Selective Cell Death Research Articles

Medicinal Plants used for the Treatment of Parkinson's Disease

Parkinsonism is single of the regular neurodegenerative diseases, which is distinguished through a discriminating and advancement deterioration of dopaminergic neurons, root a sequence of sign which might finally persuade automatic cell demise, although the etiology of parkinsonism ruins unknown, current studies have optional that oxidative stress(OS), generate apoptosis which results in mitochondrial defects, neuroinflammation may also play important roles in its pathogenesis. Various agents as 6-Hydroxydopomine(6-OHDA), 1-Methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine, Rotenone a neurotoxin commonly and many more are used in models of PD, induces selective catecholaminergic cell death, mediated by reactive oxygen species(ROS) and mitochondrial defects.

The Mathematical Model of Disturbance of Energy Metabolism in Brain during Development of Neurodegenerative Diseases: A Proposed Mechanism of Cell Death

The paper presents a theoretical study of the failure of energy metabolism in nervous tissue on a detailed biophysical model of metabolic coupling between neurons and astrocytes under afferent stimulation simulating by the release of glutamate into the presynaptic cleft. The main result of the model study is the detection of the phenomenon of adenylate collapse, the essence of which is the irreversible drop of ATP concentration after reaching the critical threshold value. This effect occurs as a result of the reversing of the reaction catalyzed by adenylate kinase. At high ATP values, the reaction equilibrium is shifted towards the formation of ADP from ATP and AMP. When the threshold is reached, the equilibrium shifts in the opposite direction, i.e., the formation of AMP and ATP from ADP occurs. If this situation is accompanied by a high consumption of ATP, the entire pool of adenine nucleotides goes into AMP. However, AMP is not phosphorylated, unlike ADP in glycolysis and oxidative phosphorylation, so the system quickly comes to depletion of ATP. Thus, adenylate collapse can be a new mechanism of the selective cell death in the development of neurodegenerative diseases.

Expression of activating transcription factor 5 (ATF5) is increased in astrocytomas of different WHO grades and correlates with survival of glioblastoma patients.

BackgroundATF5 suppresses differentiation of neuroprogenitor cells and is overexpressed in glioblastoma (GBM). A reduction of its expression leads to apoptotic GBM cell death. Data on ATF5 expression in astrocytoma WHO grade II (low-grade astrocytoma [LGA]) are scarce and lacking on recurrent GBM.Patients and methodsATF5 mRNA was extracted from frozen samples of patients’ GBM (n=79), LGA (n=40), and normal brain (NB, n=10), quantified by duplex qPCR and correlated with retrospectively collected clinical data. ATF5 protein expression was evaluated by measuring staining intensity on immunohistochemistry.ResultsATF5 mRNA was overexpressed in LGA (sevenfold, P<0.001) and GBM (tenfold, P<0.001) compared to NB, which was confirmed on protein level. Although ATF5 mRNA expression in GBM showed a considerable fluctuation range, groups of varying biological behavior, that is, local/multifocal growth or primary tumor/relapse and the tumor localization at diagnosis, were not significantly different. ATF5 mRNA correlated with the patients’ age (r=0.339, P=0.028) and inversely with Ki67-staining (r=−0.421, P=0.007). GBM patients were allocated to a low and a high ATF5 expression group by the median ATF5 overexpression compared to NB. Kaplan–Meier analysis and Cox regression indicated that ATF5 mRNA expression significantly correlated with short-term survival (t,12 months, median survival 18 vs 13 months, P=0.022, HR 2.827) and progression-free survival (PFS) (12 vs 6 months, P=0.024). This advantage vanished after 24 months (P=0.084).ConclusionATF5 mRNA expression could be identified as an additional, though not independent factor correlating with overall survival and PFS. Since its inhibition might lead to the selective death of glioma cells, it might serve as a potential ubiquitous therapeutic target in astrocytic tumors.

