Pro-apoptotic Bcl-2 Proteins Research Articles

Effect of Lamotrigine on Refractory Epilepsy: Clinical Outcomes and EEG Changes.

Refractory epilepsy poses significant challenges in clinical management due to its resistance to standard antiepileptic therapies, necessitating the exploration of more effective treatment regimens. Lamotrigine, with its proven efficacy and tolerability, offers potential benefits when combined with traditional medications like valproate, though its comprehensive impact on clinical outcomes and neurological markers requires further study. To analyze the improvement effect of combined application of lamotrigine on refractory epilepsy patients and its impact on patients' EEG and neurological function. This retrospective cohort study analyzed the clinical data of 93 patients with refractory epilepsy who were admitted to our hospital between January 2023 and June 2024. Based on the treatment interventions received, patients were divided into a control group (n=46, treated with valproate) and an observation group (n=47, treated with lamotrigine in addition to valproate). The clinical treatment effects, EEG (δ, θ, α, β) power levels, neurological function indicators [brain-derived neurotrophic factor (BDNF), nerve growth factor (NGF), pro-apoptotic protein Bcl-2, Bax], inflammatory response indicators [interleukin-1β (IL-1β), interleukin-6 (IL-6), prostaglandin E2 (PGE2)], and the incidence of adverse reactions were compared between the two groups. The clinical treatment effect in the observation group was significantly better than that in the control group, with a higher total effective rate (93.62% vs 76.09%, P<0.05). The monthly seizure frequency was significantly reduced in both groups after treatment (P < 0.05). The observation group demonstrated a significantly greater reduction in seizure frequency compared to the control group (P = 0.014). Regarding EEG power levels, both groups showed decreases in δ and θ power levels and increases in α and β power levels after treatment, with the observation group exhibiting more pronounced changes (P<0.05). Neurological function indicators revealed that Bcl-2 levels decreased, while BDNF, NGF, and Bax levels increased in both groups after treatment, with the observation group showing more significant improvements (P<0.05). Similarly, inflammatory response indicators, including IL-1β, IL-6, and PGE2, decreased in both groups, with the observation group demonstrating greater reductions (P<0.05). The incidence of adverse reactions was comparable between the two groups, with no significant difference observed (23.40% vs 17.39%, P>0.05). Compared to valproate treatment alone, the combined application of lamotrigine can further enhance the efficacy in refractory epilepsy patients, Lower the seizure frequency, improve EEG power levels and neurological function, reduce inflammatory responses, and does not increase the risk of related adverse reactions.

Chaetocin inhibits the progression of neuroblastoma by targeting JAK2/STAT3 signaling pathway in SH-SY5Y cells.

Chaetocin is a fungal mycotoxin that extensively found in various natural products and has anticancer and anti‑inflammatory activities. Herein, the anticancer effects of chaetocin against the progression of neuroblastoma were studied with SHSY-5Y human neuroblastoma cells and examined the underlying molecular mechanisms. The effects of chaetocin on cellular viability, apoptosis, cell migration, and invasion were investigated. The underlying mechanism of anticancer effects of chaetocin was found to mediate via activating JAK2/STAT3 signaling pathway. Furthermore, when SHSY-5Y cells were exposed to a higher concentration of chaetocin, the induction of cell apoptosis significantly increased by enhancing the expression of pro-apoptotic protein Bcl-2, resulting in anticancer activity against neuroblastoma. In addition, chaetocin significantly decreased the SHSY-5Y cell invasion and migration at 50μM treatment. Moreover, it was shown that increasing chaetocin treatments greatly decreased the activity of proteins connected to the JAK2/STAT3 signaling pathway. In conclusion, chaetocin exhibits a diverse range of actions on neuroblastoma cells, including the inhibition of proliferation, induction of apoptosis, perturbation of cellular morphology, and modulation of critical signaling pathways, with a specific focus on the JAK/STAT3 pathway. These results contribute valuable insights that underscore the potential therapeutic utility of chaetocin in the context of neuroblastoma treatment, suggesting its multifaceted impact on key cellular processes involved in cancer progression.

Oral Administration of Carotenoid-Rich Dunaliella salina Powder Inhibits Colon Carcinogenesis via Modulation of Wnt/β-catenin Signaling Cascades in a Rat Model.

