Abstract 3761: Ubiquitin carboxyl-terminal esterase L1 (UCHL1) role in mediating MYC family of transcription factors and regulation of cell cycle in neuroblastoma cancer stem-like cells.

Proceedings: AACR 104th Annual Meeting 2013; Apr 6-10, 2013; Washington, DC Neuroblastoma (NB) is the most common extra-cranial nervous system malignancy of infants and children under 5 years of age. NB likely arises from the neural crest cells of the developing sympathetic system. The majority of NB occurs in or around the adrenal glands, which are located above the kidney. Patients with high risk NB have a long-term survival of less than 40% and over half of the survivors will relapse. Emerging evidence supports the concept that a small pool of cells displaying stem-like properties is the underlying cause for clinical relapse; therefore, targeting these cancer stem-like cells (CSC) may lead to improved therapies. Previous work identified that ubiquitin carboxyl-terminal esterase L1 (UCHL1) was differentially expressed in the CSC pool of human NB cell line, SK-N-BE(2), suggesting a potential connection between UCHL1 and CSCs. qRT-PCR revealed that inhibiting UCHL1 hydrolase activity in the CSC fraction lead to differential expression of MYC and MYCN. MYCN amplification has been correlated with poor prognosis and is associated with the aggressive disease phenotype. Our study is focused on determining how UCHL1 mediates MYCN and its downstream target genes in the aggressive form of this malignancy. We performed in vitro analyses on three NB cell lines: SK-N-BE(2), SK-N-AS, and SK-N-DZ. These cell lines vary in P53 mutation and MYCN amplification status. Sphere formation assay was used to enrich for CSCs. We used small molecule inhibitor, LDN-57444, to block UCHL1 hydrolase activity as well as knocking down UCHL1 expression by transduction with lentivirus expressing short hairpin RNA (shRNA) specific to UCHL1. The effect of LDN-57444 and UCHL1 knockdown (KD) on cellular viability was measured by MTS assay, and sphere formation (readout for self-renewal) was determined by quantification of spheres. Cell cycle distribution was analyzed by FACS analysis. The regulation of MYC and MYCN and its downstream targets by UCHL1 in the knocked down cells was quantified by qRT-PCR. Decreased gene expression of MYC and MYCN was observed following knockdown of UCHL1. The effect of knocking down UCHL1 on targets downstream of MYC and MYCN are currently being investigated. We investigated the in vivo therapeutic effect of UCHL1 inhibition using a xenograft mouse model, and this study is ongoing. Our data suggest that UCHL1 may function by mediating MYCN and/or MYC and their downstream targets at the level of transcription. It may play a role in promoting tumor cell survival, potentially by maintaining the CSC population through the cell cycle. Citation Format: Judy C. Chang, Jaclyn Y. Hung. Ubiquitin carboxyl-terminal esterase L1 (UCHL1) role in mediating MYC family of transcription factors and regulation of cell cycle in neuroblastoma cancer stem-like cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3761. doi:10.1158/1538-7445.AM2013-3761