Soluble curcumin prevents cadmium cytotoxicity in primary rat astrocytes by improving a lack of GFAP and glucose-6-phosphate-dehydrogenase

Cadmium (Cd) is a heavy metal which is widespread in various environmental components. Moreover several occupational diseases have the complications that are related to Cd cytotoxicity. Low doses of Cd exposure could induce pathogenetic disturbances in several sensitive cells as result of its long biological half-life and accumulation in vital tissue types. Prolonged Cd exposure was determined as main factor in accumulation of this metal ion over time in the liver, kidneys and neural tissue cells. The neurotoxic effect of Cd was presented in several articles which reported both in vivo and in vitro study. One of the main causes of Cd neurotoxicity is the ability of this ion to increase the permeability of the blood brain barrier. Despite a focus of attention on Cd cytotoxicity over the last few decades, the effect of Cd in neural tissue cells has been presented in a limited number of articles. The neurotoxic effect of Cd is accompanied by biochemical changes as well as a lack of functional activity of the central nervous system. Taking into account that the cytotoxic effect of Cd is associated with oxidative stress, inflammation and selective cell death, antioxidants could be used to protect neural tissue cells against both chronic and acute Cd exposure. Antioxidants protect diverse cell types against metal induced cytotoxicity. Curcumin is a natural polyphenol which exhibits antioxidant and anti-inflammatory effect. Soluble forms of cucrcumin can penetrate the blood brain barrier and protect neural tissue cells against detrimental effects of cytotoxic compounds. Glial cells are the most numerous cell population in CNS. Astrocytes possess the ability to protect the neuronal cells against cytotoxicity and maintain CNS functions. The cytoskeleton of astrocytes is constructed with glial fibrillary acidic protein (GFAP). GFAP is involved in essential functions of astrocytes and reflects astrocyte reactivity. The molecular mechanisms of the neurotoxic effect of Cd on glial cytoskeleton remain unknown. Primary astrocyte cell culture was used as model to assess the gliotoxic effect of Cd as well as the potency of low doses of soluble curcumin to ameliorate the neurotoxic effect of Cd. The obtained results demonstrated depletion of GFAP and glucose-6-phosphate-dehydrogenase (G6PD) in astrocytes treated with 10 µM Cd. The exposure to 5 µM curcumin ameliorated the expression both of GFAP and G6PD in Cd suppressed astrocytes. Moreover, low doses of soluble curcumin significantly prevented the detrimental effects of Cd on cell viability and indices of oxidative stress. The obtained results are evidence that soluble forms of curcumin improve astrocyte viability, cytoskeleton depletion and glucose utilization pathway. Thus, soluble curcumin possesses a neuroprotective effect directed on astrocyte cytoskeleton and metabolic energy production.

Piperlongumine Induces Apoptosis in Colorectal Cancer HCT 116 Cells Independent of Bax, p21 and p53 Status

Colorectal cancer is a common type of cancer with reported resistance to treatment, in most cases due to loss of function of apoptotic and cell-cycle proteins. Piperlongumine (PPLGM) is a natural alkaloid isolated from Piper species, with promising anti-cancer properties. This study investigated whether PPLGM is able to induce cell death in colorectal carcinoma HCT 116 cells expressing wild-type or deficient in Bax, p21 or p53. PPLGM was extracted from roots of Piper tuberculatum. Cell viability was determined by reduction of 3-(4,5-dimethilthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and clonogenic assay. Cell death was evaluated by acridine orange/ethidium bromide staining and flow cytometry. Plasmid cleavage activity and circular dichroism DNA interaction were also analyzed. PPLGM induced selective cell death in all cell lines (IC50 range from 10.7 to 13.9 μM) with an increase in the number of late apoptotic cells and different profiles in cell-cycle distribution. Plasmid DNA analysis showed that PPLGM does not interact directly with DNA. This paper suggests that PPLGM may be a promising candidate in colorectal cancer therapy.