The present study aims to investigate the oral therapeutic and molecular role of carotenoid-rich Dunaliella salina powder (DSP) against 1,2-dimethylhydrazine (DMH)-triggered colon carcinogenesis. In this study, thirty six male Wistar rats were categorized into six distinct groups (G1-G6): G1 group with no intervention, G2 group received only DSP (1000mg/kg), G3 group received only DMH carcinogen (20mg/kg), and G4-G6 group received both DMH and DSP at various phases (pre-initiation, post-initiation and entire phases) for 32weeks. Body weight, tumor incidence, tumor volume, histopathological examination, antioxidants, and detoxification enzymes activities were analyzed in the experimental rats. In addition, the protein expression profile of components involved in the Wnt/β-catenin signaling pathway was determined by western blot analysis. Matrix metalloproteinases (MMP-7 and MMP-9), proliferation marker (PCNA), and pro-apoptotic (Bcl-2 and Bax) proteins were analyzed using immunohistochemistry. Colorimetric assay was used to determine the levels of anti-inflammatory (iNOS and COX-2) and apoptotic proteins (Caspase-3 and Caspase-9). Results showed that concomitant administration of DSP with DMH significantly reduced tumor progression and prevented colon carcinogenesis in rats. However, treatment with DSP before or after DMH exposure did not significantly prevent colon carcinogenesis. DMH and DSP treatment group showed increased activities of antioxidant enzymes with significant reduction in the oxidative stress. Additionally, the detoxification enzymes and colonic histopathology of those rats were restored to that of control rats. The administration of DSP to rats exposed to DMH exhibited antitumor effects via inhibition of the Wnt/β-catenin signaling pathway with induced apoptosis through the Bcl-2/Bax/caspases signaling cascades. Moreover, the same group also showed significant anti-inflammatory activity via regulating iNOS and COX-2 biomarkers. Our findings revealed molecular chemopreventive activity of carotenoid-rich DSP through regulating Wnt/beta-catenin and intrinsic apoptotic pathways. Thus, DSP is propound to function as a potent antioxidant, anti-proliferative, and anti-inflammatory therapeutic agent against colon carcinogenesis.

BCL-2 and BOK regulate apoptosis by interaction of their C-terminal transmembrane domains

The Bcl-2 family controls apoptosis by direct interactions of pro- and anti-apoptotic proteins. The principle mechanism is binding of the BH3 domain of pro-apoptotic proteins to the hydrophobic groove of anti-apoptotic siblings, which is therapeutically exploited by approved BH3-mimetic anti-cancer drugs. Evidence suggests that also the transmembrane domain (TMD) of Bcl-2 proteins can mediate Bcl-2 interactions. We developed a highly-specific split luciferase assay enabling the analysis of TMD interactions of pore-forming apoptosis effectors BAX, BAK, and BOK with anti-apoptotic Bcl-2 proteins in living cells. We confirm homotypic interaction of the BAX-TMD, but also newly identify interaction of the TMD of anti-apoptotic BCL-2 with the TMD of BOK, a peculiar pro-apoptotic Bcl-2 protein. BOK-TMD and BCL-2-TMD interact at the endoplasmic reticulum. Molecular dynamics simulations confirm dynamic BOK-TMD and BCL-2-TMD dimers and stable heterotetramers. Mutation of BCL-2-TMD at predicted key residues abolishes interaction with BOK-TMD. Also, inhibition of BOK-induced apoptosis by BCL-2 depends specifically on their TMDs. Thus, TMDs of Bcl-2 proteins are a relevant interaction interface for apoptosis regulation and provide a novel potential drug target.

Synergistic Effect of Doxorubicin and Blue Light Irradiation on the Antitumor Treatment of HepG2 Cells in Liver Cancer.

Doxorubicin (DOX) has been an effective antitumor agent for human liver cancer cells; however, an overdose might lead to major side effects appearing in clinical applications. In this work, we present a strategy of combining DOX and blue light (BL) irradiation for the antitumor treatment of HepG2 cells (one typical human liver cancer cell line). It is demonstrated that synergetic DOX and BL can significantly reduce cell proliferation and increase the apoptotic rate of HepG2 cells in comparison to individual DOX treatment. The additional BL irradiation is further helpful for enhancing the inhibition of cell migration and invasion. Analyses of reactive oxygen species (ROS) level and Western blotting reveal that the strategy results in more ROS accumulation, mitochondrial damage, and the upregulation of proapoptotic protein (Bcl-2) and downregulation of antiapoptotic protein (Bax). In addition to the improved therapeutic effect, the non-contact BL irradiation is greatly helpful for reducing the dosage of DOX, and subsequently reduces the side effects caused by the DOX drug. These findings offer a novel perspective for the therapeutic approach toward liver cancer with high efficiency and reduced side effects.

