-
Journal of Virology Nov 2021Targeting host factors is a promising strategy to develop broad-spectrum antiviral drugs. Drugs targeting anti-apoptotic Bcl-2 family proteins that were originally...
Targeting host factors is a promising strategy to develop broad-spectrum antiviral drugs. Drugs targeting anti-apoptotic Bcl-2 family proteins that were originally developed as tumor suppressors have been reported to inhibit multiplication of different types of viruses. However, the mechanisms whereby Bcl-2 inhibitors exert their antiviral activity remain poorly understood. In this study, we have investigated the mechanisms by which obatoclax (OLX) and ABT-737 Bcl-2 inhibitors exhibited a potent antiviral activity against the mammarenavirus lymphocytic choriomeningitis virus (LCMV). OLX and ABT-737 potent anti-LCMV activity was not associated with their proapoptotic properties but rather with their ability to induce cell arrest at the G0/G1 phase. OLX- and ABT-737-mediated inhibition of Bcl-2 correlated with reduced expression levels of thymidine kinase 1 (TK1), cyclin A2 (CCNA2), and cyclin B1 (CCNB1) cell cycle regulators. In addition, small interfering RNA (siRNA)-mediated knockdown of TK1, CCNA2, and CCNB1 resulted in reduced levels of LCMV multiplication. The antiviral activity exerted by Bcl-2 inhibitors correlated with reduced levels of viral RNA synthesis at early times of infection. Importantly, ABT-737 exhibited moderate efficacy in a mouse model of LCMV infection, and Bcl-2 inhibitors displayed broad-spectrum antiviral activities against different mammarenaviruses and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Our results suggest that Bcl-2 inhibitors, actively being explored as anticancer therapeutics, might be repositioned as broad-spectrum antivirals. Antiapoptotic Bcl-2 inhibitors have been shown to exert potent antiviral activities against various types of viruses via mechanisms that are currently poorly understood. This study has revealed that Bcl-2 inhibitors' mediation of cell cycle arrest at the G0/G1 phase, rather than their proapoptotic activity, plays a critical role in blocking mammarenavirus multiplication in cultured cells. In addition, we show that Bcl-2 inhibitor ABT-737 exhibited moderate antimammarenavirus activity and that Bcl-2 inhibitors displayed broad-spectrum antiviral activities against different mammarenaviruses and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Our results suggest that Bcl-2 inhibitors, actively being explored as anticancer therapeutics, might be repositioned as broad-spectrum antivirals.
Topics: A549 Cells; Animals; Antiviral Agents; Apoptosis; Apoptosis Regulatory Proteins; Arenaviridae; Biphenyl Compounds; COVID-19; Cell Cycle; Cell Cycle Checkpoints; Cells, Cultured; Chlorocebus aethiops; Cyclin A2; Cyclin B1; G1 Phase; Humans; Indoles; Mice; Mice, Inbred C57BL; Nitrophenols; Piperazines; Proto-Oncogene Proteins c-bcl-2; Pyrroles; Resting Phase, Cell Cycle; SARS-CoV-2; Sulfonamides; Thymidine Kinase; Vero Cells; COVID-19 Drug Treatment
PubMed: 34586865
DOI: 10.1128/JVI.01399-21 -
Molecules (Basel, Switzerland) Feb 2023In the present study, the binding affinity of 52 bioactive secondary metabolites from towards the anti-apoptotic B-cell lymphoma-2 (Bcl-2) protein (PDB: 2W3L) structure...
Computational Studies of Bioactive Secondary Metabolites from against Anti-Apoptotic B-Cell Lymphoma-2 (Bcl-2) Protein Associated with Cancer Cell Survival and Resistance.