Different contributions of autophagy to retinal ganglion cell death in the diabetic and glaucomatous retinas

Diabetes mellitus and glaucoma are the two major causes of selective retinal ganglion cell (RGC) death. To determine the relationship between autophagy and RGC death, we compared autophagy and the related molecular pathways in diabetic and glaucomatous retinas and examined their effect on RGC survival. Biochemical analysis of microtubule-associated protein light chain 3 (LC3)-II and beclin-1 were observed. To determine the pathways involved in autophagy induction, adenosine monophosphate-activated protein kinase (AMPK) and the mechanistic target of rapamycin (mTOR) were also explored. Beclin-1 and the LC3B-II to LC3B-I ratio significantly elevated at 4 and 8 weeks after glaucoma induction; however, only a slight increase was apparent in the diabetic retina. Significant upregulation of phosphorylated AMPK and downregulation of phosphorylated mTOR was evident in the diabetic retina. After autophagy was inhibited with 3-methyladenine (3-MA), apoptosis of RGCs was significantly increased in the diabetic retinas. However, 3-MA inhibition of autophagy decreased the apoptosis of RGCs in glaucomatous retinas. Therefore, our results suggest that RGC death is differentially regulated by autophagy and that the pathways involved differ depending on the triggering injury.

Relationship between Liver Pathology and Disease Progression in a Murine Model of Amyotrophic Lateral Sclerosis

Background: Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease that causes selective motor neuron cell death and accompanying skeletal muscle atrophy and structural deformities. In both patients with ALS and animal models, there appears to be spinal cord and muscle pathology. This pathology can be modeled in hSOD1^G93A mice, which have a point mutation in the gene for superoxide dismutase 1. Similar to patients with ALS, hSOD1^G93A mice present hepatic abnormalities and lymphocytic infiltration in the liver. However, the relationship between liver function and disease progression is not well understood. Objective: The goal of this study was to investigate the molecular mechanisms relating liver pathology to disease progression in hSOD1^G93A mice. Methods: Liver tissues were harvested from control (nontransgenic) mice, presymptomatic hSOD1^G93A mice, and symptomatic hSOD1^G93A mice. Results: In the liver, the expression of proteins related to inflammation and oxidative stress increased with disease progression in hSOD1^G93A mice. Furthermore, histone deacetylase 4, DNA-damage-inducible 45α, and platelet-derived growth factor β, which are associated with liver fibrosis, were upregulated in the livers of presymptomatic hSOD1^G93A mice. Conclusions: Taken together, these findings suggest that liver dysfunction in hSOD1^G93A transgenic mice is mediated by increased inflammation and oxidative stress as well as the upregulation of fibrosis-related proteins.

Enzymatic Assemblies Disrupt the Membrane and Target Endoplasmic Reticulum for Selective Cancer Cell Death.

The endoplasmic reticulum (ER) is responsible for the synthesis and folding of a large number of proteins, as well as intracellular calcium regulation, lipid synthesis, and lipid transfer to other organelles, and is emerging as a target for cancer therapy. However, strategies for selectively targeting the ER of cancer cells are limited. Here we show that enzymatically generated crescent-shaped supramolecular assemblies of short peptides disrupt cell membranes and target ER for selective cancer cell death. As revealed by sedimentation assay, the assemblies interact with synthetic lipid membranes. Live cell imaging confirms that the assemblies impair membrane integrity, which is further supported by lactate dehydrogenase (LDH) assays. According to transmission electron microscopy (TEM), static light scattering (SLS), and critical micelle concentration (CMC), attaching an l-amino acid at the C-terminal of a d-tripeptide results in the crescent-shaped supramolecular assemblies. Structure-activity relationship suggests that the crescent-shaped morphology is critical for interacting with membranes and for controlling cell fate. Moreover, fluorescent imaging indicates that the assemblies accumulate on the ER. Time-dependent Western blot and ELISA indicate that the accumulation causes ER stress and subsequently activates the caspase signaling cascade for cell death. As an approach for in situ generating membrane binding scaffolds (i.e., the crescent-shaped supramolecular assemblies), this work promises a new way to disrupt the membrane and to target the ER for developing anticancer therapeutics.