Abstract 565: PDE7 modulates EMT, cells morphology and mitochondrial cristae in high grade serous ovarian cancer

Abstract Background: Due to the acquisition of chemoresistant phenotype by epithelial ovarian cancer (EOC) cells and, consequently, disease relapse, the identification of novel therapeutic strategies is urgent. Herein, we targeted phosphodiesterase 7A (PDE7A) in EOC cells, based on data derived from RNA-sequencing assays comparing high-grade serous ovarian carcinoma (HGSOC) and fallopian tube samples. Methods: To understand the role of PDE inhibition, we evaluated the effects of its inhibitor BRL50481 alone or in combination with paclitaxel (PTX) on untreated A2780 EOC cells and multi-resistant high grade serous ovarian cancer (HGSOC) OVCAR3 cells. To do so, we pursued with the: validation of data obtained by high throughput RNA sequencing assays using qRT-PCR; evaluation of PDE7 expression in normal and EOC cells, as well as in normal Fallopian tube cells by qRT-PCR; immunoblotting; metabolic cell viability (MCV) by MTT assays; cytokine dosage by qRT-PCR and ELISA; scanning electron microscopy; transmission electron microscopy; statistical analysis (GraphPad software). Results: Drug-sensitive A2780 cells, but not OVCAR3 cells, exhibited lower MCV when treated with the PDE7 inhibitor; however, the combinatory therapy decreased MCV in both cell lines. Of clinical relevance, pre-treatment of A2780 and OVCAR3 with the PDE7 inhibitor BRL50481 followed by PTX lowered its IC50 by 103- and 625-fold, respectively. While the phosphatidylinositol 3-kinase (PI3K)/protein kinase B(AKT)/mammalian target of rapamycin (mTOR) pathway was inhibited in both cell lines, the pro-apoptotic protein Bcl-2 Associated X-protein (BAX) was overexpressed in A2780 cells only. Analysis of mRNA and protein expression in OVCAR3 pretreated with BRL50481 showed a 2-fold increase in IL-6 cytokine expression. Pre-treatment of OVCAR3 with BRL50481 followed by the combination of BRL50481 and PTX inhibited the expression of vimentin and Octamer-binding transcription factor 4 (OCT4) by 2-fold. BRL50481 promoted changes in EOC cells morphology and mitochondrial cristae. Conclusions: Altogether, our data led us to postulate that inhibition of PDE7 in EOC cells, specifically in those presenting the chemoresistant phenotype, is a promising strategy to fight this yet highly lethal disease. Citation Format: Leticia B. A. Rangel, Nayara Gusmão Tessarollo, Isabella dos Santos Guimarães, Diandra Zipinotti dos Santos, Taciane Barbosa Henriques, Marcele Lorentz de Souza, Leide Laura Figueiredo Maciel, João Carlos Aquino Almeida, Tatiana Massariol Pimenta, Josiany Carlos de Souza, Ranna Batista Wanzeler, Barbara Silva Martins, Solenny Maria Silva Butzene, Jose Matheus Simões Padilha. PDE7 modulates EMT, cells morphology and mitochondrial cristae in high grade serous ovarian cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 565.

Melatonin enhances the effect of ABT-737 in acute monocytic leukemia THP-1 cells

Melatonin (N-acetyl-5-methoxytryptamine, MEL) is a hormone synthesized by the pineal gland. Due to its oncostatic effect, it can be considered as an antitumor agent and used for combination therapy. ABT-737, a Bcl-2 inhibitor, promotes cell death after treatment with agents that induce pro-apoptotic signals. In the present study, the combined effect of MEL and ABT-737 on changes in proliferative and mitotic activity, mitochondrial membrane potential, intracellular production of reactive oxygen species (ROS) and cytosolic Ca2+ was studied. Moreover, changes in the expression of anti- and pro-apoptotic proteins (Bcl-2 and Bax), autophagy markers (LC3A/B (I, II)), endoplasmic reticulum stress markers (chaperones BIP and PDI, CHOP) were studied under these conditions. The effect of MEL together with ABT-737 led to an increase in the level of cytosolic Ca2+, intracellular production of ROS, and a decrease in the membrane potential of mitochondria. Under these conditions, the content of Bcl-2 increased, while the level of Bax decreased. The activation of CHOP stimulated autophagy and led to a decrease in the expression of BIP and PDI chaperones. These results suggest that MEL is able to enhance the effects of other chemotherapeutic agents and can be used in strategies in the treatment of cancer.