In the present study, the binding affinity of 52 bioactive secondary metabolites from towards the anti-apoptotic B-cell lymphoma-2 (Bcl-2) protein (PDB: 2W3L) structure was identified by using in silico molecular docking and molecular dynamics simulation. The molecular docking results demonstrated that the binding energies of docked compounds with Bcl-2 protein ranged from -5.3 kcal/mol to -10.1 kcal/mol. However, the lowest binding energy (-10.1 kcal/mol) was offered by Friedelin against Bcl-2 protein when compared to other metabolites and the standard drug Obatoclax (-8.4 kcal/mol). The molecular dynamics simulations revealed that the Friedelin-Bcl-2 protein complex was found to be stable throughout the simulation period of 100 ns. Overall, the predicted Absorption, Distribution, Metabolism, Excretion, and Toxicity (ADMET) properties of Friedelin are relatively better than Obatoclax, with the most noticeable differences in many parameters where Friedelin has no AMES toxicity, hepatotoxicity, and skin sensitization. The ADMET profiling of selected compounds supported their in silico drug-likeness properties. Based on the computational analyses, the present study concluded that Friedelin of was found to be the potential inhibitor of the Bcl-2 protein, which merits attention for further in vitro and in vivo studies before clinical trials.
Topics: Humans; Apoptosis Regulatory Proteins; Cell Survival; Molecular Docking Simulation; Molecular Dynamics Simulation; Neoplasms; Proto-Oncogene Proteins c-bcl-2; Wedelia; Phytochemicals
PubMed: 36838574
DOI: 10.3390/molecules28041588 -
BioRxiv : the Preprint Server For... Aug 2021Targeting host factors is a promising strategy to develop broad-spectrum antiviral drugs. Drugs targeting anti-apoptotic Bcl-2 family proteins that were originally...
UNLABELLED
Targeting host factors is a promising strategy to develop broad-spectrum antiviral drugs. Drugs targeting anti-apoptotic Bcl-2 family proteins that were originally developed as tumor suppressors have been reported to inhibit multiplication of different types of viruses. However, the mechanisms whereby Bcl-2 inhibitors exert their antiviral activity remain poorly understood. In this study, we have investigated the mechanisms by which obatoclax (OLX) and ABT-737 Bcl-2 inhibitors exhibited a potent antiviral activity against the mammarenavirus lymphocytic choriomeningitis virus (LCMV). OLX and ABT-737 potent anti-LCMV activity was not associated with their pro-apoptotic properties, but rather their ability of inducing cell arrest at G0/G1 phase. OLX and ABT-737 mediated inhibition of Bcl-2 correlated with reduced expression levels of thymidine kinase 1 (TK1), cyclin A2 (CCNA2), and cyclin B1 (CCNB1) cell cycle regulators. In addition, siRNA-mediated knock down of TK1, CCNA2, and CCNB1 resulted in reduced levels of LCMV multiplication. The antiviral activity exerted by Bcl-2 inhibitors correlated with reduced levels of viral RNA synthesis at early times of infection. Importantly, ABT-737 exhibited moderate efficacy in a mouse model of LCMV infection, and Bcl-2 inhibitors displayed broad-spectrum antiviral activities against different mammarenaviruses and SARS-CoV-2. Our results suggest that Bcl-2 inhibitors, actively being explored as anti-cancer therapeutics, might be repositioned as broad-spectrum antivirals.
IMPORTANCE
Anti-apoptotic Bcl-2 inhibitors have been shown to exert potent antiviral activities against various types of viruses via mechanisms that are currently poorly understood. This study has revealed that Bcl-2 inhibitors mediated cell cycle arrest at the G0/G1 phase, rather than their pro-apoptotic activity, plays a critical role in blocking mammarenavirus multiplication in cultured cells. In addition, we show that Bcl-2 inhibitor ABT-737 exhibited moderate anti-mammarenavirus activity , and that Bcl-2 inhibitors displayed broad-spectrum antiviral activities against different mammarenaviruses and SARS-CoV-2. Our results suggest that Bcl-2 inhibitors, actively being explored as anti-cancer therapeutics, might be repositioned as broad-spectrum antivirals.
