Multi-Omics Integration to Elucidate the Cis-Regulatory Network Re-Wiring upon NSD2 Degradation in t(4;14) Multiple Myeloma

• Published in: Blood 2024-11, Vol.144, p.1881-1881
• Main Authors: Hu, Bo, Modi, Hardik, Jankeel, Diana, Edwards, Jacob T., Zhao, Junfei, Bjorklund, Chad C., Jain, Gaurav, Hagner, Patrick R., Ortiz, Maria, Mo, Zhongying, Fontanillo, Celia, Zapf, Christoph W., Lopez-Girona, Antonia, Bence, Neil, Wang, Kai, Gandhi, Anita K., Rolfe, Mark, Mortensen, Deborah S., Groocock, Lynda
• Format: Article
• Language: English
• ISSN: 0006-4971
• DOI: 10.1182/blood-2024-203621

Introduction. The t(4;14) translocation activates nuclear receptor binding SET domain protein 2 (NSD2) to define a molecularly distinct subset of multiple myeloma patients with especially unfavorable prognosis. Despite the clear delineation of t(4;14) as a segment with NSD2 activation as the driver, an incomplete understanding of the mechanistic cascade towards myelomagenesis in these patients has been a major hurdle. We thus developed a tool CRL4CRBN E3 ubiquitin ligase-dependent ligand-directed degrader (LDD) that rapidly catalyzes proximity-induced degradation of NSD2 to facilitate the multi-omics characterization of NSD2's role in myeloma, uncovering direct impacts on both the local activity and long-range organization of cis-regulatory elements governing critical programs. Methods. t(4;14) and non-t(4;14) multiple myeloma preclinical models, at baseline and with NSD2-LDD treatment, were profiled using: CUT&RUN/ChIP-Seq (CTCF, H3K27ac/me3, H3K36me2/3, H3K4me1/3), Hi-C, ATAC-seq, RNA-seq, histone mass spectrometry. Soft agar colony formation, fibronectin cell adhesion, and FBS chemotaxis migration assays were used to validate the phenotypic consequences in vitro, whereas cell-line derived xenograft models were used to assess tumor control in vivo. Additionally, a co-culture system of myeloma cells with bone marrow stromal cells was used to evaluate both paracrine signaling and cell adhesion-mediated drug resistance. Results. NSD2 degradation-dependent therapeutic benefit was demonstrated across multiple in vivo models: up to 73% tumor volume reduction in mice bearing subcutaneous KMS34 tumors as well as intrafemoral engraftment of luciferase-labelled KMS34 cells that not only saw significant survival benefit (p = 0.034) but also absence of extra-nodal growth - contrasting CNS-resident and contra-lateral metastases in the vehicle group. Seeing that the in vivo efficacy of NSD2 degradation coincided with deep NSD2 degradation (-94%) as well as loss of its direct catalytic product H3K36me2 (-74%), we turned to in vitro models to elucidate NSD2's trans-omics mechanistic cascade. Consistent with H3K36me2 loss (9x) and H3K27me3 gain (3x) being the most significant changes among the 40 histone post-translational modifications detected by mass spectrometry, cells treated with NSD2-LDD exhibit global restoral of transcriptional control across multiple modalities with: (1) 3x excess number of down-regulated genes based on RNA-seq, (2) 6x more sites with reduced a