In vitro evaluation of Neosetophomone B inducing apoptosis in cutaneous T cell lymphoma by targeting the FOXM1 signaling pathway.

BACKGROUND: Cutaneous T cell lymphoma (CTCL) is a T cell-derived non-Hodgkin lymphoma primarily affecting the skin, with treatment posing a significant challenge and low survival rates. OBJECTIVE: In this study, we investigated the anti-cancer potential of Neosetophomone B (NSP-B), a fungal-derived secondary metabolite, on CTCL cell lines H9 and HH. METHODS: Cell viability was measured using Cell counting Kit-8 (CCK8) assays. Apoptosis was measured by annexin V/PI dual staining. Immunoblotting was performed to examine the expression of proteins. Applied Biosystems' high-resolution Human Transcriptome Array 2.0 was used to examine gene expression. RESULTS: NSP-B induced apoptosis in CTCL cells by activating mitochondrial signaling pathways and caspases. We observed downregulated expression of BUB1B, Aurora Kinases A and B, cyclin-dependent kinases (CDKs) 4 and 6, and polo-like kinase 1 (PLK1) in NSP-B treated cells, which was further corroborated by Western blot analysis. Notably, higher expression levels of these genes showed reduced overall and progression-free survival in the CTCL patient cohort. FOXM1 and BUB1B expression exhibited a dose-dependent reduction in NSP-B-treated CTCL cells.FOXM1 silencing decreased cell viability and increased apoptosis via BUB1B downregulation. Moreover, NSP-B suppressed FOXM1-regulated genes, such as Aurora Kinases A and B, CDKs 4 and 6, and PLK1. The combined treatment of Bortezomib and NSP-B showed greater efficacy in reducing CTCL cell viability and promoting apoptosis compared to either treatment alone. CONCLUSION: Our findings suggest that targeting the FOXM1 pathway may provide a promising therapeutic strategy for CTCL management, with NSP-B offering significant potential as a novel treatment option.

Inhibition of EZH2 and activation of ERRγ synergistically suppresses gastric cancer by inhibiting FOXM1 signaling pathway.

BACKGROUND: Gastric cancer (GC) is a leading cause of cancer-related mortality worldwide, because of the low efficacy of current therapeutic strategies. Estrogen-related receptor γ (ERRγ) was previously showed as a suppressor of GC. However, the mechanism and effective therapeutic method based on ERRγ is yet to be developed. METHODS: The expression levels of ERRγ, EZH2, and FOXM1 were detected by immunohistochemistry, qRT-PCR, and western blot. The regulatory mechanisms of ERRγ and FOXM1 were analyzed by ChIP, EMSA, and siRNA. The effects of EZH2 inhibitor (GSK126) or/and ERRγ agonist (DY131) on the tumorigenesis of gastric cancer cell lines were examined by cell proliferation, transwell migration, wound healing, and colony formation assays. Meanwhile, the inhibitory effects of GSK126 or/and DY131 on tumor growth were analyzed by xenograft tumor growth assay. RESULTS: The expression of ERRγ was suppressed in tumor tissues of GC patients and positively correlated with prognosis, as opposed to that of EZH2 and FOXM1. EZH2 transcriptionally suppressed ERRγ via H3K27me3, which subsequently activated the expression of master oncogene FOXM1. The combination of GSK126 and DY131 synergistically activated ERRγ expression, which subsequently inhibited the expression of FOXM1 and its regulated pathways. Synergistic combination of GSK126 and DY131 significantly inhibited the tumorigenesis of GC cell lines and suppressed the growth of GC xenograft. CONCLUSION: The FOXM1 signaling pathway underlying the ERRγ-mediated gastric cancer suppression was identified. Furthermore, combined treatment with EZH2 inhibitor and ERRγ agonist synergistically suppressed GC progression by inhibiting this signaling pathway, suggesting its high potential in treating GC patients.

Prenylated xanthones from Metaxya rostrata suppress FoxM1 and induce active cell death by distinct mechanisms.