Annonacin promotes selective cancer cell death via NKA-dependent and SERCA-dependent pathways

In the healthcare sector, phytocompounds are known to be beneficial by contributing or alleviating a variety of diseases. Studies have demonstrated the progressive effects of phytocompounds on immune-related diseases and to exhibit anticancer effects. Graviola tree is an evergreen tree with its extracts (leafs and seeds) been reported having anticancer properties, but the precise target of action is not clear. Using an in silico approach, we predicted that annonacin, an Acetogenin, the active agent found in Graviola leaf extract (GLE) to potentially act as a novel inhibitor of both sodium/potassium (NKA) and sarcoplasmic reticulum (SERCA) ATPase pumps. We were able to validate and confirm the in silico studies by showing that GLE inhibited NKA and SERCA activity in intact cells. In the present study, we also demonstrated the antiproliferative and anticancer effects of GLE in a variety of cancer cell lines with limited toxic effects on non-transformed cells. Moreover, our results revealed that known inhibitors of both NKA and SERCA pumps could also promote cell death in several cancer cell lines. In addition, a mouse xenograft cancer model showed GLE as able to reduce tumor size and progression. Finally, bioprofiling studies indicated a strong correlation between overexpression of both NKA and SERCA gene expression vs. survival rates. Overall, our results demonstrated that GLE can promote selective cancer cell death via inhibiting NKA and SERCA, and thus can be considered as a potential novel treatment for cancer. After molecular analysis of GLE by liquid chromatography–mass spectrometry and ESI–QTOF–MS analysis, it was found that the MS spectrum of the high abundant chromatographic peak purified sample highly consisted of annonacin.

Abstract 3219: Sensitizing hypoxic small cell lung cancer cells to radiation and hydrogen peroxide-producing agents using CuATSM

Abstract Cancer cells have increased steady state levels of reactive oxygen species (ROS; O2•- and H2O2) compared to normal cells. It has been proposed that using redox active agents that further increase ROS levels will result is selective cancer cell death. This metabolic frailty can be targeted using drugs deemed safe for human use, ascorbate (ASC) and disulfiram (DSF), via a mechanism of H2O2 production. CuATSM is a drug being used in clinical trials to treat ALS disease. Imaging studies have shown that CuATSM preferentially concentrates in hypoxic tissues, releasing its Cu after entering cells. Copper (Cu) can participate in oxidation reactions that result in highly toxic hydroxyl radicals. Tumors often have areas of hypoxic tissue that exhibit resistance to ionizing radiation (IR). Our hypothesis is that CuATSM can be used to sensitize hypoxic regions of tumors to IR, ASC and/or DSF. Method: H2O2 flux after addition of ASC or DSF was measured in small cell lung cancer (SCLC) cells DMS53 using the 3-aminotriazole method. The Cu uptake of these cells was measured in normoxia and hypoxia after treatment with CuATSM. DMS53 and DMS273 (SCLC lines) were treated with ASC, CuATSM, and/or DSF+CuSO4 at varying oxygen tensions with and without IR and clonogenic assays were performed. Results: Pharmacologically relevant dosing of DSF and ASC resulted in significantly higher fluxes of H2O2 compared to control. Both DSF and ASC enhanced IR clonogenic cell death in SCLC lines. Treatment with CuATSM resulted in increased intracellular Cu and enhanced cell death from ASC and IR in hypoxic conditions. Conclusion: These observations support the hypothesis that the differences in steady-state level of ROS in small cell lung cancer cells can be exploited to develop effective therapies using ASC and DSF. Furthermore CuATSM can enhance responses in hypoxic tumor tissues to both IR and ASC. (Supported by P50 CA174521, T32 CA078586, P30 CA086862 and Carver Research Program of Excellence in Redox Biology and Medicine.) Citation Format: Sebastian Sciegienka, Samuel Rodman, Ann Tomanek-Chalkley, Kelly C. Falls, Douglas R. Spitz, Melissa A. Fath. Sensitizing hypoxic small cell lung cancer cells to radiation and hydrogen peroxide-producing agents using CuATSM [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3219.

