Abstract B48: A LIN28B-PDZ kinase axis promotes neuroblastoma metastasis
Authors: Chen Dongdong , Cox Julie , Annam Jayabhargav , Weingart Melanie , Essien Grace , Rathi Komal , Khurana Priya , Cuya Selma , Harenza Jo Lynne , Bosse Kristopher , Pilgrim Adeiye , Maris John M. , Schnepp Robert W. 
Published: August 2020

Background: Neuroblastoma, an aggressive cancer of the developing sympathetic nervous system, continues to cause significant morbidity and mortality, highlighting the need to identify novel therapeutic vulnerabilities. We previously demonstrated that the RNA binding protein (RBP) LIN28B is an oncogenic driver and induces neuroblastoma proliferation, in part by regulating a RAN GTPase-Aurora kinase A axis. LIN28B blocks the processing of the let-7 family of tumor suppressors and binds mRNAs directly. In addition to alterations in cell cycle/apoptosis, metastatic dissemination is a hallmark of cancer that is incompletely understood, and neuroblastoma exhibits a striking proclivity for widespread metastases. In these studies, we investigated how LIN28B shapes neuroblastoma metastasis.

Methods: We generated GFP-luciferase expressing neuroblastoma cell line models in which LIN28B levels were manipulated, injected these models into the tail veins of NSG mice, and tracked dissemination using an IVIS Spectrum system. We used gain- and loss-of-function approaches (siRNAs, shRNAs, microRNA mimetics) to manipulate transcripts of interest in neuroblastoma cells and measured effects on self-renewal, invasion, and downstream signaling. To discover LIN28B-associated pathways, we assessed clinically annotated mRNA expression datasets.

Results: Mice injected with LIN28B-depleted neuroblastoma cells exhibit a delayed onset of tumor metastasis, reduced tumor burden, and extended survival (103 days vs. 50 days, p&lt;0.0001), compared to mice bearing neuroblastoma cells expressing control scrambled shRNA. We next demonstrated that LIN28B promotes, and let-7 opposes, self-renewal and migration, two hallmarks of metastasis. As we discovered that Aurora kinase A is a novel LIN28B target, we speculated that LIN28B might positively regulate diverse oncogenic kinases to promote metastasis. We evaluated the TARGET dataset of neuroblastoma tumors and found LIN28B mRNA expression to be robustly correlated with PDZ-binding kinase mRNA expression (PBK; r=0.67; p=3.2 × 10-33), a kinase with roles in cell proliferation/survival, self-renewal, and metastasis, and is overexpressed in multiple malignancies. We demonstrated that LIN28B directly promotes, and let-7 opposes, the expression of PBK protein, and, indeed, that PBK is a novel and direct let-7 target. Moreover, we reveal that MYCN binds to the promoter of PBK and positively regulates PBK RNA and protein expression. Finally, PBK depletion mimics the effects of LIN28B depletion, with respect to self-renewal and invasion.

Conclusions: Taken together, our findings suggest that LIN28B/let-7 shapes neuroblastoma metastasis, in part through influencing PBK, a kinase not previously implicated in the pathogenesis of aggressive pediatric solid tumors. Current studies are defining whether PBK, a therapeutically tractable target for which clinically relevant inhibitors exist, represents a novel therapeutic vulnerability in metastatic neuroblastoma.

Citation Format: Dongdong Chen, Julie Cox, Jayabhargav Annam, Melanie Weingart, Grace Essien, Komal Rathi, Priya Khurana, Selma Cuya, Jo Lynne Harenza, Kristopher Bosse, Adeiye Pilgrim, John M. Maris, Robert W. Schnepp. A LIN28B-PDZ kinase axis promotes neuroblastoma metastasis [abstract]. In: Proceedings of the AACR Special Conference on the Advances in Pediatric Cancer Research; 2019 Sep 17-20; Montreal, QC, Canada. Philadelphia (PA): AACR; Cancer Res 2020;80(14 Suppl):Abstract nr B48.