NRF1 enhancer inhibition sensitizes MM cells to bortezomib in vitro and in vivo. (A) qRT-PCR of Kms18 treated or not with 5 nM BTZ and analyzed for eNRF1 expression. Data are presented as mean ± SD of 3 independent experiments and error bars represent the SD. Student t test ∗∗P < .01. (B) Evaluation of mRNA level of eNRF1 in 6 patients with NDMM treated or not treated with 5 nM of BTZ for 24 hours. Student t test ∗∗∗P < .005. (C) Evaluation of Kms27 cell line proliferation transiently transfected with siControl or si-eNRF1 after BTZ treatment or no treatment. Data are presented as mean ± SD of 3 independent experiments with values normalized to actin expression. Error bars represent the SD. Student t test ∗∗∗P < .005; ∗∗∗∗P < .001. (D) Schematic representation of in vivo experiments for silencing the NRF1 enhancer using ASO-eNRF1. (E) Tumor volumes over time in ASO scramble (blue, n = 5), ASO scramble + BTZ (purple, n = 5), ASO-eNRF1 (pink, n = 5), and ASO-eNRF1 + BTZ (green, n = 5) were calculated from caliper measurements every 3 to 4 days. Diamonds represent the timing of ASO administration. ANOVA test ∗∗∗∗P < .001. (F) Representative excised tumors (left) and tumor weight (right) at the termination of the experiments. The Kruskal-Wallis test followed by the Wilcoxon signed-rank test was applied ∗∗∗P < .001. (G) Survival analysis of previously described mice. N°, number of cells.