Conceptual frameworks of synthetic lethality in clear cell carcinoma of the ovary.

Targeting non-oncogenes may result in the selective death of cancer cells. Clear cell carcinoma of the ovary (CCC) may exhibit resistance against conventional chemotherapy and is associated with poor prognosis. The aim of the present report was to review synthetic lethality-based therapies for CCC. Previous English-language studies were reviewed to accumulate preclinical and clinical data on targeting synthetic lethal partners. Synthetic lethal interactions have a variety of types, involving components of a backup or parallel pathway with overlapping functions, components encoded by paralogous pairs, subunit components that form heteromeric complexes and components that are arranged in a single linear pathway. A set of candidate gene targets potentially resulting in synthetic lethality have been previously identified. HNF class homeobox, AT-rich interaction domain 1A, ATR serine/threonine kinase, ATM serine/threonine kinase, checkpoint kinase 1 and phosphatase and tensin homolog may be the key partner genes. A variety of loss of function genes in CCC are driver or passenger events and may function as synthetic lethal pairs under replication stress conditions. Further clinical studies will be required to investigate the safety and therapeutic effect of synthetic lethality pairs in CCC tumor types with replication stress.

5-Azacitidine Induces Cell Death in a Tissue Culture of Brachypodium distachyon.

Morphological and histological observations revealed that, at a concentration of 50 µM, 5-azacitidine (5-azaC) totally inhibited the induction of embryogenic masses (EM), while the cultivation of explants (zygotic embryos; ZEs) in the presence of 5 µM of 5-azaC led to the formation of a callus with EM in 10% of the cases. Transmission electron microscopy (TEM) analyzes revealed the presence of the morphological and ultrastructural features that are typical for the vacuolar type of cell death in the callus cells that were treated. A TUNEL assay confirmed the presence of DNA double-strand breaks for the callus cells that had been treated with both 5 and 50 µM 5-azaC concentrations. Analysis of the gene expression of selected cell death markers demonstrated a reduced expression of metacaspase, protein executer 1 (EX1), and thioredoxin (TRX) in the callus cells that had been treated compared to the control culture. The strongest increase in the gene activity was characteristic for glutathione S-transferase (GST). Our studies also included an analysis of the distribution of some arabinogalactan proteins (AGPs) and extensin epitopes, which can be used as markers of cells that are undergoing death in a Brachypodium distachyon tissue culture.

The Amyloid-β Oligomer Hypothesis: Beginning of the Third Decade.

The amyloid-β oligomer (AβO) hypothesis was introduced in 1998. It proposed that the brain damage leading to Alzheimer’s disease (AD) was instigated by soluble, ligand-like AβOs. This hypothesis was based on the discovery that fibril-free synthetic preparations of AβOs were potent CNS neurotoxins that rapidly inhibited long-term potentiation and, with time, caused selective nerve cell death (Lambert et al., 1998). The mechanism was attributed to disrupted signaling involving the tyrosine-protein kinase Fyn, mediated by an unknown toxin receptor. Over 4,000 articles concerning AβOs have been published since then, including more than 400 reviews. AβOs have been shown to accumulate in an AD-dependent manner in human and animal model brain tissue and, experimentally, to impair learning and memory and instigate major facets of AD neuropathology, including tau pathology, synapse deterioration and loss, inflammation, and oxidative damage. As reviewed by Hayden and Teplow in 2013, the AβO hypothesis “has all but supplanted the amyloid cascade.” Despite the emerging understanding of the role played by AβOs in AD pathogenesis, AβOs have not yet received the clinical attention given to amyloid plaques, which have been at the core of major attempts at therapeutics and diagnostics but are no longer regarded as the most pathogenic form of Aβ. However, if the momentum of AβO research continues, particularly efforts to elucidate key aspects of structure, a clear path to a successful disease modifying therapy can be envisioned. Ensuring that lessons learned from recent, late-stage clinical failures are applied appropriately throughout therapeutic development will further enable the likelihood of a successful therapy in the near-term.

