Search Results (16)

Mantle cell lymphoma (MCL) is an aggressive B-cell non-Hodgkin lymphoma characterized by the t(11;14)(q13;q32) cytogenetic abnormality and cyclin D1 overexpression. We have found evidence that Forkhead box M1 (FOXM1), a transcription factor with oncogenic potential, is important in the pathogenesis of MCL. Relatively high levels of FOXM1 proteins were detectable in all six MCL cell lines examined. By immunohistochemistry, we consistently found a subset of FOXM1-positive cells in MCL tumors. Analysis of two Gene Expression Omnibus (GEO) datasets from MCL patients showed that elevated FOXM1 levels significantly correlate with a worse clinical outcome. In MCL cell lines, inhibition of FOXM1 using thiostrepton or shRNA effectively triggered apoptosis and significantly reduced cell growth. FOXM1 forms a positive feedback loop with NFκB in MCL cells. Specifically, inhibition of FOXM1 dramatically decreased the protein level/transcription activity of p65, while enforced FOXM1 expression upregulated p65 and downregulated IκBα, a key NFκB inhibitor. Conversely, curcumin-mediated NFκB inhibition decreased the protein level/DNA binding of FOXM1, while transduction of a constitutively active IKKα construct into MCL cells significantly dampened the inhibitory effects of thiostrepton. Confocal microscopy revealed that FOXM1 and p65 colocalize with each other. In conclusion, FOXM1 and NFκB work collaboratively in promoting the growth and drug resistance of MCL, and FOXM1 may be a potentially useful therapeutic target. Full article

Oxydifficidin is a natural polyketide antibiotic that has long been recognized as a ribosome-targeting agent that inhibits protein synthesis. In this paper, we describe Bacillus velezensis strain EV17 and compare its complete genome sequence with that of the previously characterized B. velezensis strain K-3618 and the difficidin biosynthetic gene cluster (BGC) combined with mass spectrometry to elucidate the production of oxydifficidin by these strains. Toeprinting and small fluorescent peptide assays showed that isolated oxydifficidin induces a generalized inhibition of translation at every step of elongation in protein biosynthesis. In previous studies, it has been demonstrated that oxydifficidin targets bL12 protein. Although spontaneous mutations conferring resistance to oxydifficidin in ribosomal protein bL12 located relatively close to the thiostrepton binding site on uL11, our data show that oxydifficidin binding does not interfere with thiostrepton, thereby refining previous findings about its putative ribosomal target. We are the first to show that this compound does not affect eukaryotic translation and has two orders of magnitude lower effect on eukaryotic cells compared to bacteria. These facts are important to further investigate its potential as a bioprotectant against phytopathogens or even as a therapeutic agent. Full article

The tumor suppressor DAB2IP, a RasGAP and cytoplasmic adaptor protein, modulates signal transduction in response to several extracellular stimuli, negatively regulating multiple oncogenic pathways. Accordingly, the loss of DAB2IP in tumor cells fosters metastasis and enhances chemo- and radioresistance. DAB2IP is rarely mutated in cancer but is frequently downregulated or inactivated by multiple mechanisms. Solid experimental evidence shows that DAB2IP reactivation reduces cancer aggressiveness in tumors driven by multiple different oncogenic mutations, making this protein an interesting target for cancer therapy. Considering this evidence, we screened a drug library to identify molecules that increase DAB2IP protein levels. We employed CRISPR/Cas9 gene editing to generate two prostate cancer cell models in which endogenous DAB2IP is fused to HiBiT, a peptide tag that enables luminescence-based detection of protein levels in a sensitive and quantitative manner. Using this approach, we identified drugs able to increase DAB2IP levels. We focused our attention on thiostrepton, a natural cyclic oligopeptide antibiotic that has been reported to inhibit the survival of various cancer cell lines. Functional experiments revealed that the cancer-inhibitory effect of thiostrepton is reduced in the absence of DAB2IP, suggesting that upregulation of this protein contributes to its action. These findings encourage further development of thiostrepton for the treatment of solid cancers and unveil a novel molecular target underlying its anti-tumoral activity. Full article