A comprehensive apoptotic assessment of niloticin in cervical cancer cells: a tirucallane-type triterpenoid from Aphanamixis polystachya (Wall.) Parker.

Pharmacologically active small organic molecules derived from natural resources are prominent drug candidates due to their inherent structural diversity. Herein, we explored one such bioactive molecule, niloticin, which is a tirucallane-type triterpenoid isolated from the stem barks of Aphanamixis polystachya (Wall.) Parker. After initial screening with other isolated compounds from the same plant, niloticin demonstrated selective cytotoxicity against cervical cancer cells (HeLa) with an IC50 value of 11.64 μM. Whereas the compound exhibited minimal cytotoxicity in normal epithelial cell line MCF-10A, with an IC50 value of 83.31 μM. Subsequently, in silico molecular docking studies of niloticin based on key apoptotic proteins such as p53, Fas, FasL, and TNF β revealed striking binding affinity, reflecting docking scores of -7.2, -7.1, -6.8, and -7.2. Thus, the binding stability was evaluated through molecular dynamic simulation. In a downstream process, the apoptotic capability of niloticin was effectively validated through in vitro fluorimetric assays, encompassing nuclear fragmentation. Additionally, an insightful approach involving surface-enhanced Raman spectroscopy (SERS) re-establishes the occurrence of DNA cleavage during cellular apoptosis. Furthermore, niloticin was observed to induce apoptosis through both intrinsic and extrinsic pathways. This was evidenced by the upregulation of upstream regulatory molecules such as CD40 and TNF, which facilitate the activation of caspase 8. Concurrently, niloticin-induced p53 activation augmented the expression of proapoptotic proteins Bax and Bcl-2 and downregulation of IAPs, leading to the release of cytochrome C and subsequent activation of caspase 9. Therefore, the reflection of mitochondrial-mediated apoptosis is in good agreement with molecular docking studies. Furthermore, the anti-metastatic potential was evidenced by wound area closure and Ki67 expression patterns. This pivotal in vitro assessment confirms the possibility of niloticin being a potent anti-cancer drug candidate, and to the best of our knowledge, this is the first comprehensive anticancer assessment of niloticin in HeLa cells.

Small molecule Mcl-1 inhibitor for triple negative breast cancer therapy.

Apoptosis is an evolutionarily conserved cell death pathway that plays a crucial role in maintaining tissue homeostasis, orchestrating organismal development, and eliminating damaged cells. Dysregulation of apoptosis can contribute to the pathogenesis of malignant tumors and neurodegenerative diseases. Anticancer drugs typically possess the capacity to induce apoptosis in tumor cells. The Bcl-2 protein family, consisting of 27 members in humans, serves as the key regulator of mitochondrial function. This family can be divided into two functional groups: anti-apoptotic proteins (e.g., Bcl-2, Bcl-xl, Mcl-1) and pro-apoptotic proteins (e.g., Bad, Bax). Mcl-1 exerts its function by binding pro-apoptotic Bcl-2 proteins thereby preventing apoptosis induction. Overexpression of Mcl-1 not only correlates closely with tumorigenesis but also associates significantly with resistance towards targeted therapy and conventional chemotherapy. Effective induction of apoptosis can be achieved through inhibition or interference with Mcl-1. Thus, this mini review discusses existing Mcl-1 inhibitors.

The role of apoptosis and oxidative stress in the pathophysiology of Acanthamoeba spp. infection in the kidneys of hosts with different immunological status