Indole-3-Carbinol (I3C) enhances the sensitivity of murine breast adenocarcinoma cells to doxorubicin (DOX) through inhibition of NF-κβ, blocking angiogenesis and regulation of mitochondrial apoptotic pathway

Various epidemiological and preclinical studies have already established the cancer chemopreventive potential of naturally occurring glucosinolate breakdown product Indole-3-Carbinol (I3C) as well as its abilities to induce selective cell death towards malignant cell. Therefore, the objective of the present study is to improve the therapeutic efficacy and prevention of doxorubicin (DOX)-induced toxicity, by the concurrent use of Indole-3-Carbinol (I3C). In this study, I3C was administered (20 mg/kg b.w., p.o.) to breast adenocarcinoma (Ehrlich ascites carcinoma) induced solid tumor bearing mice alone as well as in combination with DOX (5 mg/kg b.w., i.p.) in concomitant and pretreatment schedule. The results showed that concurrent administration of I3C and DOX significantly (P < 0.05) improved therapeutic efficacy as evidenced by reduction of tumor size and enhancement of host survivability. Oral administration of I3C significantly (P < 0.05) inhibited the expression of NF-κβ in both tumor cells and cardiac tissue as well as maximizes the therapeutic outcome in terms of tumor cell killing and toxicity. In addition, I3C sensitized tumor cells to DOX-therapy by down-regulating the expression of anti-apoptotic protein Bcl-2 and by up-regulating molecules like Bax, cytochrome c, caspases, which led to PARP cleavage and apoptosis. Significant inhibition of angiogenesis along with reduction in the serum levels of VEGF-A and MMP-9 further contribute to the sensitization accomplished by I3C. Moreover, we also found that I3C provided additional host survival advantages by attenuated DOX-induced toxicities through modulation of Nrf2/ARE pathway and promoted expression of cytoprotective proteins HO-1, NQO1 and GSTπ in cardiac tissue. In addition, I3C significantly attenuated DOX-induced inflammation by down-regulation of NF-kβ, iNOS, COX-2 and IL-6 in cardiac tissue. Thus, the present study clearly suggested therapeutic benefit of I3C in combination with DOX by augmenting anticancer efficacy and diminishing toxicity to the host.

Development of melittin-based anti-cancer drug for targeting tumor-associated macrophages

Abstract Tumor-associated macrophages (TAMs) are the major component of tumor infiltrating immune cells. Macrophages are broadly categorized as M1 or M2 types and TAMs have been known to express M2-like phenotype. TAMs promote tumor progression and contribute to resistant to chemotherapies. Therefore, M2-like TAMs are potential targets for the treatment of cancers. In this study, we targeted M2-like TAMs using a hybrid peptide MEL-dKLA composed of melittin (MEL) peptide, which binds preferentially to M2-like TAMs and d-KLAKLAKKLAKLAK (dKLA) peptide, which induces apoptotic cell death after cell membrane penetration. We showed that the MEL-dKLA induced selective cell death of M2 macrophages by disrupting mitochondrial membrane in vitro while MEL did not show the correlation with mitochondrial apoptosis. We also showed that MEL-dKLA selectively targeted the M2-like TAMs without affecting other leukocytes such as T cells and dendritic cells in vivo. It resulted in the inhibition of tumor growth rate and tumor weight, and angiogenesis in vivo. Importantly, although both MEL and MEL-dKLA reduced the CD206+ M2-like TAMs in the tumor, only MEL-dKLA induced the apoptosis of CD206+ M2-like TAMs whereas the MEL did not show the cell death. Taken together, our study describes MEL-dKLA can be used as the promising therapeutic agent for targeting M2-like TAMs.

