Abstract 6449: Unique site-specific phosphorylation of PML co-ordinates disease aggravation of neuroblastoma

Abstract Neuroblastoma a fatal infant malignancy accounts for 15% of pediatric cancer deaths. The standard treatment for children with neuroblastoma is intensive and include induction, consolidation, maintenance, and salvage therapy. Despite the largely evolving molecular targets for maintenance therapy, retinoic acid (RA) remains the most promiscuous therapy. Restoration of promyelocytic leukemia protein nuclear body (PML-NB) is a crucial cofactor for RA mediated signal transduction, potentially improving the treatment response and outcomes. Phosphorylation at PAS-NA is one of the (if not the only one) crucial phosphorylation site of PML that direct PML degradation through ubiquitin pathway thereby modulating abundance and stability of PML-NB. Herein we investigated the vital association between phosphorylation of PML at PAS-NA and the clinical outcomes of neuroblastoma patients (n=124). To study the spatio-temporal expression of PML and pPML (PAS-NA) at specific tumor sites determined by pathologists, we custom archived the tumors in triplicate in a tissue microarray and effectuated a chromogenic IHC using our custom synthesized antibodies. The analytical endeavor was accomplished by using the modified Multiplex IHC 3.4.9 module integrated into Indica Lab’s HALO platform. We observed a strong and definitive inverse correlation of PML to pPML (PAS-NA) expression in the tumor tissues. Negative correlation of pPML (PAS-NA) with patient survival (overall-, relapse-free, progression-free survival) indicating phosphorylation of PAS-NA as a key regulatory mechanism mediating the stability of PML-NBs and impacting the survival outcomes. While PML exhibits an inverse association to disease progression, we observed a proportional increase in pPML (PAS-NA) to the advanced disease stages in neuroblastoma. Furthermore, increased phosphorylation of PML at PAS-NA in patients presenting with metastatic disease or relapse state also emphasizes the critical role of its site-specific phosphorylation in regulating disease aggravation, dissemination and recurrence in neuroblastoma which is mediated by perpetuating PML-NB stability. Together, these results unveil a prominent molecular mechanism that mediates the evolution of neuroblastoma. Critically, these outcomes would have major implications on developing novel molecular targeted therapy and would pave way for achieving desirable clinical outcomes for children affected with this disease. Funding: This work was partially or in full, funded by Department of Defense CA-210339; OCAST-HR19-04; NIH-P20GM103639 to Dr. Aravindan. The work was also supported by NCI Support Grant [P30CA225520], NIGMS [P30GM154635] and a grant from the Oklahoma Tobacco Settlement Endowment Trust [R23-03] to OU Health Stephenson Cancer Center. Citation Format: Sreenidhi Mohanvelu, Poorvi Subramanian, Madhi Oli Ramamurthy, Sheeja Aravindan, Natarajan Aravindan. Unique site-specific phosphorylation of PML co-ordinates disease aggravation of neuroblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 6449.