BackgroundAcanthamoeba spp. are opportunistic pathogens that cause inflammation, mostly in the brain, lungs and cornea. Recent reports indicate kidney dysfunction in hosts with systemic acanthamoebiasis. The aim of the study was to analyze the gene expression and protein concentration of NADPH oxidase 2 and 4 (NOX2 and NOX4, respectively) and nuclear erythroid 2-related factor (Nrf2) in the kidneys of hosts with systemic acanthamoebiasis. We also aimed to determine the protein and gene expressions of Bcl2, Bax, caspases 3 and 9.Methods Mice were divided into four groups based on their immunological status and Acanthamoeba sp. infection: A, immunocompetent Acanthamoeba sp.-infected mice; AS, immunosuppressed Acanthamoeba sp.- infected mice; C, immunocompetent uninfected mice; CS, immunosuppressed uninfected mice. NOX2, NOX4 and Nrf2 were analyzed by quantitative reverse transcription PCR (qRT-PCR) and ELISA methods, while pro-apoptotic and anti-apoptotic proteins (Bax and Bcl-2, respectively), Cas9, Cas3 were analyzed by qRT-PCR and western blot methods. ResultsIncreased gene expression and/or protein concentration of NOX2 and NOX4 were found in both immunocompetent and immunosuppressed mice infected with Acanthamoeba sp. (groups A and AS, respectively). Gene expression and/or protein concentration of Nrf2 were higher in group A than in control animals. Compared to control mice, in the AS group the expression of the Nrf2 gene was upregulated while the concentration of Nrf2 protein was decreased. Additionally in A group, higher gene and protein expression of Bcl-2, and lower gene as well as protein expression of Bax, caspases 3 and 9 were noted. In contrast, the AS group showed lower gene and protein expression of Bcl-2, and higher gene as well as protein expression of Bax, caspases 3 and 9.ConclusionsThis study is the first to address the mechanisms occurring in the kidneys of hosts infected with Acanthamoeba sp. The contact of Acanthamoeba sp. with the host cell surface and/or the oxidative burst caused by elevated levels of NOXs lead to an antioxidant response enhanced by the Nrf2 pathway. Acanthamoeba sp. have various strategies concerning apoptosis. In immunocompetent hosts, amoebae inhibit the apoptosis of kidney cells, and in immunosuppressed hosts, they lead to increased apoptosis by the intrinsic pathway and thus to a more severe course of the disease.Graphical

Simplified Synthesis of Renieramycin T Derivatives to Target Cancer Stem Cells via β-Catenin Proteasomal Degradation in Human Lung Cancer.

Cancer stem cells (CSCs) found within cancer tissue play a pivotal role in its resistance to therapy and its potential to metastasize, contributing to elevated mortality rates among patients. Significant strides in understanding the molecular foundations of CSCs have led to preclinical investigations and clinical trials focused on CSC regulator β-catenin signaling targeted interventions in malignancies. As part of the ongoing advancements in marine-organism-derived compound development, it was observed that among the six analogs of Renieramycin T (RT), a potential lead alkaloid from the blue sponge Xestospongia sp., the compound DH_32, displayed the most robust anti-cancer activity in lung cancer A549, H23, and H292 cells. In various lung cancer cell lines, DH_32 exhibited the highest efficacy, with IC50 values of 4.06 ± 0.24 μM, 2.07 ± 0.11 μM, and 1.46 ± 0.06 μM in A549, H23, and H292 cells, respectively. In contrast, parental RT compounds had IC50 values of 5.76 ± 0.23 μM, 2.93 ± 0.07 μM, and 1.52 ± 0.05 μM in the same order. Furthermore, at a dosage of 25 nM, DH_32 showed a stronger ability to inhibit colony formation compared to the lead compound, RT. DH_32 was capable of inducing apoptosis in lung cancer cells, as demonstrated by increased PARP cleavage and reduced levels of the proapoptotic protein Bcl2. Our discovery confirms that DH_32 treatment of lung cancer cells led to a reduced level of CD133, which is associated with the suppression of stem-cell-related transcription factors like OCT4. Moreover, DH_32 significantly suppressed the ability of tumor spheroids to form compared to the original RT compound. Additionally, DH_32 inhibited CSCs by promoting the degradation of β-catenin through ubiquitin-proteasomal pathways. In computational molecular docking, a high-affinity interaction was observed between DH_32 (grid score = -35.559 kcal/mol) and β-catenin, indicating a stronger binding interaction compared to the reference compound R9Q (grid score = -29.044 kcal/mol). In summary, DH_32, a newly developed derivative of the right-half analog of RT, effectively inhibited the initiation of lung cancer spheroids and the self-renewal of lung cancer cells through the upstream process of β-catenin ubiquitin-proteasomal degradation.

