Absence Of Caspase Activation Research Articles

Spontaneous Overactivation of Xenopus Frog Eggs Triggers Necrotic Cell Death.

The excessive activation of frog eggs, referred to as overactivation, can be initiated by strong oxidative stress, leading to expedited calcium-dependent non-apoptotic cell death. Overactivation also occurs spontaneously, albeit at a low frequency, in natural populations of spawned frog eggs. Currently, the cytological and biochemical events of the spontaneous process have not been characterized. In the present study, we demonstrate that the spontaneous overactivation of Xenopus frog eggs, similarly to oxidative stress- and mechanical stress-induced overactivation, is characterized by the fast and irreversible contraction of the egg's cortical layer, an increase in egg size, the depletion of intracellular ATP, a drastic increase in the intracellular ADP/ATP ratio, and the degradation of M phase-specific cyclin B2. These events manifest in eggs in the absence of caspase activation within one hour of triggering overactivation. Importantly, substantial amounts of ATP and ADP leak from the overactivated eggs, indicating that plasma membrane integrity is compromised in these cells. The rupture of the plasma membrane and acute depletion of intracellular ATP explicitly define necrotic cell death. Finally, we report that egg overactivation can occur in the frog's genital tract. Our data suggest that mechanical stress may be a key factor promoting egg overactivation during oviposition in frogs.

Oxidative Stress-Induced Overactivation of Frog Eggs Triggers Calcium-Dependent Non-Apoptotic Cell Death.

Excessive activation of frog eggs (overactivation) is a pathological process that renders eggs unfertilizable. Its physiological inducers are unknown. Previously, oxidative stress was shown to cause time- and dose-dependent overactivation of Xenopus laevis frog eggs. Here, we demonstrate that the oxidative stress-induced egg overactivation is a calcium-dependent phenomenon which can be attenuated in the presence of the selective calcium chelator BAPTA. Degradation of cyclin B2, which is known to be initiated by calcium transient in fertilized or parthenogenetically activated eggs, can also be observed in the overactivated eggs. Decline in mitochondrial membrane potential, ATP depletion and termination of protein synthesis manifest in the eggs within one hour of triggering overactivation. These intracellular events occur in the absence of caspase activation. Furthermore, plasma membrane integrity is compromised in the overactivated eggs, as evidenced by ATP leakage and egg swelling. In sum, our data demonstrate that oxidative stress-induced overactivation of frog eggs causes fast and dramatic disruption of cellular homeostasis, resulting in robust and expedited cell death by a calcium-dependent non-apoptotic mechanism.

Diversity of cell death signaling pathways in macrophages upon infection with modified vaccinia virus Ankara (MVA)

Regulated cell death frequently occurs upon infection by intracellular pathogens, and extent and regulation is often cell-type-specific. We aimed to identify the cell death-signaling pathways triggered in macrophages by infection with modified vaccinia virus Ankara (MVA), an attenuated strain of vaccinia virus used in vaccination. While most target cells seem to be protected by antiapoptotic proteins encoded in the MVA genome, macrophages die when infected with MVA. We targeted key signaling components of specific cell death-pathways and pattern recognition-pathways using genome editing and small molecule inhibitors in an in vitro murine macrophage differentiation model. Upon infection with MVA, we observed activation of mitochondrial and death-receptor-induced apoptosis-pathways as well as the necroptosis-pathway. Inhibition of individual pathways had a little protective effect but led to compensatory death through the other pathways. In the absence of mitochondrial apoptosis, autocrine/paracrine TNF-mediated apoptosis and, in the absence of caspase-activity, necroptosis occurred. TNF-induction depended on the signaling molecule STING, and MAVS and ZBP1 contributed to MVA-induced apoptosis. The mode of cell death had a substantial impact on the cytokine response of infected cells, indicating that the immunogenicity of a virus may depend not only on its PAMPs but also on its ability to modulate individual modalities of cell death. These findings provide insights into the diversity of cell death-pathways that an infection can trigger in professional immune cells and advance our understanding of the intracellular mechanisms that govern the immune response to a virus.

Cytotoxicity of [HuArgI (co)-PEG5000]-induced arginine deprivation to ovarian Cancer cells is autophagy dependent.

