Table 1.
Patient and tumor characteristics.
Fig 1.
Expression level of BRD4 in human neurofibroma and MPNST samples and cell lines.
(A) qRT-PCR was used to determine mRNA levels of BRD4 in paired plexiform neurofibroma (NF, blue, n = 9) and MPNST (red, n = 9) formalin-fixed paraffin-embedded tumor samples, each pair being derived from the same NF1 patient. Asterisk indicates P<0.05. (B) qRT-PCR was used to determine mRNA levels of BRD4 in fresh frozen MPNST (red, n = 11), plexiform neurofibroma (blue, n = 7) and atypical neurofibroma (grey, n = 4). (C) qRT-PCR was used to determine mRNA levels of BRD4 in a cell line panel: Hs53.T neurofibroma cell line (blue) and STS26T, sNF96.2, ST88-14, T265 and 90-8TL MPNST cell lines (red) (D) Western blot displaying BRD4 protein expression in cell line panel and HEK293T. ẞ-actin levels are shown as a loading control.
Fig 2.
Sensitivity of neurofibroma and MPNST cell lines to the BET bromodomain inhibitor JQ1.
(A) An in vitro cytotoxicity assay (SRB assay) was used to determine IC50 values (nM) for the BET bromodomain inhibitor JQ1 of neurofibroma and MPNST cell lines after a 72h exposure to the drug. Graphs show cell viability as a function of JQ1 concentration. Depicted is the average viability (n = 4) of a representative experiment. (B) Listing of calculated IC50 values and correlation plot, with BRD4 protein expression levels on the Y-axis and IC50 values for JQ1 on the X-axis. Pearson correlation coefficient is depicted in the graph.
Fig 3.
Expression level of EZH2 in human neurofibroma and MPNST samples and cell lines.
(A) qRT-PCR was used to determine mRNA levels of EZH2 in paired plexiform neurofibroma (NF, blue, n = 9) and MPNST (red, n = 9) formalin-fixed paraffin-embedded tumor samples, each pair being derived from the same NF1 patient. Asterisk indicates P<0.05. (B) qRT-PCR was used to determine mRNA levels of EZH2 in fresh frozen MPNST (red, n = 11), plexiform neurofibroma (blue, n = 7) and atypical neurofibroma (grey, n = 4). (C) qRT-PCR was used to determine mRNA levels of EZH2 in a cell line panel: Hs53.T neurofibroma cell line (blue) and STS26T, sNF96.2, ST88-14, T265 and 90-8TL MPNST cell lines (red). (D) Western blot displaying EZH2 protein expression in cell line panel and HEK293T. ẞ-actin levels are shown as a loading control.
Fig 4.
siRNA mediated knockdown of EZH2 and its effect on cell proliferation.
(A) Western blot showing the effect of EZH2 siRNA (si+) or a scrambled control siRNA (si-) on EZH2 protein levels in T265 and 90-8TL at 48h and 72 h post-transfection. (B) Cell proliferation monitored in time after transfection of T265 and 90-8TL with EZH2 siRNA (si+) or a scrambled control siRNA (si-). ẞ-actin levels are shown as a loading control.
Fig 5.
Expression level of TOP2A in human neurofibroma and MPNST samples and cell lines.
(A) qRT-PCR was used to determine mRNA levels of TOP2A in paired plexiform neurofibroma (NF, blue, n = 9) and MPNST (red, n = 9) formalin-fixed paraffin-embedded tumor samples, each pair being derived from the same NF1 patient. Asterisk indicates P<0.05. (B) qRT-PCR was used to determine mRNA levels of TOP2A in fresh frozen MPNST (red, n = 11), plexiform neurofibroma (blue, n = 7) and atypical neurofibroma (grey, n = 4). (C) qRT-PCR was used to determine mRNA levels of TOP2A in a cell line panel: Hs53.T neurofibroma cell line (blue) and STS26T, sNF96.2, ST88-14, T265 and 90-8TL MPNST cell lines (red). (D) Western blot displaying TOP2A protein expression in cell line panel and HEK293T. ẞ-actin levels are shown as a loading control.
Fig 6.
Sensitivity of neurofibroma and MPNST cell lines to doxorubucin.
(A) An in vitro cytotoxicity assay (SRB assay) was used to determine IC50 values (ng/ml) for doxorubucin of neurofibroma and MPNST cell lines after a 48h exposure to the drug. Graphs show cell viability as a function of doxorubucin concentration. Depicted is the average viability (n = 4) of a representative experiment. (B) Listing of calculated IC50 values and correlation plot, with TOP2A protein expression levels on the Y-axis and IC50 values for doxorubicin on the X-axis. Pearson correlation coefficient is depicted in the graph.