Abstract Many human cancers overexpress the E3 ubiquitin ligase MDM2. The cancer genome atlas (TCGA) found that increased MDM2 expression is one of four gene pathways that correlate with all breast cancer subtypes. An increasing body of literature strongly suggests that overexpression of MDM2 can drive breast oncogenesis in the presence of mutant p53 (mtp53), as well as through molecular pathways that do not require wild-type p53 degradation. Therefore, the canonical wild-type p53 degradation pathway by MDM2 is clearly not the only important pathway. MDM2 expression has been shown to influence breast cancer cell invasion and metastasis. One potential mechanism for this is through the activation of SLUG (a transcription repressor of E-Cadherin) and the process of E-cadherin degradation. To investigate the p53-independent influence of MDM2 we engineered human breast cancer clones with inducible shRNA to knockdown MDM2 in MDA-MB-231 (mtp53 R280K), T47D (mtp53 L194F) and MCF-7 (wild-type p53) cells. Wound healing assays showed that MDM2 knockdown in all three cell lines reduced cell migratory capacity without altering p53 protein levels. In T47D.shmdm2 cells, a decrease in SLUG expression was observed with MDM2 knockdown indicating SLUG is a p53-independent target of MDM2. The knockdown of MDM2 also decreased slug and mmp9 mRNA detected by quantitative RT-PCR. To determine the biological significance of MDM2 in a system to recapitulate breast cancer architecture we examined T47D.shmdm2 cells in a 3-dimensional (3D) cell culture model. Importantly, the depletion of MDM2 in T47D.shmdm2 cells grown in 3D culture promoted luminal clearance. Our findings suggest that MDM2 can influence SLUG and MMP9 levels associated with breast cancer metastasis through a mechanism that does not require degradation of wild-type p53. Moreover our work suggests that MDM2 may promote changes to breast cancer architecture that drive metastasis. Acknowledgements. This work was supported by a grant to JB from the Breast Cancer Research Foundation. Citation Format: Chong Gao, Nandini Kundu, Jill Bargonetti. MDM2 promotes p53-independent breast cancer metastatic phenotypes. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1220. doi:10.1158/1538-7445.AM2015-1220