In this study, we assess arginine auxotrophy in ovarian cancer cells and attempt to target them using arginine deprivation induced by a pegylated recombinant human Arginase I cobalt [HuArgI (Co)-PEG5000]. Ovarian cancer cells were sensitive to [HuArgI (Co)-PEG5000]-induced arginine deprivation with IC50 values in the low pM range. Addition of excess L-citrulline rescued only one of three cell lines tested, indicating that the majority of cell lines are completely auxotrophic for arginine. The expression pattern of argininosuccinate synthetase (ASS1) confirmed the degree of auxotrophy of ovarian cancer cell lines with completely auxotrophic cells not expressing ASS1 and partially auxotrophic cells expressing the enzyme. Ovarian cancer cell lines were negative for annexinV staining while showing loss of membrane integrity and absence of caspase activation, indicating caspase-independent, non-apoptotic cell death. [HuArgI (Co)-PEG5000]-induced arginine deprivation led to extensive and prolonged activation of autophagy, which proved to be deleterious to cell survival since its inhibition led to a significant decrease in cytotoxicity. This indicates that the activation of autophagy following arginine-deprivation, rather than being protective, mediates cell cytotoxicity leading to death by autophagy.

Mitochondrial inner membrane permeabilisation enables mtDNA release during apoptosis.

During apoptosis, pro‐apoptotic BAX and BAK are activated, causing mitochondrial outer membrane permeabilisation (MOMP), caspase activation and cell death. However, even in the absence of caspase activity, cells usually die following MOMP. Such caspase‐independent cell death is accompanied by inflammation that requires mitochondrial DNA (mtDNA) activation of cGAS‐STING signalling. Because the mitochondrial inner membrane is thought to remain intact during apoptosis, we sought to address how matrix mtDNA could activate the cytosolic cGAS‐STING signalling pathway. Using super‐resolution imaging, we show that mtDNA is efficiently released from mitochondria following MOMP. In a temporal manner, we find that following MOMP, BAX/BAK‐mediated mitochondrial outer membrane pores gradually widen. This allows extrusion of the mitochondrial inner membrane into the cytosol whereupon it permeablises allowing mtDNA release. Our data demonstrate that mitochondrial inner membrane permeabilisation (MIMP) can occur during cell death following BAX/BAK‐dependent MOMP. Importantly, by enabling the cytosolic release of mtDNA, inner membrane permeabilisation underpins the immunogenic effects of caspase‐independent cell death.

Mitochondrial permeabilization engages NF-κB-dependent anti-tumour activity under caspasedeficiency.

Apoptosis represents a key anti-cancer therapeutic effector mechanism. During apoptosis, mitochondrial outer membrane permeabilisation (MOMP) typically kills cells even in the absence of caspase activity. Caspase activity can also have a variety of unwanted consequences that include DNA-damage. We therefore investigated whether MOMP-induced caspase-independent cell death (CICD) might be a better way to kill cancer cells. We find that cells undergoing CICD display potent pro-inflammatory effects relative to apoptosis. Underlying this, MOMP was found to stimulate NF-κB activity through the down-regulation of inhibitor of apoptosis (IAP) proteins. Strikingly, engagement of CICD displays potent anti-tumorigenic effects, often promoting complete tumour regression in a manner dependent on intact immunity. Our data demonstrate that by activating NF-κB, MOMP can exert additional signalling functions besides triggering cell death. Moreover, they support a rationale for engaging caspase-independent cell death in cell-killing anti-cancer therapies.

Biological activity of tumor-treating fields in preclinical glioma models

Glioblastoma is the most common and aggressive form of intrinsic brain tumor with a very poor prognosis. Thus, novel therapeutic approaches are urgently needed. Tumor-treating fields (TTFields) may represent such a novel treatment option. The aim of this study was to investigate the effects of TTFields on glioma cells, as well as the functional characterization of the underlying mechanisms. Here, we assessed the anti-glioma activity of TTFields in several preclinical models. Applying TTFields resulted in the induction of cell death in a frequency- and intensity-dependent manner in long-term glioma cell lines, as well as glioma-initiating cells. Cell death occurred in the absence of caspase activation, but involved autophagy and necroptosis. Severe alterations in cell cycle progression and aberrant mitotic features, such as poly- and micronucleation, preceded the induction of cell death. Furthermore, exposure to TTFields led to reduced migration and invasion, which are both biological hallmarks of glioma cells. The combination of TTFields with irradiation or the alkylating agent, temozolomide (TMZ), resulted in additive or synergistic effects, and the O6-methyl-guanine DNA methyltransferase status did not influence the efficacy of TTFields. Importantly, TMZ-resistant glioma cells were responsive to TTFields application, highlighting the clinical potential of this therapeutic approach. In summary, our results indicate that TTFields induce autophagy, as well as necroptosis and hamper the migration and invasiveness of glioma cells. These findings may allow for a more detailed clinical evaluation of TTFields beyond the clinical data available so far.