PubMed: 34426809
DOI: 10.1101/2021.08.16.456587 -
American Journal of Cancer Research 2019Sphingosine kinase 2 (SPHK2) is a key factor within sphingolipid metabolism, responsible for the conversion of pro-apoptotic sphingosine to the pro-survival...
Sphingosine kinase 2 (SPHK2) is a key factor within sphingolipid metabolism, responsible for the conversion of pro-apoptotic sphingosine to the pro-survival sphingosine-1-phosphate. We have previously shown that ABC294640, a first-in-class SPHK2 inhibitor, inhibits growth of cholangiocarcinoma cells. In a Phase I study of ABC294640 in tumors, the best response was achieved in a cholangiocarcinoma patient. These data suggest SPHK2 as a novel therapeutic target of cholangiocarcinoma. However, the antitumor mechanism of ABC294640 in cholangiocarcinoma remains not clear. In the current study, we found that ABC294640 upregulated expression of pro-apoptotic NOXA. In cholangiocarcinoma patients, high NOXA mRNA expression was associated with better overall survival. Also, SPHK2 mRNA expression was negatively correlated with NOXA mRNA expression. NOXA is known to degrade MCL1, an anti-apoptotic BCL2 protein. We showed that ABC294640 directed MCL1 for proteasome degradation. Knockdown of NOXA prevented ABC294640-induced MCL1 degradation and apoptosis. In addition, ABC294640 had a synergistic effect with BCL2/BCL-XL inhibitors ABT-263 and Obatoclax in inhibiting cell growth. Combined treatment with ABC294640 and BCL2/BCL-XL inhibitors induced potent apoptosis. Silencing of MCL1 also potentiated ABT-263-induced cytotoxicity. Furthermore, we found that both SPHK2 and MCL1 protein expression were significantly higher in cholangiocarcinoma than that in nontumoral bile ducts. SPHK2 expression correlated significantly with MCL1 expression. Our study reveals that ABC294640 inhibits cholangiocarcinoma cell growth and sensitizes the antitumor effect of BCL2/BCL-XL inhibitors through NOXA-mediated MCL1 degradation. Combinations of ABC294640 with BCL2/BCL-XL inhibitors may provide novel strategies for the treatment of cholangiocarcinoma.
PubMed: 30949409
DOI: No ID Found -
Journal of Experimental & Clinical... Apr 2017Osteosarcoma is the most frequent primary malignant bone tumor. Although survival has distinctly increased due to neoadjuvant chemotherapy in the past, patients with...
BACKGROUND
Osteosarcoma is the most frequent primary malignant bone tumor. Although survival has distinctly increased due to neoadjuvant chemotherapy in the past, patients with metastatic disease and poor response to chemotherapy still have an adverse prognosis. Hence, development of new therapeutic strategies is still of utmost importance.
METHODS
Anticancer activity of KP46 against osteosarcoma cell models was evaluated as single agent and in combination approaches with chemotherapeutics and Bcl-2 inhibitors using MTT assay. Underlying mechanisms were tested by cell cycle, apoptosis and autophagy assays.
RESULTS
KP46 exerted exceptional anticancer activity at the nanomolar to low micromolar range, depending on the assay format, against all osteosarcoma cell models with minor but significant differences in IC values. KP46 treatment of osteosarcoma cells caused rapid loss of cell adhesion, weak cell cycle accumulation in S-phase and later signs of apoptotic cell death. Furthermore, already at sub-cytotoxic concentrations KP46 reduced the migratory potential of osteosarcoma cells and exerted synergistic effects with cisplatin, a standard osteosarcoma chemotherapeutic. Moreover, the gallium compound induced signs of autophagy in osteosarcoma cells. Accordingly, blockade of autophagy by chloroquine but also by the Bcl-2 inhibitor obatoclax increased the cytotoxic activity of KP46 treatment significantly, suggesting autophagy induction as a protective mechanism against KP46.
CONCLUSION
Together, our results identify KP46 as a new promising agent to supplement standard chemotherapy and possible future targeted therapy in osteosarcoma.