Defective immuno- and thymoproteasome assembly causes severe immunodeficiency

By N-ethyl-N-nitrosourea (ENU) mutagenesis, we generated the mutant mouse line TUB6 that is characterised by severe combined immunodeficiency (SCID) and systemic sterile autoinflammation in homozygotes, and a selective T cell defect in heterozygotes. The causative missense point mutation results in the single amino acid exchange G170W in multicatalytic endopeptidase complex subunit-1 (MECL-1), the β2i-subunit of the immuno- and thymoproteasome. Yeast mutagenesis and crystallographic data suggest that the severe TUB6-phenotype compared to the MECL-1 knockout mouse is caused by structural changes in the C-terminal appendage of β2i that prevent the biogenesis of immuno- and thymoproteasomes. Proteasomes are essential for cell survival, and defective proteasome assembly causes selective death of cells expressing the mutant MECL-1, leading to the severe immunological phenotype. In contrast to the immunosubunits β1i (LMP2) and β5i (LMP7), mutations in the gene encoding MECL-1 have not yet been assigned to human disorders. The TUB6 mutant mouse line exemplifies the involvement of MECL-1 in immunopathogenesis and provides the first mouse model for primary immuno- and thymoproteasome-associated immunodeficiency that may also be relevant in humans.

Probing the interaction interface of the GADD45β/MKK7 and MKK7/DTP3 complexes by chemical cross-linking mass spectrometry

GADD45β is selectively and constitutively expressed in Multiple Myeloma cells, and this expression correlates with an unfavourable clinical outcome. GADD45β physically interacts with the JNK kinase, MKK7, inhibiting its activity to enable the survival of cancer cells. DTP3 is a small peptide inhibitor of the GADD45β/MKK7 complex and is able to restore MKK7/JNK activation, thereby promoting selective cell death of GADD45β-overexpressing cancer cells.Enzymatic MS foot-printing and diazirine-based chemical cross-linking MS (CX-MS) strategies were applied to study the interactions between GADD45β and MKK7 kinase domain (MKK7_KD) and between DTP3 and MKK7_KD. Our data show that the binding between GADD45β and MKK7 largely occurs between GADD45β loop 2 (region 103–117) and the kinase enzymatic pocket. We also show that DTP3 interferes with this GADD45β/MKK7 interaction by contacting the MKK7 peptides, 113–136 and 259–274. Accordingly, an MKK7_KD Δ(101–136) variant lacking Trp135 did not produce a fluorescence quenching effect upon the binding of DTP3. The assessment of the interaction between GADD45β and MKK7 and the elucidation of the recognition surfaces between DTP3 and MKK7 significantly advance the understanding of the mechanism underlying the inhibition of the GADD45β/MKK7 interaction by DTP3 and pave the way to the design of small-molecule DTP3 analogues.

Targeting Pyruvate Kinase M2, β Catenin Signaling by Juglone Silver Nano Framework for Selective Cancer Cell Death

AbstractThe modern biomedical nanotechnology necessitates functionalized nanoparticles for targeted action on diseased cells with excellent physico chemical properties. The present study revealed the potential of fabricating juglone silver nano framework for selective cancer cell death by altering the redox states in the cancer cell and selectively inhibiting its growth and proliferation. Conjugation of Juglone with silver nanoparticle reduced the toxicity against normal cells and specifically only inhibited the proliferation of cancer cells. The mechanism of the selective cytotoxic and apoptotic effect on cancer cell was due to down‐regulation of NADPH oxidase 4, Hypoxia‐inducible factor 1α and subdued Pyruvate kinase M2 by arresting Wnt/β catenin signaling through ROS dependent caspase‐3 pathway. Collectively, our findings epitomized that functionalization of nanoparticle gives up a new perspective for adding‐in and tuning the therapeutic values by altering the redox states in cancer cells. Implications of this study opens up a new horizon in modern biomedicine for tuning therapeutic entities to meet the needs of practical applications in complex biological systems.

