNNG MEF PARP inhibitors, SP600125 [10,30]

Imatinib K562 TPCK [18]

Honokiol HL-60, MCF7, HEK293 CsA [16]

Shikonin MCF7, HEK293 Necrostatin-1 [17]

Resveratrol MCF7 - [15]

facilitate cellular uptake) can induce selective necrosis in p53-

2. SIGNALING PATHWAYS AND MODULATORS OF

2.1. Receptors and Protein Kinase Cascades

Fig. (2). Necrosis as a modulator of immune response.

Received: June 18, 2009 Accepted: June 30, 2009

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