Supplementary Table S1-S5.docx

Duchenne muscular dystrophy (<i>DMD</i>) gene responsible for the fatal inherited neuromuscular disorder Duchenne muscular dystrophy (DMD) is regarded as a likely tumor suppressor in all major cancers. Spindle cell sarcoma SCS) is very rare in muscular dystrophy (mdx) model mouse or DMD patients while Rhabdomyosarcoma (RMS) has been readily detected. Six aged mdx mice from our colony developed tumors (3 SCS and 3 RMS) spontaneously. Here, we performed a comparative metabolomic analysis of SCS and RMS to understand the rarity of SCS from a metabolomic point of view since metabolism is considered significant in tumor development and progress.Among 25 significantly altered metabolites in SCS compared to RMS, the alanine, aspartate and glutamate metabolic pathway was very impacted with four components that were significantly downregulated in SCS.Functional assays with alanine supplementation in the culture medium of SCS tumor derived primary cell line indicated increase in growth, EMT and invasion. Moreover, chemical knockdown of the alanine transporter Slc1a5 with O-Benzyl-O-Serine demonstrated growth reduction. Whereas, Slc1a5 knockdown with specific shRNA showed suppressed EMT and invasion. We also observed that reduction in intracellular alanine triggered the energy sensor AMPK which functions in modulating the energy utilization within the cell.We propose that differential metabolic secretion could be a driving force for tumor development.