Topics: Antineoplastic Combined Chemotherapy Protocols; Autophagy; Bone Neoplasms; Cell Cycle; Cell Line, Tumor; Cell Movement; Drug Screening Assays, Antitumor; Drug Synergism; Humans; Indoles; Molecular Targeted Therapy; Organometallic Compounds; Osteosarcoma; Oxyquinoline; Pyrroles
PubMed: 28403890
DOI: 10.1186/s13046-017-0527-z -
Tumour Biology : the Journal of the... Mar 2017Multiple factors including tumor heterogeneity and intrinsic or acquired resistance have been associated with drug resistance in lung cancer. Increased stemness and the...
Multiple factors including tumor heterogeneity and intrinsic or acquired resistance have been associated with drug resistance in lung cancer. Increased stemness and the plasticity of cancer cells have been identified as important mechanisms of resistance; therefore, treatments targeting cancer cells independent of stemness phenotype would be much more effective in treating lung cancer. In this article, we have characterized the anticancer effects of the antibiotic Nigericin in cells displaying varying degrees of stemness and resistance to anticancer drugs, arising from (1) routine culture conditions, (2) prolonged periods of serum starvation. These cells are highly resistant to conventional anticancer drugs such as Paclitaxel, Hydroxyurea, Colchicine, Obatoclax, Wortmannin, and LY294002, and the multidrug-resistant phenotype of cells growing under prolonged periods of serum starvation is likely the result of extensive rewiring of signaling pathways, and (3) lung tumorspheres that are enriched for cancer stem-like cells. We found that Nigericin potently inhibited the viability of cells growing under routine culture conditions, prolonged periods of serum starvation, and lung tumorspheres. In addition, we found that Nigericin downregulated the expression of key proteins in the Wnt canonical signaling pathway such as LRP6, Wnt5a/b, and β-catenin, but promotes β-catenin translocation into the nucleus. The antitumor effects of Nigericin were potentiated by the Wnt activator HLY78 and by therapeutic levels of the US Food and Drug Administration-approved drug Digitoxin and its novel synthetic analog MonoD. We believe that Nigericin may be used in a co-therapy model in combination with other novel chemotherapeutic agents in order to achieve potent inhibition of cancers that display varying degrees of stemness, potentially leading to sustained anticancer effects.
Topics: Antineoplastic Agents; Benzodioxoles; Cell Line, Tumor; Cell Proliferation; Drug Resistance, Multiple; Drug Resistance, Neoplasm; Humans; Lung Neoplasms; Neoplastic Stem Cells; Nigericin; Phenanthridines; Wnt Signaling Pathway
PubMed: 28351327
DOI: 10.1177/1010428317694310 -
Frontiers in Cellular and Infection... 2018Neglected tropical diseases, especially those caused by parasites, are significantly underserved by current drug development efforts, mostly due to the high costs and...
Neglected tropical diseases, especially those caused by parasites, are significantly underserved by current drug development efforts, mostly due to the high costs and low economic returns. One method for lowering the costs of drug discovery and development for these diseases is to repurpose drugs developed for other indications. Here, we present the results of a screen of five repurposed drug libraries to identify potential new lead compounds to treat amebiasis, a disease that affects tens of millions of people and causes ~100,000 deaths annually. , the causative agent of amebiasis, has two major life cycle stages, the trophozoite and the cyst. The current primary treatment for amebiasis, nitroimidazole compounds, do not eliminate parasites from the colonic lumen, necessitating a multi-drug treatment regimen. We aimed to address this problem by screening against both life stages, with the aim of identifying a single drug that targets both. We successfully identified eleven compounds with activity against both cysts and trophozoites, as well as multiple compounds that killed trophozoites with improved efficacy over existing drugs. Two lead compounds (anisomycin and prodigiosin) were further characterized for activity against metronidazole (MNZ) resistant parasites and mature cysts. Anisomycin and prodigiosin were both able to kill MNZ resistant parasites while prodigiosin and its analog obatoclax were active against mature cysts. This work confirms the feasibility of identifying drugs that target both trophozoites and cysts, and is an important step toward developing improved treatment regimens for infection.