Contribution of caspase-independent pathway to apoptosis in malignant glioma induced by carbon ion beams.

High linear energy transfer (LET) carbon ion beam (CIB) is becoming the best tool for external radiotherapy of inoperable tumors because of its greater cell killing than conventional low LET gamma or X-rays. In the present study, whether the caspase-independent pathway exerts the important contribution in CIB-induced cell apoptosis was explored. Herein we showed, despite the absence of caspase activity using a pan caspase inhibitor Z-VAD-FMK, that apoptosis induced by high LET CIB were clearly observed in the glioma cells. Simultaneously, the increased 8-OHdG level, PARP-1 activity and AIF translocation occurred in response to CIB irradiation. Moreover, it was distinctly higher in the nuclear translocation frequency along with PARP-1 activation when the caspase protease cascade was suppressed in the irradiated glioma cells. Nuclear colocalization between PARP-1 and AIF as well as a positive association of the PARP-1 mRNA expression with AIF translocation frequency indicated that PARP-1 activation controlled the translocation of AIF to the nucleus. Our findings strongly demonstrated that caspase-independent cell apoptosis provided a prominent compensation in the glioma cell death involving the PARP-1/AIF signaling pathway at 24h after CIB exposure, and likely triggered by oxidative damage to DNA. The knowledge on the molecular mechanism of AIF-mediated cell death may be very useful for the improvement of the therapeutic efficacy of malignant gliomas with heavy charged particles.

K45A mutation of RIPK1 results in poor necroptosis and cytokine signaling in macrophages, which impacts inflammatory responses in vivo.

Receptor interacting protein kinase 1 (RIPK1) participates in several cell signaling complexes that promote cell activation and cell death. Stimulation of RIPK1 in the absence of caspase signaling induces regulated necrosis (necroptosis), which promotes an inflammatory response. Understanding of the mechanisms through which RIPK1 promotes inflammation has been unclear. Herein we have evaluated the impact of a K45A mutation of RIPK1 on necroptosis of macrophages and the activation of inflammatory response. We show that K45A mutation of RIPK1 results in attenuated necroptosis of macrophages in response to stimulation with LPS, TNFα and IFNβ in the absence of caspase signaling. Impairment in necroptosis correlated with poor phosphorylation of RIPK1, RIPK3 and reduced trimerization of MLKL. Furthermore, K45A mutation of RIPK1 resulted in poor STAT1 phosphorylation (at S727) and expression of RANTES and MIP-1α following TNF-R engagement in the absence of caspase activation. Our results further indicate that in the absence of stimulation by pathogen-associated molecular patterns (PAMPs), cellular inhibitors of apoptotic proteins (cIAPs) prevent the K45-dependent auto-phosphorylation of RIPK1, leading to resistance against necroptosis. Finally, RIPK1(K45A) mice displayed attenuated inflammatory response in vivo as they were significantly resistant against endotoxin shock, but highly susceptible against a challenge with Salmonella typhimurium. This correlated with reduced expression of IL-1β and ROS, and poor processing of caspase 8 by RIPK1(K45A) macrophages. Overall, these results indicate that K45 mediated kinase activity of RIPK1 is not only important for necroptosis but it also has a key role in promoting cytokine signaling and host response to inflammatory stimuli.

The interplay between human herpes simplex virus infection and the apoptosis and necroptosis cell death pathways.

