The Study Of The Effect And Mechanism Of A Novel Agent DMAMCL On Tumor Growth In Rhabdomyosarcoma

Purpose: Rhabdomyosarcoma(RMS)is a common soft tissue sarcoma which making it the third most prevalent extracranial solid tumor in children with poor survival.New treatment approaches are urgently needed to improve the treatment efficacy in RMS patients.DMAMCL is a novel agent from Asteraceae family,which tested in phase Ⅱ clinical trials in Australia and in China in adult glioma.In this study,the effect of DMAMCL on tumor growth,alone or in combination with vincristine(VCR)or Epirubicin,was evaluated in vitro and in vivo in RMS.Experimental Design: Five RMS cell lines(RD,RH18,RH28,RH30 and RH41)were used.The anti-tumor effect of DMAMCL,alone or in combination with VCR or Epirubicin,was studied by MTS assay or with IncuCyte Zoom through dynamically live cell imaging in vitro,and further validated by assessing the tumor growth in nude mice inoculated with RMS cells in vivo.The caspase-3/7 activities and cell-cycle changes after DMAMCL treatment were investigated.Small interfering RNAs(siRNA)of pro-apoptosis protein Bim were used to study the role of Bim in DMAMCL-induced cell death.The expression of Bim was measured by Western Blotting or by RT-qPCR.Results: In vitro,DMAMCL treatment induced a dose-dependent cell death which could be blocked by a pan-caspase inhibitor Z-VAD-FMK in the five RMS cells;The percent of cells in the SubG1 phase and the activities of caspase 3/7 increased after DMAMCL treatment;A combination of DMAMCL with VCR or Epirubicin induced significantly increased-cell death compared to each reagent alone.In vivo,DMAMCL inhibited the tumor growth and prolonged the survival of mice bearing xenograft RMS tumors(RD,RH18,RH30 and RH41)at the dose of 75mg/kg or 100 mg/kg.Compared to treatment with DMAMCL or VCR,a combination of DMAMCL with VCR caused a significant inhibition of tumor growth(RD and RH41),even after the treatment was stopped.An increased expression of Bim after DMAMCL treatment was detected in RMS cells and xenograft tumor tissues.Transfection of Bim siRNA into RMS cells blocked the increase of DMAMCL-induced Bim and partially attenuated the DMAMCL-induced cell death.Conclusion: DMAMCL had an anti-tumor growth effect in vitro and in vivo.A combination of DMAMCL with chemotherapeutic drugs significantly increased the treatment efficacy.Bim may be a potential target that mediated the DMAMCL effect.Our study supports future clinical evaluation of DMAMCL in combination with conventional chemotherapy.