BACKGROUND: Metaxya rostrata C.Presl (Metaxyaceae) is a tree fern widespread in Central and South America and the dried rhizome is used in ethnic medicine against intestinal ulcers or tumors. An activity-guided isolation resulted in two structurally related xanthones: 2-deprenyl-rheediaxanthone B (XB) and 2-deprenyl-7-hydroxy-rheediaxanthone B (OH-XB). HYPOTHESIS/PURPOSE: This study analyzed the cytotoxic activity and underlying cellular mechanisms of OH-XB for the first time in comparison to XB. METHODS: We exposed the colorectal cancer cell line SW480 and F331 fibroblasts to XB and OH-XB and determined cell viability by neutral red uptake and nuclear morphology by staining with Hoechst dye. Cell cycle distribution and the mechanism of cell death were analyzed by FACS and western blot. Knockdown of FoxM1 expression was performed with siRNA. RESULTS: OH-XB was at least as cytotoxic as XB in the induction of cell cycle arrest and active cell death. While both compounds strongly inhibited the transcription factor FoxM1, the cellular mechanisms of growth arrest and cell death induction differed widely: OH-XB induced S-phase cell cycle arrest in contrast to a G2-M-phase arrest by XB. It caused morphological modifications typical for classical apoptosis with increased caspase 7 activity and enhanced cleavage of PARP, while XB caused caspase 2 activation and mitotic catastrophe. After knockdown of FoxM1 expression no induction of caspase activity could be observed. CONCLUSION: In summary, our data clearly showed that XB and OH-XB are promising new lead compounds for cancer therapy with distinct cellular mechanisms. Both compounds are candidates for further pre-clinical and clinical investigations.

Intranasal wnt3a Attenuates Neuronal Apoptosis through Frz1/PIWIL1a/FOXM1 Pathway in MCAO Rats.

After ischemic stroke, apoptosis of neurons is a primary factor in determining outcome. Wnt3a is a naturally occurring protein that has been shown to have protective effects in the brain for traumatic brain injury. Although wnt3a has been investigated in the phenomena of neurogenesis, anti-apoptosis, and anti-inflammation, it has never been investigated as a therapy for stroke. We hypothesized that the potential neuroprotective agent wnt3a would reduce infarction and improve behavior following ischemic stroke by attenuating neuronal apoptosis and promoting cell survival through the Frizzled-1/PIWI1a/FOXM1 pathway in middle cerebral artery occlusion (MCAO) rats. A total of 229 Sprague Dawley rats were assigned to male, female, and 9-month-old male MCAO or sham groups followed by reperfusion 2 h after MCAO. Animals assigned to MCAO were either given wnt3a or its control. To explore the downstream signaling of wnt3a, the following interventions were given: Frizzled-1 siRNA, PIWI1a siRNA, and PIWI1a-clustered regularly interspaced short palindromic repeats, along with the appropriate controls. Post-MCAO assessments included neurobehavioral tests, infarct volume, Western blot, and immunohistochemistry. Endogenous levels of wnt3a and Frizzled-1/PIWI1a/FOXM1 were lowered after MCAO. The administration of intranasal wnt3a, 1 h after MCAO, increased PIWIL1a and FOXM1 expression through Frizzled-1, reducing brain infarction and neurological deficits at 24 and 72 h. Frizzled-1 and PIWI1a siRNAs reversed the protective effects of wnt3a after MCAO. Restoration of PIWI1a after knockdown of Frizzled-1 increased FOXM1 survival protein and reduced cleaved caspase-3 levels. In summary, wnt3a decreases neuronal apoptosis and improves neurological deficits through Frizzled-1/PIWI1a/FOXM1 pathway after MCAO in rats. Therefore, wnt3a is a novel intranasal approach to decrease apoptosis after stroke.SIGNIFICANCE STATEMENT Only 5% of patients receive recombinant tissue plasminogen activator after stroke, and few qualify for mechanical thrombectomy. No neuroprotective agents have been successfully translated to promote neuronal survival in stroke. Thus, using a clinically relevant rat model of stroke, middle cerebral artery occlusion, we explored a novel intranasal administration of wnt3a. wnt3a naturally occurs in the body and crosses the blood-brain barrier, supporting the clinically translatable approach of intranasal administration. Significant neuronal apoptosis occurs during stroke, and wnt3a shows promise due to its antiapoptotic effects. We investigated whether wnt3a mediates its poststroke effects via Frizzled-1 and the impact on its downstream signaling molecules, PIWI1a and FOXM1, in apoptosis. Elucidating the mechanism of wnt3a will identify additional pharmacological targets and further understanding of stroke.

CC-401 Promotes ß-Cell Replication via Pleiotropic Consequences of DYRK1A/B Inhibition.