Human herpes simplex virus (HSV) is a ubiquitous human pathogen that establishes a lifelong latent infection and is associated with mucocutaneous lesions. In multicellular organisms, cell death is a crucial host defense mechanism that eliminates pathogen-infected cells. Apoptosis is a well-defined form of programmed cell death executed by a group of cysteine proteases, called caspases. Studies have shown that HSV has evolved strategies to counteract caspase activation and apoptosis by encoding anti-apoptotic viral proteins such as gD, gJ, Us3, LAT, and the ribonucleotide reductase large subunit (R1). Recently, necroptosis has been identified as a regulated form of necrosis that can be invoked in the absence of caspase activity. Receptor-interacting kinase 3 (RIP3 or RIPK3) has emerged as a central signaling molecule in necroptosis; it is activated via interaction with other RIP homotypic interaction motif (RHIM)-containing proteins such as RIP1 (or RIPK1). There is increasing evidence that HSV R1 manipulates necroptosis via the RHIM-dependent inactivation or activation ofRIP3 in a species-specific manner. This review summarizes the current understanding of the interplay between HSV infection and cell death pathways, with an emphasis on apoptosis and necroptosis.

BNIP3 promotes calcium and calpain-dependent cell death

Loss of cardiac muscle by programmed cell death contributes to the progression of ischemic heart disease. Hypoxia, metabolite waste buildup and energy depletion are components of ischemia which may initiate caspase dependent and independent cell death pathways. Previous work from our laboratory has shown that combined hypoxia with acidosis, a hallmark of ischemia promotes cardiac myocyte injury with increasing severity as the pH declines. Hypoxia-acidosis was demonstrated to activate the pro-apoptotic Bcl-2 protein BNIP3 which initiated opening of the mitochondrial permeability transition pore and cell death in the absence of caspase activation. Because calpains are known to contribute to ischemic myocardial damage in some models, we hypothesized that they are intermediates in the BNIP3-mediated death caused by hypoxia-acidosis. Neonatal rat cardiac myocytes were subjected to hypoxia with and without acidosis and the contribution of calpains to hypoxia-acidosis cell death determined. Here we report that the death pathway activated by hypoxia-acidosis is driven by a combination of calcium-activated calpains and pro-death factors (DNases) secreted by the mitochondria. Cytochrome c accumulated in the cytoplasm during hypoxia-acidosis but caspase activity was repressed through a calpain-dependent process that prevents the cleavage of procaspase 3. Calpain inhibitors provide vigorous protection against hypoxia-acidosis-induced programmed death. Knockdown of BNIP3 with siRNA prevented calpain activation confirming a central role of BNIP3 in this pathway. The results implicate BNIP3 and calpain as dependent components of cardiac myocyte death caused by hypoxia-acidosis.

8-p-Hdroxybenzoyl Tovarol Induces Paraptosis Like Cell Death and Protective Autophagy in Human Cervical Cancer HeLa Cells

8-p-Hdroxybenzoyl tovarol (TAW) is a germacrane-type sesquiterpenoid that can be isolated from the roots of Ferula dissecta (Ledeb.) Ledeb. In this study, the growth inhibitory effects induced by TAW were screened on some types of tumor cells, and the mechanism was investigated on TAW-induced growth inhibition, including paraptosis and autophagy in human cervical cancer HeLa cells. TAW-induced paraptosis involved extensive cytoplasmic vacuolization in the absence of caspase activation. Additionally, TAW evoked cell paraptotic death mediated by endoplasmic reticulum (ER) stress and unfolded protein response (UPR). Autophagy induced by TAW was found to antagonize paraptosis in HeLa cells. This effect was enhanced by rapamycin and suppressed by the autophagy inhibitor, 3-methyladenine (3MA). Loss of beclin 1 (an autophagic regulator) function led to promote ER stress. Taken together, these results suggest that TAW induces paraptosis like cell death and protective autophagy in HeLa cells, which would provide a new clue for exploiting TAW as a promising agent for the treatment of cervical cancer.

GSK3A is redundant with GSK3B in modulating drug resistance and chemotherapy-induced necroptosis.

Glycogen Synthase Kinase-3 alpha (GSK3A) and beta (GSK3B) isoforms are encoded by distinct genes, are 98% identical within their kinase domain and perform similar functions in several settings; however, they are not completely redundant and, depending on the cell type and differentiative status, they also play unique roles. We recently identified a role for GSK3B in drug resistance by demonstrating that its inhibition enables necroptosis in response to chemotherapy in p53-null drug-resistant colon carcinoma cells. We report here that, similarly to GSK3B, also GSK3A silencing/inhibition does not affect cell proliferation or cell cycle but only abolishes growth after treatment with DNA-damaging chemotherapy. In particular, blocking GSK3A impairs DNA repair upon exposure to DNA-damaging drugs. As a consequence, p53-null cells overcome their inability to undergo apoptosis and mount a necroptotic response, characterized by absence of caspase activation and RIP1-independent, PARP-dependent AIF nuclear re-localization. We therefore conclude that GSK3A is redundant with GSK3B in regulating drug-resistance and chemotherapy-induced necroptosis and suggest that inhibition of only one isoform, or rather partial inhibition of overall cellular GSK3 activity, is enough to re-sensitize drug-resistant cells to chemotherapy.