Monkeypox virus C7L is a Bcl-2-fold protein that binds human pro-apoptotic Bcl-2 proteins

Monkeypox virus C7L is a Bcl-2-fold protein that binds human pro-apoptotic Bcl-2 proteins

Astragaloside IV against Alzheimer's disease via microglia-mediated neuroinflammation using network pharmacology and experimental validation

Alzheimer's disease (AD) is one of the most prevalent neurodegenerative diseases in the world. The effective therapeutic methods and drugs are still not clear. Astragaloside IV (AS-IV), a triterpenoid saponin isolated from the root of Huangqi, has a beneficial effect in the treatment of AD. However, whether AS-IV alters microglia in the inflammation of AD is still ambiguous. In our study, 99 common targets were collected between AS-IV and AD. BCL2 apoptosis regulator (Bcl-2), pro-apoptotic BCL-2 protein BAX, epidermal growth factor receptor (EGFR), and receptor tyrosine phosphatase type C (PTPRC) were screened for inflammation and microglia in the above targets by network pharmacology. Interleukin-1β (IL-1β) and EGFR both interact with signal transducer and activator of transcription 3 (STAT3) by a protein interaction network, and IL-1β had a higher affinity for AS-IV based on molecular docking. Enrichment revealed targets involved in the regulation of neuronal cell bodies, growth factor receptor binding, EGFR tyrosine kinase inhibitor resistance., etc. Besides, AS-IV alleviated the reduced cell proliferation in amyloid-beta (Aβ)-treated microglial BV2 cells. AS-IV affected BV2 cell morphological changes and decreased cluster of differentiation 11b (CD11b) gene, IL-1β, and EGFR mRNA levels increment during lipopolysaccharide (LPS) injury in BV2 cell activation. Therefore, AS-IV may regulate microglial activation and inflammation via EGFR-dependent pathways in AD. EGFR and IL-1β are vital targets that may relate to each other to coregulate downstream molecular functions in the cure of AD. Our study provides a candidate drug and disease target for the treatment of neurodegenerative diseases in the clinic.

Amsacrine downregulates BCL2L1 expression and triggers apoptosis in human chronic myeloid leukemia cells through the SIDT2/NOX4/ERK/HuR pathway

Accumulating evidence indicates that the anticancer activity of acridine derivatives is mediated through the regulation of anti-apoptotic and pro-apoptotic BCL2 protein expression. Therefore, we investigated whether the cytotoxicity of amsacrine with an acridine structural scaffold in human chronic myeloid leukemia (CML) K562 cells was mediated by BCL2 family proteins. Amsacrine induced apoptosis, mitochondrial depolarization, and BCL2L1 (also known as BCL-XL) downregulation in K562 cells. BCL2L1 overexpression inhibited amsacrine-induced cell death and mitochondrial depolarization. Amsacrine treatment triggered SIDT2-mediated miR-25 downregulation, leading to increased NOX4-mediated ROS production. ROS-mediated inactivation of ERK triggered miR-22 expression, leading to increased HuR mRNA decay. As HuR is involved in stabilizing BCL2L1 mRNA, downregulation of BCL2L1 was noted in K562 cells after amsacrine treatment. In contrast, amsacrine-induced BCL2L1 downregulation was alleviated by restoring ERK phosphorylation and HuR expression. Altogether, the results of this study suggest that amsacrine triggers apoptosis in K562 cells by inhibiting BCL2L1 expression through the SIDT2/NOX4/ERK-mediated downregulation of HuR. Furthermore, a similar pathway also explains the cytotoxicity of amsacrine in CML MEG-01 and KU812 cells.

Molecular docking analysis of MCL-1 inhibitors for breast cancer management.

Myeloid leukemia 1 (MCL-1), a BCL-2 protein family member, acts as an anti-apoptotic protein by interacting with pro-apoptotic BCL-2 proteins. Its overexpression is frequently observed in numerous cancer types including breast cancer, and is closely linked to the initiation and progression of tumors as well as poor prognosis and resistance to therapeutic interventions. Here, a database of 3402 chemicals with established therapeutic activity against various diseases was chosen and systematically screened against the MCL-1 protein. Visual inspection and binding energy analysis revealed that the compounds OSU-03012, Raltitrexed, Ostarine (MK-2866), Dovitinib (TKI-258), and Varespladib (LY315920) had strong binding affinity for the MCL-1 protein. Notably, their binding affinity was higher than that of the control compounds. These compounds exhibited strong interactions with critical amino acid residues of the MCL-1 protein. Furthermore, these compounds shared several common amino acid residue interactions with the control compounds. These findings suggest that these compounds may be useful as MCL-1 inhibitors in the treatment of breast cancer. However, additional experimental validation is required to confirm these findings.

