Fig 2: Proposed nongenomic signaling pathways involved in Cd-exposed ht-UtLM cells. Cd-induced proliferative effects observed in ht-UtLM cells have been shown to occur through the membrane/cytosolic GPER receptor via activation of the nongenomic GPER/EGFR/MAPK signaling pathway. Cd concurrently increases the expression of FOXM1 and Cyclin D1 downstream of MAPK44/42, which in turn upregulates the kinase Aurora B, leading to Histone H3 phosphorylation at the serine 10 site, and resulting in human uterine leiomyoma cell proliferation

Fig 3: RNA-Seq analysis of the effects of compounds on gene expression in breast cancer cells. a Regulation of FOXM1-signature genes by NB-73 (4 µM) in MCF7 cells at 9 h and 24 h, and by NB-73 treatment (1.5 µM, 24 h) in MDA-MB-231 cells. b Venn diagram showing overlap of genes with p < 0.05 and regulated more than 2-fold by NB-73, NB-55 and FDI-6 (20 µM) in MCF7 cells, 9 h treatment. c Gene Set Enrichment Analysis (GSEA) showing Enrichment Scores. MDA-MB-231 cells treated with NB-73 (1.5 µM, 24 h) or siFOXM1 (25 nM, 72 h). Gene sets used FOXM1 genes derived from four independent datasets of FOXM1 cistromes.19,52 NES, normalized enrichment score. d REVIGO (Reduce and Visualize Gene Ontologies) analyses showing biological process gene ontologies most impacted by MDA-MB-231 cell treatment with NB-73 or siFOXM1. Circle sizes represent GO term gene count; colors represent similarity along Semantic Space Y.

Fig 4: Compounds NB-55, NB-68, NB-71, and NB-73 suppress breast tumor xenograft growth and the expression of FOXM1-regulated genes. Human breast cancer DT22 cells were injected into the mammary fat pad of intact 7-week-old NSG mice and mice bearing DT22 tumors were dosed in a daily and then every other day from day 7 on with 100 mg/kg of NB-55 or control vehicle by oral gavage; or b with NB-68, NB-71, or NB-73 by s.c. injection at 40 mg/kg daily for 4 days and then 20 mg/kg every third day until day 13 and then 10 mg/kg every third day, or with control vehicle. Tumor volumes in Veh and compound treated animals were monitored (2-way ANOVA, Dunnett’s post-test, **P < 0.01). In c, animals were treated with low doses of NB-73 (5 or 10 mg/kg s.c. daily and then every other day after day 21). Volumes of tumors in Veh and compound treated animals were monitored (2-way ANOVA, Bonferroni post-test, ****P < 0.0001, n = 8 per group, error bars are ± SD). d At the end of treatments, tumors from panel C were processed for gene expression analysis by q-PCR. (t-test, *p < 0.05; **p < 0.01; ***p < 0.001; ****P < 0.0001, n = 8 per group, error bars are ± SD).

Fig 5: miR-199a-3p increased palbociclib efficacy on human HCC cells in vitro and in vivo(A) Hep3B cells were transduced with an Adeno Associated Virus expressing miR-199a-3p (AAVV-199) or with a control AAVV (AAVV-CTRL) at MOI = 100. A group of cells received no treatment (NT). Palbociclib (PB) (20 µM) was added to cell culture 72 h before cell collection. The levels of miR-199a-3p were measured 120 h after transduction. (B) Viability and apoptosis were evaluated 120 h after transduction. Data are represented as mean +SD. (C) Western blot analysis and quantification of RB1, AKT, FOXM1, and their phosphorylated forms in Hep3B-treated cells. The values were normalized on the GAPDH expression. Because Hep3B cells exhibit a low level of full-length protein, digital images of RB1 and p-RB1 were acquired with an exposure time of 300 s instead of 30 s. (D) Tumor nodule volumes in DEN-treated TG221 mice of the following groups: (1) palbociclib (PB) (n = 11); (2) miR-199a-3p mimics (miR-199) (n = 9); (3) palbociclib + miR-199a-3p mimics (PB + miR-199) (n = 11); (4) sorafenib (SF) (n = 9); (5) vehicle/scramble oligonucleotides (CTRL) (n = 9). Experimental therapies started at 6 months, when all mice presented one or more tumor nodules in their livers. Single tumor nodules were monitored by ultrasound at the beginning and end of treatment. (E) Distribution of tumor nodule size measured at the time of euthanization. (F) Mice weight was monitored during treatment to check drug toxicity. Data are represented as mean ± SD. *p value =0.05; **p value =0.01; ***p value =0.001; ****p value =0.0001.