Asparagine bioavailability is critical for maintaining MYC protein levels post-transcriptionally in lymphoid malignancies, rendering its depletion as a potential strategy to target MYC-driven cancers. Sankalp Srivastava1,2, Jiang Jie2, Changlin Wan3, Jaganath Misra1, Chi Zhang3, Ron Wek1, and Ji Zhang1,2 1Department of Biochemistry and Molecular Biology, Indiana University School of Medicine, 2Herman B. Wells Center for Pediatric Research, Indiana University School of Medicine, 3Center for Computational Biology and Bioinformatics L-Asparaginase has been used for decades to treat acute lymphoblastic leukemia (ALL) successfully. This bacterial enzyme-derived chemotherapeutic exploits the auxotrophic nature of ALL cells for the amino acid asparagine, and depletes it in circulation. Recent publications have shown the importance of this non-essential amino acid asparagine in rescuing tumor cell survival and proliferation during glutamine starvation, a situation frequently observed in the tumor microenvironment. We recently showed that ALL cells are sensitive to L-Asparaginase therapy due to the lack of expression of asparagine synthetase (ASNS), the rate-limiting enzyme responsible for the de novo biosynthesis of asparagine. However, certain ALL cells can induce ASNS expression following L-Asparaginase treatment to cause therapeutic resistance. This process requires the activation of the General Control Nonderepressible 2 (GCN2) mediated amino acid response pathway and the promoter demethylation of the ASNS gene. Using our ALL model, we have discovered a novel regulation of MYC, a key oncogenic driver in ALL, governed by asparagine bioavailability. Asparagine withdrawal from the growth medium causes an acute depletion of MYC protein levels without changing the MYC mRNA levels. Our data also indicates that this depletion of MYC protein levels is predominantly at the level of translation and not degradation. We have also found that the Mechanistic Target of Rapamycin complex 1 (mTORC1) and GCN2 pathways that intersect amino acid bioavailability and translation control, are not responsible for suppressing MYC mRNA translation following asparagine depletion. Using a novel polysome profiling assay coupled with a high throughput RNA sequencing approach, we aim to identify other mRNAs whose translation is similarly regulated by asparagine depletion as MYC. In addition, our data suggest that inhibition of ASNS expression can be used to limit asparagine bioavailability and therapeutically target MYC-driven cancers in vivo.