Effects of novel pyrrolomycin MP1 in MYCN amplified chemoresistant neuroblastoma cell lines alone and combined with temsirolimus

Abstract

Background: The activity of MP1, a pyrrolomycin, was studied in MYCN amplified neuroblastoma (NB) alone and combined with temsirolimus (TEM). Methods: Activity of MP1 was tested in MYCN amplified (BE-2c, IMR) and non amplified (SKN-AS) NB cells. The effect of MP1 on MYCN, MCL-1, cleaved PARP, LC3II/LC3I, bcl-2, BAX, and BRD-4 were determined by western blot and RNAseq. The effect of MP1 on metabolism, mitochondrial morphology, and cell cycle was determined. Toxicology and efficacy of MP1 plus TEM were evaluated. Results: The IC50 of MP1 was 0.096 μM in BE-2c cells compared to 0.89 μM in IMR, and >50 μM in SKN-AS. The IC50 of MP1 plus TEM in BE-2c cells was 0.023 μM. MP1 inhibited metabolism leading to quiescence and produced a decline in cell cycle S-phase. Electron microscopy showed cristae loss and rounding up of mitochondria. Gene and protein expression for MYCN and MCL-1 declined while LCII and cleaved PARP increased. Protein expression of BAX, bcl-2, and BRD-4 were not significantly changed after MP1 treatment. The in-vivo concentrations of MP1 in blood and tumor were sufficient to produce the biologic effects seen in-vitro. MP1 plus TEM produced a complete response in 3 out of 5 tumor bearing mice. In a second mouse study, the combination of MP1 and TEM slowed tumor growth compared to control. Conclusions: MP1 has a potent inhibitory effect on the viability of MYCN amplified NB. Inhibition of metabolism by MP1 induced quiescence and autophagy with a favorable toxicology and drug distribution profile. When combined with TEM anti-tumor activity was potentiated in-vitro and in-vivo.