NSC12 targets WM cells within the context of the BM microenvironment both in vitro and in vivo. (A) Modulation of cell proliferation of BCWM.1 cells exposed to NSC12 (0 to 6 μM; 48 hours) cocultured in the presence or absence of WM patient-derived BMSCs was assessed by CellTiter-Glo. (B) BCWM.1 and MWCL.1 cells were cultured in the presence or absence of WM patient’s derived BMSC and exposed to NSC12 (0 to 6 μM; 6 hours). WM cells were then harvested, and cell lysates were subjected to western blotting, with the use of antibodies anti-p-FGFR3, -p-AKT, -p-GSK3β, -p-.ERK, p-STAT3, -p-Cofilin, p-SRC, and -GAPDH (C) Adhesion of BCWM.1, MWCL.1, MEC.1, WMWSU cells to WM patient-derived BMSCs exposed to NSC12 (0 to 6 μM; 4 hours) was evaluated by an in vitro adhesion assay, using Calcein AM labeling of WM cells, with degree of fluorescence measured on a spectrophotometer (λ485 nm to λ520 nm). (D-E) SCID/Bg mice were injected with BCWM.1-mCherry⁺/Luc⁺ cells and treated with either NSC12 (7.5 mg/kg, every other day) or vehicle control. Detection of tumor growth was performed by measuring bioluminescence imaging (BLI) intensity at different time points post-WM cell injection (days 7, 10, 14). P, P value. (F) WM cell BM colonization was evaluated ex vivo from harvested femurs, using immunostaining for human (h)-CD20. H.E., hematoxylin-eosin staining (×20). (G) BM mononuclear cells were harvested ex vivo from femurs, subjected to RNA extraction, and evaluated for MYD88, HCK mRNA levels by using qRT-PCR (2^−ΔΔCt), with normalization to GAPDH (*P < .05).