Topics: Anisomycin; Antiprotozoal Agents; Cell Survival; Drug Evaluation, Preclinical; Drug Repositioning; Drug Resistance; Entamoeba; High-Throughput Screening Assays; Life Cycle Stages; Metronidazole; Prodigiosin; Spores, Protozoan; Trophozoites
PubMed: 30175074
DOI: 10.3389/fcimb.2018.00276 -
Oncotarget Jun 2018The activated JAK2-V617F mutant is very frequently found in myeloproliferative neoplasms (MPNs), and its inhibitor ruxolitinib has been in clinical use, albeit with...
Mechanisms for mTORC1 activation and synergistic induction of apoptosis by ruxolitinib and BH3 mimetics or autophagy inhibitors in JAK2-V617F-expressing leukemic cells including newly established PVTL-2.
The activated JAK2-V617F mutant is very frequently found in myeloproliferative neoplasms (MPNs), and its inhibitor ruxolitinib has been in clinical use, albeit with limited efficacies. Here, we examine the signaling mechanisms from JAK2-V617F and responses to ruxolitinib in JAK2-V617F-positive leukemic cell lines, including PVTL-2, newly established from a patient with post-MPN secondary acute myeloid leukemia, and the widely used model cell line HEL. We have found that ruxolitinib downregulated the mTORC1/S6K/4EBP1 pathway at least partly through inhibition of the STAT5/Pim-2 pathway with concomitant downregulation of c-Myc, MCL-1, and BCL-xL as well as induction of autophagy in these cells. Ruxolitinib very efficiently inhibited proliferation but only modestly induced apoptosis. However, inhibition of BCL-xL/BCL-2 by the BH3 mimetics ABT-737 and navitoclax or BCL-xL by A-1331852 induced caspase-dependent apoptosis involving activation of Bak and Bax synergistically with ruxolitinib in HEL cells. On the other hand, the putative pan-BH3 mimetic obatoclax as well as chloroquine and bafilomycin A1 inhibited autophagy at its late stage and induced apoptosis in PVTL-2 cells synergistically with ruxolitinib. The present study suggests that autophagy as well as the anti-apoptotic BCL-2 family members, regulated at least partly by the mTORC1 pathway downstream of STAT5/Pim-2, protects JAK2-V617F-positive leukemic cells from ruxolitinib-induced apoptosis depending on cell types and may contribute to development of new strategies against JAK2-V617F-positive neoplasms.
PubMed: 29928488
DOI: 10.18632/oncotarget.25515 -
PloS One 2014Altered expression of Mcl-1, an anti-apoptotic member of the Bcl-2 family, has been linked to the progression and outcome of a variety of malignancies. We have...
BACKGROUND
Altered expression of Mcl-1, an anti-apoptotic member of the Bcl-2 family, has been linked to the progression and outcome of a variety of malignancies. We have previously reported the overexpression of Mcl-1 protein in human oral cancers. The present study aimed to evaluate the clinicopathological significance of the expression of three known Mcl-1 isoforms in oral tumors and the effect of targeting Mcl-1L isoform on chemosensitivity of oral cancer cells.
METHODS
The expression of Mcl-1 isoforms- Mcl-1L, Mcl-1S & Mcl-1ES was analyzed in 130 paired oral tumors and 9 oral cell lines using quantitative real-time PCR & protein by western blotting. The Mcl-1 mRNA levels were correlated with clinicopathological parameters and outcome of oral cancer patients. The effect of Mcl-1L shRNA or Obatoclax (a small molecule Mcl-1 inhibitor), in combination with Cisplatin on chemosensitivity of oral cancer cells was also assessed.