Independent role of caspases and Bik in augmenting influenza A virus replication in airway epithelial cells and mice

Caspases and poly (ADP-ribose) polymerase 1 (PARP1) have been shown to promote influenza A virus (IAV) replication. However, the relative importance and molecular mechanisms of specific caspases and their downstream substrate PARP1 in regulating viral replication in airway epithelial cells (AECs) remains incompletely elucidated. Here, we targeted caspase 2, 3, 6, and PARP1 using specific inhibitors to compare their role in promoting IAV replication. Inhibition of each of these proteins caused significant decline in viral titer, although PARP1 inhibitor led to the most robust reduction of viral replication. We previously showed that the pro-apoptotic protein Bcl-2 interacting killer (Bik) promotes IAV replication in the AECs by activating caspase 3. In this study, we found that as compared with AECs from wild-type mice, bik-deficiency alone resulted in ~ 3 logs reduction in virus titer in the absence of treatment with the pan-caspase inhibitor (Q-VD-Oph). Inhibiting overall caspase activity using Q-VD-Oph caused additional decline in viral titer by ~ 1 log in bik-/- AECs. Similarly, mice treated with Q-VD-Oph were protected from IAV-induced lung inflammation and lethality. Inhibiting caspase activity diminished nucleo-cytoplasmic transport of viral nucleoprotein (NP) and cleavage of viral hemagglutinin and NP in human AECs. These findings suggest that caspases and PARP1 play major roles to independently promote IAV replication and that additional mechanism(s) independent of caspases and PARP1 may be involved in Bik-mediated IAV replication. Further, peptides or inhibitors that target and block multiple caspases or PARP1 may be effective treatment targets for influenza infection.

Enhanced Apoptosis and Loss of Cell Viability in Melanoma Cells by Combined Inhibition of ERK and Mcl-1 Is Related to Loss of Mitochondrial Membrane Potential, Caspase Activation and Upregulation of Proapoptotic Bcl-2 Proteins.

Targeting of MAP kinase pathways by BRAF inhibitors has evolved as a key therapy for BRAF-mutated melanoma. However, it cannot be applied for BRAF-WT melanoma, and also, in BRAF-mutated melanoma, tumor relapse often follows after an initial phase of tumor regression. Inhibition of MAP kinase pathways downstream at ERK1/2, or inhibitors of antiapoptotic Bcl-2 proteins, such as Mcl-1, may serve as alternative strategies. As shown here, the BRAF inhibitor vemurafenib and the ERK inhibitor SCH772984 showed only limited efficacy in melanoma cell lines, when applied alone. However, in combination with the Mcl-1 inhibitor S63845, the effects of vemurafenib were strongly enhanced in BRAF-mutated cell lines, and the effects of SCH772984 were enhanced in both BRAF-mutated and BRAF-WT cells. This resulted in up to 90% loss of cell viability and cell proliferation, as well as in induction of apoptosis in up to 60% of cells. The combination of SCH772984/S63845 resulted in caspase activation, processing of poly (ADP-ribose) polymerase (PARP), phosphorylation of histone H2AX, loss of mitochondrial membrane potential, and cytochrome c release. Proving the critical role of caspases, a pan-caspase inhibitor suppressed apoptosis induction, as well as loss of cell viability. As concerning Bcl-2 family proteins, SCH772984 enhanced expression of the proapoptotic Bim and Puma, as well as decreased phosphorylation of Bad. The combination finally resulted in downregulation of antiapoptotic Bcl-2 and enhanced expression of the proapoptotic Noxa. In conclusion, combined inhibition of ERK and Mcl-1 revealed an impressive efficacy both in BRAF-mutated and WT melanoma cells, and may thus represent a new strategy for overcoming drug resistance.

P53-Bad* Fusion Gene Therapy Induces Apoptosis In Vitro and Reduces Zebrafish Tumor Burden in Hepatocellular Carcinoma.

