Abstract 3196: MPP8 inhibits PRKN signaling through interactions with SIRT1 and ZEB1 to promote bladder cancer proliferation and metastasis

Abstract Background: Metastatic bladder cancer (mBC) remains incurable, and epigenetic dysregulation is associated with disease progression. We hypothesize that the H3K9me3 reader protein MPP8 complexes with class III histone deacetylase SIRT1 and transcription factor ZEB1 to drive proliferation and metastasis. Here, we explore how MPP8 knockdown (MPP8KD) disrupts MPP8-SIRT1-ZEB1 interactions, thereby altering differentially expressed genes (DEGs) and gene networks and abrogating proliferation and metastasis. Methods: UM-UC-3 mBC cells were used to compare MPP8KD and untransfected (MPP8WT) cells. RNA-seq quantified differential expression. DEGs were analyzed in R v4.1.1 using DESeq2.3. Gene enrichment was assessed with GSEA v4.2.3 (Hallmark sets) and IPA pathway analyses (Qiagen). Co-IP WBs evaluated MPP8-SIRT1-ZEB1 protein complexes. CellTiter-Glo™ evaluated differences in viability. Clonogenic assays assessed proliferation while migration/invasion assays evaluated metastatic potential. MPP8KD and MPP8WT cells were injected into NSG mouse flanks, and changes to both tumor volume and TBW were measured until tumors were ≥ 2 cm3. Results: Over-representation analysis of DEGs indicated MPP8KD reduced mBC cell growth and DNA replication, as well as increased inhibition of cell migration. GSEA identified downregulated E2F (NES=-1.9, q=0.02) and enrichment of TNFα signaling via NFKB gene sets (NES=1.71, q=0.01). IPA revealed activation of the Parkinson’s signaling pathway (z-score=1.54, P=1.1E-03), and consistent with GSEA analyses, predicted reduced proliferation and cellular stress responses. A custom IPA gene network of top DEGs included Parkinson’s signaling pathway genes (e.g., PRKN) and demonstrated that MPP8, SIRT1, ZEB1 all act upstream of pathway genes. Moreover, SIRT1 and PRKN were predicted to interact through protein-protein interactions, while MPP8KD directly increased PRKN expression (LFC=1.658, q=2.39E-03). Functionally, we demonstrated MPP8 forms a complex with SIRT1 and ZEB1 in MPP8WT cells, while the MPP8-SIRT1-ZEB1 interactions were lost in MPP8KD cells. MPP8KD reduced viability (44% vs. 84% after 48 h; P=0.0001), colony growth (13% vs. 36% plate covered with colonies at 10 d; P=0.0001), number of migrated cells (97 vs. 470 cells after 18 h; P<0.0001), and number of invading cells (44 vs. 130 cells after 18 h; P=0.02). By D+32 post-injection, all mice with MPP8WT cells were sacrificed (mean vol. 2,859 ± 761 mm3; n=8). At D+32, tumor volume was significantly smaller in MPP8KD1 (mean vol. 234 ± 447 mm3, P<0.0001; n=8) and MPP8KD2 mice (mean vol. 1,341 ± 785 mm3, P=0.002; n=8). At D+32, changes in TBW were not observed. Conclusions: Our data suggest the MPP8-SIRT1-ZEB1 axis is a key driver of mBC, and MPP8KD results in PRKN activation that could halt mBC proliferation and metastasis. Citation Format: Stephany Gonzalez Tineo, Ryan M. Kemper, Surya K. Tripathi, Daniel J. Crona. MPP8 inhibits PRKN signaling through interactions with SIRT1 and ZEB1 to promote bladder cancer proliferation and metastasis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 3196.