Involvement of poly(ADP-ribose) polymerase in paraptotic cell death of D. discoideum

Paraptosis is mediated by several proteins, poly(ADP-ribose) polymerase being one of them. D. discoideum lacks caspases thus providing a better system to dissect out the role of PARP in paraptosis. The cell death phenotype in unicellular eukaryote, D. discoideum is similar to the programmed cell death phenotype of multicellular animals. However, the events downstream to the death signal of PCD in D. discoideum are yet to be understood. Our results emphasize that oxidative stress in D. discoideum lacking caspases leads to PARP activation, mitochondrial membrane potential changes, followed by the release of apoptosis inducing factor from mitochondria. AIF causes large scale DNA fragmentation, a hallmark feature of paraptosis. The role of PARP in paraptosis is reiterated via PARP inhibition by benzamide, PARG inhibition by gallotannin and PARP down-regulation, which delays paraptosis. PARP, PARG and AIF interplay is quintessential in paraptosis of D. discoideum. This is the first report to establish the involvement of PARP in the absence of caspase activity in D. discoideum which could be of evolutionary significance and gives a lead to understand the caspase independent paraptotic mechanism in higher organisms.

Mitochondrial Regulation of Cell Death

Although required for life, paradoxically, mitochondria are often essential for initiating apoptotic cell death. Mitochondria regulate caspase activation and cell death through an event termed mitochondrial outer membrane permeabilization (MOMP); this leads to the release of various mitochondrial intermembrane space proteins that activate caspases, resulting in apoptosis. MOMP is often considered a point of no return because it typically leads to cell death, even in the absence of caspase activity. Because of this pivotal role in deciding cell fate, deregulation of MOMP impacts on many diseases and represents a fruitful site for therapeutic intervention. Here we discuss the mechanisms underlying mitochondrial permeabilization and how this key event leads to cell death through caspase-dependent and -independent means. We then proceed to explore how the release of mitochondrial proteins may be regulated following MOMP. Finally, we discuss mechanisms that enable cells sometimes to survive MOMP, allowing them, in essence, to return from the point of no return.

DsDNA ASCs for caspase 8-mediated apoptosis

dsDNA ASCs for caspase 8-mediated apoptosis

Melatonin induces apoptosis through a caspase‐dependent but reactive oxygen species‐independent mechanism in human leukemia Molt‐3 cells

Melatonin is a naturally occurring indoleamine synthesized in the pineal gland that exhibits an extensive repertoire of biological activities. An increasing number of studies indicate that melatonin protects normal cells, while it reducing cancer cell proliferation. In this study, we investigated the effect of melatonin on the growth of the human leukemia cells and found that it efficiently reduced the number of cells in a concentration- and time-dependent manner. Thus, incubation with the indoleamine increased the percentage of cells with a hypodiploid DNA content, augmented the number of annexin V-positive cells, and also provoked ultrastructural changes that are features of apoptotic cell death. Evaluation of caspases revealed that caspase-3, caspase-6, caspase-7, and caspase-9, but not caspase-8 and caspase-2, were quickly activated (3-6hr). The increase in the activity of these proteases was associated with up-regulation of the pro-apoptotic factor Bax and also with the release of cytochrome c from mitochondria. Pretreatment of the cells with the general caspase inhibitor z-VAD-fmk, reduced melatonin-induced apoptosis, but it did not block cell death suggesting that melatonin activates an alternative cell death modality in the absence of caspase activity. Thus, the activation of caspases was preceded by a fast (<30min) increase in reactive oxygen species (ROS). Rotenone and antimycin A reduced the levels of ROS stimulated by melatonin, indicating that the complex I and the complex III of the mitochondrial electron transport chain are important sources of these chemical species. However, the role of ROS in melatonin-induced cell death remains elusive because anti-oxidants that were shown to decrease ROS levels (glutathione, N-acetyl-l-cysteine and Trolox) were unable to abrogate melatonin-induced cell death.

