Kynurenine Metabolism Mediates Tumor Progression in Renal Cell Carcinoma

Renal cell carcinoma (RCC) is characterized by dysregulation of the kynurenine pathway (KP), which converts tryptophan to NAD+ while generating immunomodulatory metabolites. Therapeutic efforts have focused on inhibiting IDO1 at the pathway entry point, but the functional consequences of targeting downstream KP enzymes remain poorly characterized. We used CRISPR/Cas9 to generate knockouts of three KP enzymes in the RENCA murine RCC model--kynurenine 3-monooxygenase (KMO), quinolinic acid phosphoribosyltransferase (QPRT), and 3-hydroxyanthranilic acid dioxygenase (HAAO)--and evaluated effects on cell migration, colony formation, tumor burden, metastasis, and survival. KMO and QPRT knockouts consistently reduced migratory capacity and colony size in vitro. However, in vivo effects were distinct: while QPRT knockout reduced tumor burden, KMO knockout did not. Notably, we did not detect metastasis in female mice receiving KMO knockout RENCA cells. HAAO knockout produced divergent effects, increasing migration and colony size in vitro, but reducing tumor burden and metastasis in vivo. Mice challenged orthotopically with all three knockout cell lines had significantly extended survival compared to mice receiving wild type cells. These results indicate that individual KP enzymes exert distinct, context-dependent effects on RCC progression. The enhanced in vitro aggressiveness coupled with reduced in vivo tumorigenicity observed in HAAO knockout RENCA cells illustrates that cell culture phenotypes do not reliably predict tumor behavior, particularly when perturbing metabolic pathways with pleiotropic effects. Our findings suggest that targeting specific KP enzymes warrants further investigation as a therapeutic strategy in RCC.