Dual targeting of DHODH and SLC29A1 overcomes metabolic resistance and enhances antitumor immune responses in PDAC

Pancreatic ductal adenocarcinoma (PDAC) displays profound metabolic plasticity that sustains rapid proliferation through heightened demands for nucleotide biosynthesis. Here, we characterize HOSU-53 (JBZ-001), a dihydroorotate dehydrogenase (DHODH) inhibitor demonstrating potent antitumor activity in preclinical PDAC models and define the metabolic determinants of sensitivity and resistance. In HOSU-53-sensitive PDAC cells, DHODH inhibition induces G₂/M cell cycle arrest and triggers apoptotic and ferroptotic cell death. In contrast, resistant cells evade metabolic stress by activating compensatory nucleoside transport and macropinocytosis pathways. Integrated functional and genetic analyses reveal that nucleoside salvage via the equilibrative nucleoside transporter SLC29A1 constitutes a dominant resistance mechanism, enabling uptake of extracellular nucleosides to bypass de novo pyrimidine synthesis. Unbiased metabolomic profiling demonstrates that combined inhibition of DHODH and SLC29A1 precipitates mitochondrial dysfunction, oxidative stress, lipid remodeling, and amino acid depletion, culminating in metabolic collapse and energetic failure. Consistent with these findings, dual inhibition markedly suppresses tumor growth and metastatic burden in xenograft and syngeneic orthotopic PDAC models while reprogramming the tumor microenvironment toward a pro inflammatory, immune activated state. Furthermore, co targeting DHODH and SLC29A1 synergizes with anti-PD 1 immune checkpoint blockade, enhancing immunotherapeutic efficacy. Together, these results establish dual targeting of de novo and salvage nucleotide metabolism as a compelling strategy to overcome metabolic resistance, remodel tumor-immune interactions and potentiate therapeutic responses in PDAC.