Pharmacologic expansion of endogenous ß cells is a promising therapeutic strategy for diabetes. To elucidate the molecular pathways that control ß-cell growth we screened ∼2400 bioactive compounds for rat ß-cell replication-modulating activity. Numerous hit compounds impaired or promoted rat ß-cell replication, including CC-401, an advanced clinical candidate previously characterized as a c-Jun N-terminal kinase inhibitor. Surprisingly, CC-401 induced rodent (in vitro and in vivo) and human (in vitro) ß-cell replication via dual-specificity tyrosine phosphorylation-regulated kinase (DYRK) 1A and 1B inhibition. In contrast to rat ß cells, which were broadly growth responsive to compound treatment, human ß-cell replication was only consistently induced by DYRK1A/B inhibitors. This effect was enhanced by simultaneous glycogen synthase kinase-3ß (GSK-3ß) or activin A receptor type II-like kinase/transforming growth factor-ß (ALK5/TGF-ß) inhibition. Prior work emphasized DYRK1A/B inhibition-dependent activation of nuclear factor of activated T cells (NFAT) as the primary mechanism of human ß-cell-replication induction. However, inhibition of NFAT activity had limited effect on CC-401-induced ß-cell replication. Consequently, we investigated additional effects of CC-401-dependent DYRK1A/B inhibition. Indeed, CC-401 inhibited DYRK1A-dependent phosphorylation/stabilization of the ß-cell-replication inhibitor p27Kip1. Additionally, CC-401 increased expression of numerous replication-promoting genes normally suppressed by the dimerization partner, RB-like, E2F and multivulval class B (DREAM) complex, which depends upon DYRK1A/B activity for integrity, including MYBL2 and FOXM1. In summary, we present a compendium of compounds as a valuable resource for manipulating the signaling pathways that control ß-cell replication and leverage a DYRK1A/B inhibitor (CC-401) to expand our understanding of the molecular pathways that control ß-cell growth.

The addition of celecoxib improves the antitumor effect of cetuximab in colorectal cancer: role of EGFR-RAS-FOXM1-ß- catenin signaling axis.

Here we showed that the addition of the COX-2 inhibitor celecoxib improved the antitumor efficacy in colorectal cancer (CRC) of the monoclonal anti-EGFR antibody cetuximab. The addition of celecoxib augmented the efficacy of cetuximab to inhibit cell proliferation and to induce apoptosis in CRC cells. Moreover, the combination of celecoxib and cetuximab was more effective than either treatment alone in reducing the tumor volume in a mouse xenograft model. The combined treatment enhanced the inhibition of EGFR signaling and altered the subcellular distribution of ß-catenin. Moreover, knockdown of FOXM1 showed that this transcription factor participates in this enhanced antitumoral response. Besides, the combined treatment decreased ß-catenin/FOXM1 interaction and reduced the cancer stem cell subpopulation in CRC cells, as indicated their diminished capacity to form colonospheres. Notably, the inmunodetection of FOXM1 in the nuclei of tumor cells in human colorectal adenocarcinomas was significantly associated with response of patients to cetuximab. In summary, our study shows that the addition of celecoxib enhances the antitumor efficacy of cetuximab in CRC due to impairment of EGFR-RAS-FOXM1-ß-catenin signaling axis. Results also support that FOXM1 could be a predictive marker of response of mCRC patients to cetuximab therapy.

miR-149 reverses cisplatin resistance of gastric cancer SGC7901/DDP cells by targeting FoxM1.

Drug resistance remains a major unresolved obstacle for gastric cancer (GC) treatment. Recently, increasing studies have showen that microRNAs (miRNAs) are involved in cancer chemotherapeutic resistance and can potentially be applied to reverse drug resistance in cancers. The relationship between miRNA-149 expression and cisplatin (DDP) resistance in GC cells is still unknown. Here, we detected miR-149 expression by using RT-PCR and found that expression of miR-149 was downregulated in SGC7901/DDP cells compared with SGC7901cells, indicating a role of miR-149 in determining cisplatin-resistance of GC cells. Then, SGC7901/DDP cells were tansfected with miR-149 mimics, MTT assay was performed to determine SGC7901/DDP cell viability, and showed that overexpression of miR-149 inhibited the cell viability after cisplatin treatment, suggesting that up-regulation of miR-149 enhanced SGC7901/DDP cell sensitivity to cisplatin. Furthermore, we confirmed that Forkhead box M1 (FoxM1) is a direct target of miR-149 in SGC7901/DDP cells by using luciferase reporter assay. Besides, we also demonstrated that miR-149 enhances SGC7901/DDP cell sensitivity to cisplatin by downregulating FoxM1 expression. In summary, our data provide new insights that miR-149 plays an important role in determining sensitivity of cisplatin-resistant GC cells by targeting FoxM1 and suggest that miR-149 could be a potential target for reversing drug resistance in GC.