Enhanced Anticancer Efficacy by Drug Chemotherapy and Cold Atmospheric Plasma Against Melanoma and Glioblastoma Cell Lines &lt;italic&gt;In Vitro&lt;/italic&gt;

Cold atmospheric plasma (CAP) interacting with tumor cells, and finally, leading to selective cell death has opened new horizons in cancer therapy. Despite significant progress in modern cancer treatment using immunologic and targeted therapies melanoma and glioblastoma (GBM) presenting as brain metastases or primary tumor with poor prognosis are still a therapeutic challenge. The aim of this paper was to evaluate the potential of CAP (indirect treatment) in combination with the chemotherapeutics bleomycin, paclitaxel, and dacarbazine to inhibit proliferation on human melanoma and GBM cells in vitro . Cell lines of mouse melanoma (B16) were exposed for 1, 4, 12, 24, and 48 h, and of human melanoma (A375) and GBM (A172) for 24 h to bleomycin, dacarbazine, and paclitaxel together with plasma activated phosphate buffered saline. Directly after exposure, cell proliferation after single (CAP and chemotherapeutic) or combined (CAP plus chemotherapeutic) treatment was assessed by cytotoxicity assay. Indirect CAP treatment of cell lines of GBM and melanoma was followed by a strong reduction of proliferation most prominent after 24 and 48 h exposure. With all drugs maximal reduction was achieved when drug-CAP combinations were used. Combination with CAP treating B16 melanoma cells reached 98% (48 h, combined with paclitaxel), A375 melanoma cells 82% (24 h, combined with bleomycin), and GBM cells 96% (24 h, combined with paclitaxel). The combination of chemotherapy with CAP as indirect treatment may be beneficial in therapy of melanoma and GBM by enhanced tumor reduction and less drug toxicity of chemotherapeutics.

Cancer terminator viruses (CTV): A better solution for viral-based therapy of cancer.

In principle, viral gene therapy holds significant potential for the therapy of solid cancers. However, this promise has not been fully realized and systemic administration of viruses has not proven as successful as envisioned in the clinical arena. Our research is focused on developing the next generation of efficacious viruses to specifically treat both primary cancers and a major cause of cancer lethality, metastatic tumors (that have spread from a primary site of origin to other areas in the body and are responsible for an estimated 90% of cancer deaths). We have generated a chimeric tropism-modified type 5 and 3 adenovirus that selectively replicates in cancer cells and simultaneously produces a secreted anti-cancer toxic cytokine, melanoma differentiation associated gene-7/Interleukin-24 (mda-7/IL-24), referred to as a Cancer Terminator Virus (CTV) (Ad.5/3-CTV). In preclinical animal models, injection into a primary tumor causes selective cell death and therapeutic activity is also observed in non-injected distant tumors, that is, "bystander anti-tumor activity." To enhance the impact and therapeutic utility of the CTV, we have pioneered an elegant approach in which viruses are encapsulated in microbubbles allowing "stealth delivery" to tumor cells that when treated with focused ultrasound causes viral release killing tumor cells through viral replication, and producing and secreting MDA-7/IL-24, which stimulates the immune system to attack distant cancers, inhibits tumor angiogenesis and directly promotes apoptosis in distant cancer cells. This strategy is called UTMD (ultrasound-targeted microbubble-destruction). This novel CTV and UTMD approach hold significant promise for the effective therapy of primary and disseminated tumors.