RESULTS
Anti-apoptotic Mcl-1L was predominantly expressed, over low or undetectable pro-apoptotic Mcl-1S and Mcl-1ES isoforms. The Mcl-1L transcripts were significantly overexpressed in all cancer cell lines and in 64% oral tumors versus adjacent normals (P<0.02). In oral cancer patients, high Mcl-1L expression was significantly associated with node positivity (P = 0.021), advanced tumor size (P = 0.013) and poor overall survival (P = 0.002). Multivariate analysis indicated Mcl-1L to be an independent prognostic factor for oral cancers (P = 0.037). Mcl-1L shRNA knockdown or its inhibition by Obatoclax in combination with Cisplatin synergistically reduced viability and growth of oral cancer cells than either treatment alone.
CONCLUSION
Our studies suggest that overexpression of Mcl-1L is associated with poor prognosis and chemoresistance in oral cancers. Mcl-1L is an independent prognostic factor and a potential therapeutic target in oral cancers.
Topics: Adolescent; Adult; Aged; Aged, 80 and over; Alternative Splicing; Antineoplastic Agents; Cell Line, Tumor; Cisplatin; Drug Resistance, Neoplasm; Drug Synergism; Female; Gene Expression Regulation, Neoplastic; Humans; Indoles; Male; Middle Aged; Mouth Neoplasms; Myeloid Cell Leukemia Sequence 1 Protein; Prognosis; Protein Isoforms; Pyrroles; Young Adult
PubMed: 25409302
DOI: 10.1371/journal.pone.0111927 -
Molecular and Cellular Biology Sep 2014In selective autophagy, the adaptor protein SQSTM1/p62 plays a critical role in recognizing/loading cargo (e.g., malfolded proteins) into autophagosomes for lysosomal...
In selective autophagy, the adaptor protein SQSTM1/p62 plays a critical role in recognizing/loading cargo (e.g., malfolded proteins) into autophagosomes for lysosomal degradation. Here we report that whereas SQSTM1/p62 levels fluctuated in a time-dependent manner during autophagy, inhibition or knockdown of Cdk9/cyclin T1 transcriptionally downregulated SQSTM1/p62 but did not affect autophagic flux. These interventions, or short hairpin RNA (shRNA) directly targeting SQSTM1/p62, resulted in cargo loading failure and inefficient autophagy, phenomena recently described for Huntington's disease neurons. These events led to the accumulation of the BH3-only protein NBK/Bik on endoplasmic reticulum (ER) membranes, most likely by blocking loading and autophagic degradation of NBK/Bik, culminating in apoptosis. Whereas NBK/Bik upregulation was further enhanced by disruption of distal autophagic events (e.g., autophagosome maturation) by chloroquine (CQ) or Lamp2 shRNA, it was substantially diminished by inhibition of autophagy initiation (e.g., genetically by shRNA targeting Ulk1, beclin-1, or Atg5 or pharmacologically by 3-methyladenine [3-MA] or spautin-1), arguing that NBK/Bik accumulation stems from inefficient autophagy. Finally, NBK/Bik knockdown markedly attenuated apoptosis in vitro and in vivo. Together, these findings identify novel cross talk between autophagy and apoptosis, wherein targeting SQSTM1/p62 converts cytoprotective autophagy to an inefficient form due to cargo loading failure, leading to NBK/Bik accumulation, which triggers apoptosis.
Topics: Adaptor Proteins, Signal Transducing; Animals; Antineoplastic Agents; Apoptosis; Apoptosis Regulatory Proteins; Autophagy; Boronic Acids; Bortezomib; Cell Line, Tumor; Cells, Cultured; Cyclin T; Cyclin-Dependent Kinase 9; Cycloheximide; Flavonoids; Gene Expression Regulation; Heat-Shock Proteins; Humans; Indoles; Leupeptins; Membrane Proteins; Mice; Mice, Knockout; Mitochondrial Proteins; Piperidines; Protein Transport; Pyrazines; Pyrroles; RNA, Small Interfering; Sequestosome-1 Protein
PubMed: 25002530
DOI: 10.1128/MCB.01383-13