With few curative treatments and a global yearly death rate of over 800,000, hepatocellular carcinoma (HCC) desperately needs new therapies. Although wild-type p53 gene therapy has been shown to be safe in HCC patients, it has not shown enough efficacy to merit approval. This work aims to show how p53 can be re-engineered through fusion to the pro-apoptotic BH3 protein Bcl-2 antagonist of cell death (Bad) to improve anti-HCC activity and potentially lead to a novel HCC therapeutic, p53-Bad*. p53-Bad* is a fusion of p53 and Bad, with two mutations, S112A and S136A. We determined mitochondrial localization of p53-Bad* in liver cancer cell lines with varying p53 mutation statuses via fluorescence microscopy. We defined the apoptotic activity of p53-Bad* in four liver cancer cell lines using flow cytometry. To determine the effects of p53-Bad* in vivo, we generated and analyzed transgenic zebrafish expressing hepatocyte-specific p53-Bad*. p53-Bad* localized to the mitochondria regardless of the p53 mutation status and demonstrated superior apoptotic activity over WT p53 in early, middle, and late apoptosis assays. Tumor burden in zebrafish HCC was reduced by p53-Bad* as measured by the liver-to-body mass ratio and histopathology. p53-Bad* induced significant apoptosis in zebrafish HCC as measured by TUNEL staining but did not induce apoptosis in non-HCC fish. p53-Bad* can induce apoptosis in a panel of liver cancer cell lines with varying p53 mutation statuses and induce apoptosis/reduce HCC tumor burden in vivo in zebrafish. p53-Bad* warrants further investigation as a potential new HCC therapeutic.

L-Theanine alleviates MPTP-induced Parkinson's disease by targeting Wnt/β-catenin signaling mediated by the MAPK signaling pathway

We evaluated the neuroprotective effect of L-theanine in Parkinson's disease and the underlying mechanism focusing on WNT/β-catenin signaling mediated by the MAPK pathway. We treated MPTP-induced SH-SY5Y cells with various concentrations of L-theanine (50, 100, 200, and 500 μg/mL), and we also treated Parkinson's model mice with L-theanine. L-theanine treatment effectively reduced the immunohistochemical hallmarks of Parkinson's disease, particularly Lewy bodies and α-synuclein, and increased the number of tyrosine hydroxylase-positive cells. L-theanine also improved the motor dysfunction in MPTP-induced Parkinson's disease model mice as measured by the rotarod test. The levels of several pro-inflammatory mediators that are overexpressed in Parkinson's disease, namely TNF-α, IL-6, COX-2, and MAC-1, were reduced following L-theanine treatment, and the levels of the pro-apoptotic proteins Bcl-2, caspase-3, p53, and PARP-1 were significantly reduced. L-theanine regulated the oxidative stress-related factors SOD-1, GST, and NOX-4 by targeting several proteins related to WNT/β-catenin signaling, i.e., β-catenin, WNT-3a, WNT-5a, TCF1/TCF7, and LEF1, via the MAPK pathway (p-JNK, p-ERK, and p-p38). Our results indicate that L-theanine is neuroprotective and has anti-inflammatory effects that could be beneficial for treating Parkinson's disease.

Effect of intraperitoneal docetaxel on ovarian function in mice

Taxanes are important chemotherapeutic agents used to manage breast cancer and gynaecological malignancies. However, ovarian toxicity induced by the taxane docetaxel (DOC) is of great concern. We investigated DOC-induced toxicity in the ovaries of female CD1 strain mice. The mice were divided into control (saline), DOC-5 (5 mg/kg DOC), and DOC-10 (10 mg/kg DOC) groups and administered saline or DOC on the first day of the study and two weeks later. Two weeks after the second dose, the ovaries were removed for analysis after inducing superovulation. Ovary weight, the number of secondary follicles, and the total number of follicles were reduced after DOC administration. Additionally, the expression levels of caspase-3 and the pro-apoptotic protein Bcl-2 interacting mediator of cell death (BIM) increased. Our findings suggest that high-dose DOC induces damage to growing follicles; however, it may not affect primordial follicles. Impact statement What is already known on this subject? Docetaxel (DOC) is one of the most effective chemotherapeutic agents used to manage various cancers. Some in-vitro studies have examined paclitaxel-induced ovarian toxicity; however, limited research on DOC is available. What do the results of this study add? We investigated DOC-induced ovarian toxicity in female CD1 strain mice at 5 mg/kg and 10 mg/kg. We found that DOC reduced ovary weight, the number of secondary follicles, and the total number of follicles, with the higher dose having a higher effect. What are the implications of these findings for clinical practice and/or further research? We believe that our study makes a significant contribution to the knowledge about the effect of DOC on ovarian function.