211 Concanavalin A-Induced Hepatitis in Mice Is Prevented by Depletion of Gut Bacteria Using Antibiotics

L or hyposmotic conditions, as measured by LDH and HMGB1 release. Upon hyposmotic stress of hepatocytes, cell death occurred in the absence of caspase activation. Similar to primary hepatocytes, K18-DE mice treated with Fas-L had elevated serum necrosis markers (LDH and HMGB1) as compared to their WT counterparts. K18-DE mutation renders the K18 obligate heteropolymer, K8, less susceptible to phosphorylation at its p38 stress kinase site K8 S79, without altering the binding of K8/K18 to p38 kinase. Notably, K8 S79 phosphorylation plays an important role in mediating keratin filament reorganization upon stress. Conclusions: Caspase digestion of K18 plays an essential role in promoting apoptosisassociated keratin filament collapse in the liver. Keratin phosphorylation-mediated filament reorganization appears to be critical in protecting hepatocytes from necroapoptosis, and provides a mechanism for susceptibility to tissue injury in human IF mutations that involve caspase cleavage sites.

Cytotoxicity of Anthrax Lethal Toxin to Human Acute Myeloid Leukemia Cells Is Nonapoptotic and Dependent on Extracellular Signal-Regulated Kinase 1/2 Activity

In this study, we attempt to target the mitogen-activated protein kinase (MAPK) pathway in acute myeloid leukemia (AML) cells using a recombinant anthrax lethal toxin (LeTx). LeTx consists of protective antigen (PrAg) and lethal factor (LF). PrAg binds cells, is cleaved by furin, oligomerizes, binds three to four molecules of LF, and undergoes endocytosis, releasing LF into the cytosol. LF cleaves MAPK kinases, inhibiting the MAPK pathway. We tested potency of LeTx on a panel of 11 human AML cell lines. Seven cell lines showed cytotoxic responses to LeTx. Cytotoxicity of LeTx was mimicked by the specific mitogen-activated protein/extracellular signal-regulated kinase kinase 1/2 (MEK1/2) inhibitor U0126, indicating that LeTx-induced cell death is mediated through the MEK1/2-extracellular signal-regulated kinase (ERK1/2) branch of the MAPK pathway. The four LeTx-resistant cell lines were sensitive to the phosphatidylinositol 3-kinase inhibitor LY294002. Co-treatment of AML cells with both LeTx and LY294002 did not lead to increased sensitivity, showing a lack of additive/synergistic effects when both pathways are inhibited. Flow cytometry analysis of MAPK pathway activation revealed the presence of phospho-ERK1/2 only in LeTx-sensitive cells. Staining for Annexin V/propidium iodide and active caspases showed an increase in double-positive cells and the absence of caspase activation following treatment, indicating that LeTx-induced cell death is caspase-independent and nonapoptotic. We have shown that a majority of AML cell lines are sensitive to the LF-mediated inhibition of the MAPK pathway. Furthermore, we have demonstrated that LeTx-induced cytotoxicity in AML cells is nonapoptotic and dependent on phospho-ERK1/2 levels.

Apoptotic-like death occurs through a caspase-independent route in colon carcinoma cells undergoing mitotic catastrophe

We have examined the relationship between chemotherapy-induced mitotic catastrophe and cell death by apoptosis in both wild-type and p53(−/−) HCT116 human colon carcinoma cells treated with nanomolar concentrations of paclitaxel (PTX), a drug that acts on tubulin altering the normal development of mitosis. After treatment, HCT116 cells entered mitosis regardless of the presence of functional p53, which resulted in changes in the distribution of cells in the different phases of the cell cycle, and in cell death. In the presence of PTX, the percentage of polyploid cells observed was higher in p53-deficient cells, indicating that mitotic slippage was favored compared to wild-type cells, with the presence of large multinucleate cells. PTX caused mitotic catastrophe and about 50–60% cells that were entering an aberrant mitosis died through an apoptotic-like pathway characterized by the presence of phosphatidylserine in the outer cell membrane, which occurred in the absence of significant activation of caspases. Lack of p53 facilitated endoreduplication and polyploidy in PTX-treated cells, but cells were still killed with similar efficacy through the same apoptotic-like mechanism in the absence of caspase activity.