Background/Objectives: Breast cancer remains the leading malignancy affecting women worldwide, with significant mortality rates. This study aimed to evaluate the prognostic significance of FOXM1 expression specifically in hormone receptor-positive, HER2-negative (HR+HER2-) breast cancer patients with high KPNA2 expression, and to identify potential FOXM1-targeted therapeutic strategies for this patient subgroup. Methods: We analyzed RNA sequencing and microarray data from three independent cohorts: Mackay Memorial Hospital patient samples, The Cancer Genome Atlas, and Gene Expression Omnibus databases. The expression levels of KPNA2, FOXM1, CCNB1, and CCNB2 were evaluated, with particular emphasis on stratifying patients based on KPNA2 expression levels. Their associations with clinical outcomes were assessed using Gene Set Enrichment Analysis and survival analyses. Results: While KPNA2 expression showed strong positive correlations with FOXM1, CCNB1, and CCNB2 across all datasets, our analysis revealed a distinct prognostic pattern in HR+HER2- breast cancer patients with high KPNA2 expressions. In this specific subgroup, low FOXM1 expression emerged as a favorable prognostic indicator, despite the generally poor prognosis associated with high KPNA2 levels. Gene Set Enrichment Analysis demonstrated significant enrichment of the G2/M checkpoint pathway in high KPNA2-expressing patients, suggesting potential therapeutic vulnerability to FOXM1 inhibition in this subgroup. Conclusions: This study establishes FOXM1 expression as a critical prognostic marker, specifically in KPNA2-high HR+HER2- breast cancer patients, where low FOXM1 levels correlate with improved survival outcomes. These findings suggest that FOXM1 inhibition could be particularly effective in patients with high KPNA2 expression, offering a novel therapeutic strategy for this specific molecular subtype. Several FOXM1 inhibitors, including thiostrepton and FDI-6, warrant investigation as potential targeted treatments for KPNA2-high HR+HER2- breast cancer patients. Full article

Due to limited drug efficacy and drug resistance, it is urgent to explore effective anti-liver cancer drugs. Repurposing drugs is an efficient strategy, with advantages including reduced costs, shortened development cycles, and assured safety. In this study, we adopted a synergistic approach combining computational and experimental methods and identified the antibacterial drug thiostrepton (TST) as a candidate for an anti-liver cancer drug. Although the anti-tumor capabilities of TST have been reported, its role and underlying mechanisms in hepatocellular carcinoma (HCC) remain unclear. TST was found here to inhibit the proliferation of HCC cells effectively, arresting the cell cycle and inducing cell apoptosis, as well as suppressing the cell migration. Further, our findings revealed that TST induced mitochondrial impairment, which was demonstrated by destroyed mitochondrial structures, reduced mitochondria, and decreased mitochondrial membrane potential (MMP). TST caused the production of reactive oxygen species (ROS), and the mitochondrial impairment and proliferation inhibition of HCC cells were completely restored by the ROS scavenger N-acetyl-L-cysteine (NAC). Moreover, we discovered that TST induced mitophagy, and autophagy inhibition effectively promoted the anti-cancer effects of TST on HCC cells. In conclusion, our study suggests TST as a promising candidate for the treatment of liver cancers, and these findings provide theoretical support for the further development and potential application of TST in clinical liver cancer therapy. Full article

With the rising prevalence of obesity, non-alcoholic fatty liver disease (NAFLD) now affects 20–25% of the global population. NAFLD, a progressive condition associated with oxidative stress, can result in cirrhosis and liver cancer in 10% and 3% of patients suffering NAFLD, respectively. Therapeutic options are currently limited, emphasizing the need for novel treatments. In this study, we examined the potential of activating the transcription factor NRF2, a crucial player in combating oxidative stress, as an innovative approach to treating NAFLD. Utilizing a CRISPR/Cas9-engineered human HEK293T cell line, we were able to monitor the expression of heme oxygenase-1 (HMOX1), an NRF2 target, using a Nanoluc luciferase tag. Our model was validated using a known NRF2 activator, after which we screened 1200 FDA-approved drugs, unearthing six compounds (Disulfiram, Thiostrepton, Auranofin, Thimerosal, Halofantrine, and Vorinostat) that enhanced NRF2 activity and antioxidant response. These compounds demonstrated protective effects against oxidative stress induced by hydrogen peroxide and lipid droplets accumulation in vitro with hepatoma HUH-7 cells. Our study underscores the utility of CRISPR/Cas9 tagging with Nanoluc luciferase in identifying potential NRF2 activators, paving the way for potential NAFLD therapeutics. Full article

