Characterization of TFDP1 As Novel Regulatory Gene in Multiple Myeloma

Abstract 569

Alteration in expression and function of transcription factors (TF) has been frequently associated with neoplastic transformation and hence represent important target for cancer therapy. We evaluated expression profile of 172 newly-diagnosed uniformly-treated multiple myeloma (MM) patients. Our oncogenomic analysis of this large annotated patient samples identified TFDP1 as one of the most important gene affecting both overall and event-free survival in MM: elevated DP1 expression was a marker predictive of adverse clinical outcome. Importantly, association of DP1 protein partners, E2F1 and RB, was not observed with poor clinical outcome. This discordance between E2F1 and DP1 levels provides the significance to further evaluate role of DP1 in MM. We have observed that expression of DP1 is pronounced and mostly nuclear in all MM cell lines and primary cells and it is further markedly upregulated upon MM-bone marrow stromal cell (BMSC) interaction. Moreover, RNAi-mediated suppression of DP1 expression inhibited MM cell growth independently of pRB and p53 status, inducing cell cycle arrest and further apoptosis, revealing that E2F/DP activity is indeed essential for MM cell proliferation and survival. Interestingly, interaction with BMSC was able to partly overcome the inhibitory effect of DP1 silencing on MM cell growth. We next evaluted differential DP1 and E2F1 DNA-binding regions and targets in MM1S and U266 MM cells using chromatin immunoprecipitation with E2F1 and DP1 specific antibodies followed by genome wide sequencing (ChIP-Seq) along with gene expression profiling (GEP) after DP1 and E2F1 silencing in same MM cell lines. E2F1 and DP1 direct targets in myeloma have being identified by integrating differential GEP information with the bound target genes defined by ChIP-seq, providing insight into DP1 and E2F1 target genes and function, and identifying exclusive DP1 binding genes in MM with potential functional consequences. We have identified 1392 common genes, 169 E2F1 genes and 221 of distinct targets of DP1 in MM. We have used same approach to identify overlapping and distinct miR targets of DP1 and E2F1.This analysis is providing insight into potential DP1 targets that may predict poor clinical outcome. As interactions between DP1 with E2Fs is central to the regulation of its activity, disruption of this interaction represents a potential therapeutic target. We have investigated a polypeptide corresponding to residues 163 to 199 of DP1 which disrupt DP1-E2F interactions and observed inhibition of MM cell proliferation as evaluated by [3H]-Thymidine uptake after treatment with the peptide in a dose and time dependent manner. In conclusion, we have validated DP1 as an important functional gene in MM, and identified, using ChIP-seq, DP1 - E2F1 regulated targets with potential functional implications. Disruption of this interaction provides a novel therapeutic approach for clinical application of DP1 targeted therapy in MM.