Nonactin

Cell lines

MTC-derived TT cells

Preparation Method

The MTC-derived TT cells were cultured in DMEM/F12 medium enriched with 15% heat-inactivated fetal bovine serum (FBS) and maintained under a humidified atmosphere with 5% CO₂ at 37°C. Cells were plated at 7,500 cells per well in a 96-well plate and allowed to adhere overnight. Following this, they were exposed to various concentrations of Nonactin (0.12-0.5µM) for up to 72h. To evaluate cell viability, a solution of 0.33mg/mL MTS dye mixed with 200µg/mL phenazine methosulfate (PMS) was used. Absorbance readings were taken at 490nm. Caspase-3 activity in TT cells was evaluated utilizing the ApoAlert Caspase Fluorescent Assay kit. And cells were collected for PCR and Western blot analyses.

Reaction Conditions

0.12-0.5µM; 72h

Applications

Treatment of medullary thyroid carcinoma TT cells with Nonactin reduces RET protein and mRNA level, suppresses pAKT/pmTOR signaling, lowers cyclin D1/E, E2F1 and BCL-2, increases γH2AX and cleaved PARP, and elevates caspase-3/7 activity.

Animal models

BALB/cA Jcl-nu mice

Preparation Method

Specific pathogen-free female nude mice (BALB/cA Jcl-nu) were used in this experiment. β-catenin-mutant HCT 116 cells (2×10⁶cells/mL) were inoculated subcutaneously into the axillar region of the nude mice on Day 0. Nonactin was dissolved in 10% dimethylacetamide, 10% NIKKOL HCO60, and saline and given daily to the animals by intraperitoneal administration (100mg/kg). Tumor-bearing nude mice were randomly grouped (six mice/group), and administration of nonactin started on Day 9 to Day 12. Tumor volumes were calculated using a microgauge according to the following equations: Tumor volume (mm³)=1/2×(tumor length)×(tumor width)². TUNEL staining of xenograft tumor tissue was performed using the FragEL DNA Fragmentation Detection Kit according to the manufacturer’s protocol. Tissue sections were viewed at 100×magnification, and images were captured with a digital camera.

Dosage form

100mg/kg/day; i.p.; 4 days

Applications

Nonactin (100mg/kg/day; i.p.; 4 days) induced marked tumor regression and robust TUNEL-positive apoptosis in β-catenin-mutant HCT 116 xenograft mice without significant body-weight loss^[7].

References:
[1] Alqahtani T, Alsubait A, Aloumi M, et al. A novel role for nonactin: interfering with G-quadruplex in RET-driven medullary thyroid cancer. BMC Cancer. 2024;24(1):1569.
[2] Shikata Y, Kiga M, Futamura Y, et al. Mitochondrial uncoupler exerts a synthetic lethal effect against β-catenin mutant tumor cells. Cancer Sci. 2017;108(4):772-784.