Considerable emphasis is being placed on combinatorial chemotherapeutic/natural treatments for breast cancer. This study reveals the synergistic anti-tumor activity of morin and Doxorubicin (Dox) co-treatment on MDA-MB-231 triple-negative breast cancer (TNBC) cell proliferation. Morin/Dox treatment promoted Dox uptake and induced DNA damage and formation of nuclear foci of p-H2A.X. Furthermore, DNA repair proteins, RAD51 and survivin, and cell cycle proteins, cyclin B1 and forkhead Box M1 (FOXM1), were induced by Dox alone but attenuated by morin/Dox co-treatment. In addition, Annexin V/7-AAD analysis revealed that necrotic cell death after co-treatment and apoptotic cell death by Dox alone were associated with the induction of cleaved PARP and caspase-7 without Bcl-2 family involvement. FOXM1 inhibition by thiostrepton showed that co-treatment caused FOXM1-mediated cell death. Furthermore, co-treatment downregulated the phosphorylation of EGFR and STAT3. Flow cytometry showed that the accumulation of cells in the G2/M and S phases might be linked to cellular Dox uptake, p21 upregulation, and cyclin D1 downregulation. Taken together, our study shows that the anti-tumor effect of morin/Dox co-treatment is due to the suppression of FOXM1 and attenuation of EGFR/STAT3 signaling pathways in MDA-MB-231 TNBC cells, which suggests that morin offers a means of improving therapeutic efficacy in TNBC patients. Full article

We have designed translational animal models to investigate cardiac profibrotic gene signatures. Domestic pigs were treated with cardiotoxic drugs (doxorubicin, DOX, n = 5 or Myocet^®, MYO, n = 5) to induce replacement fibrosis via cardiotoxicity. Reactive interstitial fibrosis was triggered by LV pressure overload by artificial isthmus stenosis with stepwise developing myocardial hypertrophy and final fibrosis (Hyper, n = 3) or by LV volume overload in the adverse remodeled LV after myocardial infarction (RemoLV, n = 3). Sham interventions served as controls and healthy animals (Control, n = 3) served as a reference in sequencing study. Myocardial samples from the LV of each group were subjected to RNA sequencing. RNA-seq analysis revealed a clear distinction between the transcriptomes of myocardial fibrosis (MF) models. Cardiotoxic drugs activated the TNF-alpha and adrenergic signaling pathways. Pressure or volume overload led to the activation of FoxO pathway. Significant upregulation of pathway components enabled the identification of potential drug candidates used for the treatment of heart failure, such as ACE inhibitors, ARB, ß-blockers, statins and diuretics specific to the distinct MF models. We identified candidate drugs in the groups of channel blockers, thiostrepton that targets the FOXM1-regulated ACE conversion to ACE2, tyrosine kinases or peroxisome proliferator-activated receptor inhibitors. Our study identified different gene targets involved in the development of distinct preclinical MF protocols enabling tailoring expression signature-based approach for the treatment of MF. Full article

Rhodothermus marinus is a halophilic extreme thermophile, with potential as a model organism for studies of the structural basis of antibiotic resistance. In order to facilitate genetic studies of this organism, we have surveyed the antibiotic sensitivity spectrum of R. marinus and identified spontaneous antibiotic-resistant mutants. R. marinus is naturally insensitive to aminoglycosides, aminocylitols and tuberactinomycins that target the 30S ribosomal subunit, but is sensitive to all 50S ribosomal subunit-targeting antibiotics examined, including macrolides, lincosamides, streptogramin B, chloramphenicol, and thiostrepton. It is also sensitive to kirromycin and fusidic acid, which target protein synthesis factors. It is sensitive to rifampicin (RNA polymerase inhibitor) and to the fluoroquinolones ofloxacin and ciprofloxacin (DNA gyrase inhibitors), but insensitive to nalidixic acid. Drug-resistant mutants were identified using rifampicin, thiostrepton, erythromycin, spiramycin, tylosin, lincomycin, and chloramphenicol. The majority of these were found to have mutations that are similar or identical to those previously found in other species, while several novel mutations were identified. This study provides potential selectable markers for genetic manipulations and demonstrates the feasibility of using R. marinus as a model system for studies of ribosome and RNA polymerase structure, function, and evolution. Full article

