Identification of CDKN1A as a Candidate Therapeutic Target in Peripheral Artery Disease Progression: Integrative...

Therapeutic strategies for advanced Peripheral Artery Disease (PAD), including angiogenic gene therapy trials, have yielded limited clinical benefit, partly because target selection has relied on empirical biological reasoning rather than genetic causal evidence. Since the progression from Intermittent Claudication (IC) to Critical Limb Ischemia (CLI) represents a critical stage of PAD progression, identifying genetically supported causal targets underlying this transition is essential for developing effective molecular interventions.

An integrative analytical framework was applied, combining bulk transcriptomics, Weighted Gene Co-expression Network Analysis (WGCNA), differential expression analysis, Protein- Protein Interaction (PPI) network construction, and machine-learning-based feature selection. Candidate genes were further characterized using single-cell RNA sequencing (scRNA-seq) and validated in an in vivo murine Hindlimb Ischemia model (HLI). Immune infiltration was assessed by CIBERSORT. Genetic causal evidence was obtained through GWAS colocalization and bidirectional Mendelian Randomization (MR). Computational gene perturbation analysis was performed to predict downstream transcriptional consequences. Phenome-Wide Association Study (PheWAS) and drug-gene interaction analyses were conducted to explore pleiotropy and therapeutic relevance.

Multi-stage screening consistently identified CDKN1A as a central hub gene associated with ischemic severity. At single-cell resolution, CDKN1A was broadly upregulated across multiple cell types within the ischemic niche and positively correlated with M2 macrophage infiltration. This ischemia-dependent induction was confirmed at the protein level in vivo. Integrative genetic analyses, including GWAS colocalization (PP.H4 = 0.90) and bidirectional MR, provided evidence supporting a causal link between genetically predicted CDKN1A expression and PAD risk, with no evidence of reverse causation. Computational gene perturbation analysis identified downstream transcriptional programs associated with extracellular matrix remodeling, TGF-beta signaling, and angiogenesis. PheWAS showed no genome-wide significant associations across 19 phenotype categories, and drug-gene interaction analysis identified resveratrol as a candidate modulator.

The current findings indicate that CDKN1A-mediated stress response represents a key mechanism in PAD progression, linking ischemic injury to maladaptive remodeling and supporting its relevance as a genetically supported candidate therapeutic target in PAD.

Our findings provide convergent transcriptomic, cellular, and genetic evidence supporting CDKN1A as a genetically supported candidate causal target associated with PAD progression. These results suggest that CDKN1A-mediated cell-cycle arrest and associated stress-response programs are associated with ischemic tissue remodeling, providing a rationale for future gene-directed or pharmacological intervention strategies aimed at attenuating the transition to CLI.