Alternative splicing as a multilayered regulatory hub in pancreatic ductal adenocarcinoma

Alternative splicing is a crucial regulator of transcriptomic plasticity in pancreatic ductal adenocarcinoma (PDAC), and its dysregulation contributes to tumor progression, therapeutic resistance, and disease heterogeneity. While mutations in spliceosomal components or splice-site sequences can directly affect splicing, evidence increasingly suggests that aberrant splicing in PDAC results more often from the coordinated disruption of multiple regulatory layers. These layers include chromatin organization, transcriptional dynamics, epigenetic and epitranscriptomic mechanisms, and RNA-binding protein networks. Together, these interconnected processes reshape the transcript isoform landscape, producing oncogenic variants that facilitate tumor progression, metabolic adaptation, immune evasion, and therapeutic resistance. This review synthesizes the current knowledge on the multilayered regulation of alternative splicing in PDAC, distinguishing mechanisms that directly determine splice-site selection from those that act indirectly by regulating splicing factors, and from RNA regulatory layers that interact with, but can operate independently of, alternative splicing. Focusing on the dynamic interplay among co-transcriptional chromatin regulation, transcriptional kinetics, RNA modifications, and non-coding RNA-mediated regulatory networks, we examine how these mechanisms collectively remodel the splicing landscape to support tumor plasticity and malignant progression, while critically assessing the strength of PDAC-specific evidence and highlighting areas where mechanistic understanding remains incomplete. Additionally, we highlight emerging opportunities for isoform-level biomarker development and splicing-directed therapeutic strategies, while outlining the major challenges that must be addressed for their successful clinical translation.