Diagnosis in an advanced state is a major hallmark of ovarian cancer and recurrence after first line treatment is common. With upcoming novel therapies, tumor markers that support patient stratification are urgently needed to prevent ineffective therapy. Therefore, the transcription factor FOXM1 is a promising target in ovarian cancer as it is frequently overexpressed and associated with poor prognosis. In this study, fresh tissue specimens of 10 ovarian cancers were collected to investigate tissue cultures in their ability to predict individual treatment susceptibility and to identify the benefit of FOXM1 inhibition. FOXM1 inhibition was induced by thiostrepton (3 µM). Carboplatin (0.2, 2 and 20 µM) and olaparib (10 µM) were applied and tumor susceptibility was analyzed by tumor cell proliferation and apoptosis in immunofluorescence microscopy. Resistance mechanisms were investigated by determining the gene expression of FOXM1 and its targets BRCA1/2 and RAD51. Ovarian cancer tissue was successfully maintained for up to 14 days ex vivo, preserving morphological characteristics of the native specimen. Thiostrepton downregulated FOXM1 expression in tissue culture. Individual responses were observed after combined treatment with carboplatin or olaparib. Thus, we successfully implemented a complex tissue culture model to ovarian cancer and showed potential benefit of combined FOXM1 inhibition. Full article

A central hallmark of tumorigenesis is metabolic alterations that increase mitochondrial reactive oxygen species (mROS). In response, cancer cells upregulate their antioxidant capacity and redox-responsive signaling pathways. A promising chemotherapeutic approach is to increase ROS to levels incompatible with tumor cell survival. Mitochondrial peroxiredoxin 3 (PRX3) plays a significant role in detoxifying hydrogen peroxide (H₂O₂). PRX3 is a molecular target of thiostrepton (TS), a natural product and FDA-approved antibiotic. TS inactivates PRX3 by covalently adducting its two catalytic cysteine residues and crosslinking the homodimer. Using cellular models of malignant mesothelioma, we show here that PRX3 expression and mROS levels in cells correlate with sensitivity to TS and that TS reacts selectively with PRX3 relative to other PRX isoforms. Using recombinant PRXs 1–5, we demonstrate that TS preferentially reacts with a reduced thiolate in the PRX3 dimer at mitochondrial pH. We also show that partially oxidized PRX3 fully dissociates to dimers, while partially oxidized PRX1 and PRX2 remain largely decameric. The ability of TS to react with engineered dimers of PRX1 and PRX2 at mitochondrial pH, but inefficiently with wild-type decameric protein at cytoplasmic pH, supports a novel mechanism of action and explains the specificity of TS for PRX3. Thus, the unique structure and propensity of PRX3 to form dimers contribute to its increased sensitivity to TS-mediated inactivation, making PRX3 a promising target for prooxidant cancer therapy. Full article

Mycobacterium abscessus is a rapid-growing, multidrug-resistant, non-tuberculous mycobacterial species responsible for a variety of human infections, such as cutaneous and pulmonary infections. M. abscessus infections are very difficult to eradicate due to the natural and acquired multidrug resistance profiles of M. abscessus. Thus, there is an urgent need for the development of effective drugs or regimens against M. abscessus infections. Here, we report the activity of a US Food and Drug Administration approved drug, thiostrepton, against M. abscessus. We found that thiostrepton significantly inhibited the growth of M. abscessus wild-type strains, subspecies, clinical isolates, and drug-resistant mutants in vitro and in macrophages. In addition, treatment of macrophages with thiostrepton significantly decreased proinflammatory cytokine production in a dose-dependent manner, suggesting an inhibitory effect of thiostrepton on inflammation induced during M. abscessus infection. We further showed that thiostrepton exhibits antimicrobial effects in vivo using a zebrafish model of M. abscessus infection. Full article

