Project Details
Projekt
In vivo models for the functional analysis of YB-1 in multiple myeloma
Applicants
Professor Dr. Ralf C. Bargou; Dr. Kurt Bommert
Subject Area
Hematology, Oncology
Term
from 2009 to 2018
Project identifier
Deutsche Forschungsgemeinschaft (DFG) - Project number 100308792
YB-1 is a DNA/RNA binding protein implicated in the regulation of DNA transcription and mRNA translation. Nuclear localization of the protein has been correlated to patient prognosis in breast carcinoma and non-small cell lung carcinoma. The underlying hypothesis of this project is that YB-1 plays a critical role in both early tumor development as well as maintenance of malignant growth in multiple myeloma (MM). In studying this question we have implemented several different mouse models and tumor cell lines, each of which characterizes different aspects of plasma cell (PC) neoplasia ranging from non-malignant PC to dedifferentiated PC tumors. By working from both ends of the disease spectrum we aim to identify therapeutic targets for this painful and deadly illness.To study the role of YB-1 in the malignant transformation of PCs we use the F.YB-1 knock-in model where the YB-1 transgene is under a late B cell promoter. Within the first 1½ years these animals did not yet show any PC tumor development. However, an excess of PCs with normal morphology was found in the bone marrow and gut villi relative to wild-type littermates. Hence, YB-1 alone is not responsible for the disease pathogenesis; therefore, other oncogenes are (additionally) required for malignant PC transformation. Based on these new findings we hypothesize that co-operation of YB-1 with MYC and/or other YB-1- independent deregulated signaling pathways (NOTCH1 and CCND1/2) is required. To address these questions we will overexpress specified proteins in wild-type and transgenic spleenocytes from F.YB-1 animals and analyze tumor development after reconstitution. We have shown that YB-1 is overexpressed in MM and that it contributes to tumor cell proliferation, survival and drug resistance in vitro. Most recently, we have demonstrated that MYC expression correlates with YB-1 expression in extramedullary PC tumors (human and mouse) and that YB-1 functions as a critical regulator of mRNA translation in malignant PCs. In human MM cell lines (HMCLs) MYC protein expression depends on YB-1-mediated mRNA translation while MYC is involved in YB-1 gene transcription. Notably, knockdown of either of these proteins induces apoptosis in HMCLs demonstrating that this transcriptional/- translational circuit is critical for the survival of dedifferentiated aggressively growing MM cells. These data have since been expanded to include in vivo analyses of YB-1 function. Using a murine model for MM (employing the MOPC315.BM.luc cell line), a partial knockdown of YB-1 delayed tumor growth in vivo, suggesting a critical role for YB-1 in tumor maintenance. These experiments led to the hypothesis that YB-1-dependent translation is critical for dedifferentiated aggressively growing MM cell survival both in vitro and in vivo. To test this hypothesis, we will use a zinc-finger nuclease-based strategy to produce a YB-1 null MM cell line and use conditional YB-1 expression to determine if the protein is critical for the survival of aggressively growing dedifferentiated tumor cells. Furthermore, we will identify YB-1-dependent translationally regulated mRNAs/proteins potentially mediating its tumorpromoting effects using Click-SILAC technology. Suitable candidates will then be functionally characterized to test their contribution to tumor maintenance using a small shRNA in vivo screen. We believe that together these approaches will help to clarify the role of YB-1 in myelomagenesis and that they also hold promise to identify critical YB-1-dependent mediators of survival that could serve as therapeutically relevant proxies for the as yet undruggable YB-1 protein itself.
DFG Programme
Clinical Research Units