Forkhead Box M1 (FOXM1) is an oncogenic transcription factor implicated in the pathogenesis of solid and hematologic cancers. In this study, we examined the significance of FOXM1 in NPM-ALK-positive anaplastic large cell lymphoma (NPM-ALK + ALCL), with a focus on how it interacts with NPM-ALK, which is a key oncogenic driver in these tumors. FOXM1 was expressed in NPM-ALK + ALCL cell lines (5/5), patient samples (21/21), and tumors arising in NPM-ALK transgenic mice (4/4). FOXM1 was localized in the nuclei and confirmed to be transcriptionally active. Inhibition of FOXM1 in two NPM-ALK + ALCL cells using shRNA and pharmalogic agent (thiostrepton) resulted in reductions in cell growth and soft-agar colony formation, which were associated with apoptosis and cell-cycle arrest. FOXM1 is functionally linked to NPM-ALK, as FOXM1 enhanced phosphorylation of the NPM-ALK/STAT3 axis. Conversely, DNA binding and transcriptional activity of FOXM1 was dependent on the expression of NPM-ALK. Further studies showed that this dependency hinges on the binding of FOXM1 to NPM1 that heterodimerizes with NPM-ALK, and the phosphorylation status of NPM-ALK. In conclusion, we identified FOXM1 as an important oncogenic protein in NPM-ALK+ ALCL. Our results exemplified that NPM-ALK exerts oncogenic effects in the nuclei and illustrated a novel role of NPM1 in NPM-ALK pathobiology. Full article

Methicillin-resistant Staphylococcus aureus (MRSA) poses a significant threat for effective treatment of several difficult-to-treat infections in humans. To identify potential new treatment options for MRSA infections, we screened a clinical compound library consisting of 1524 compounds using a growth inhibition assay in 96-well plates. We identified 34 agents which are either bacteriostatic or bactericidal against log-phase clinical MRSA strain USA300. Among them, 9 candidates (thonzonium, cetylpyridinium, trilocarban, benzododecinium, bithionol, brilliant green, chlorquinaldol, methylbenzethonium and green violet) are known antiseptics, 11 candidates are known antibiotics currently recommended for the treatment of MRSA. We identified 9 new drug candidates, 5 of which (thiostrepton, carbomycin, spiramycin, clofazimine and chloroxine) are antibiotics used for treating other infections than S. aureus infections; 4 of which (quinaldine blue, closantel, dithiazanine iodide and pyrvinium pamoate) are drugs used for treating parasitic diseases or cancer. We ranked these new drug candidates according to their MICs against the MRSA strain USA300. Our findings may have implications for more effective treatment of MRSA infections. Full article

Lyme disease is a leading vector-borne disease in the United States. Although the majority of Lyme patients can be cured with standard 2–4 week antibiotic treatment, 10%–20% of patients continue to suffer from prolonged post-treatment Lyme disease syndrome (PTLDS). While the cause for this is unclear, persisting organisms not killed by current Lyme antibiotics may be involved. In our previous study, we screened an FDA drug library and reported 27 top hits that showed high activity against Borrelia persisters. In this study, we present the results of an additional 113 active hits that have higher activity against the stationary phase B. burgdorferi than the currently used Lyme antibiotics. Many antimicrobial agents (antibiotics, antivirals, antifungals, anthelmintics or antiparasitics) used for treating other infections were found to have better activity than the current Lyme antibiotics. These include antibacterials such as rifamycins (3-formal-rifamycin, rifaximin, rifamycin SV), thiostrepton, quinolone drugs (sarafloxacin, clinafloxacin, tosufloxacin), and cell wall inhibitors carbenicillin, tazobactam, aztreonam; antifungal agents such as fluconazole, mepartricin, bifonazole, climbazole, oxiconazole, nystatin; antiviral agents zanamivir, nevirapine, tilorone; antimalarial agents artemisinin, methylene blue, and quidaldine blue; antihelmintic and antiparasitic agents toltrazuril, tartar emetic, potassium antimonyl tartrate trihydrate, oxantel, closantel, hycanthone, pyrimethamine, and tetramisole. Interestingly, drugs used for treating other non-infectious conditions including verteporfin, oltipraz, pyroglutamic acid, pidolic acid, and dextrorphan tartrate, that act on the glutathione/γ-glutamyl pathway involved in protection against free radical damage, and also the antidepressant drug indatraline, were found to have high activity against stationary phase B. burgdorferi. Among the active hits, agents that affect cell membranes, energy production, and reactive oxygen species production are more active against the B. burgdorferi persisters than the commonly used antibiotics that inhibit macromolecule biosynthesis. Future studies are needed to evaluate and optimize the promising active hits in drug combination studies in vitro and also in vivo in animal models. These studies may have implications for developing more effective treatments of Lyme disease. Full article