Elevated systemic phenylalanine disrupts glutamate-dependent STING signaling in neutrophils and impairs cancer immunotherapy

Immune checkpoint inhibitors (ICIs) have transformed cancer therapy, yet acquired immune resistance remains a major barrier to durable response. Here, we identify a previously unrecognized immunometabolic axis in which elevated systemic phenylalanine (Phe) - driven by dietary intake or ICI-induced hepatic dysfunction - promotes immune evasion. Mechanistically, ICI-induced IL-6 and IFN-γ trigger proteasomal degradation of phenylalanine hydroxylase (PAH) via a TOPK-HECTD1 pathway in hepatocytes, leading to increased circulating Phe. Excess Phe suppresses antitumor immunity by impairing STING activation in tumor-associated neutrophils (TANs), through antagonism of glutamate-GRIK1-CaMKIV signaling. We uncover a key STING phosphorylation site (S324/326) essential for ER-to-Golgi trafficking and type I interferon responses, which is disrupted by Phe. These effects are independent of cGAS-cGAMP and define a glutamate-sensing checkpoint in neutrophil immunity. Therapeutically, restoring PAH function using liver-targeted synthetic RNA mimetics of HULC - a noncoding RNA shown to allosterically enhance PAH enzymatic activity - or activating GRIK1 with SYM2081 reprograms the tumor microenvironment, enhances CD8⁺ T cell function, and overcomes ICI resistance. This work reveals hepatic metabolic reprogramming as a central mechanism of immune escape and highlights new therapeutic opportunities to boost immunotherapy efficacy. This study utilized single-cell RNA sequencing (scRNA-seq) to investigate how elevated systemic phenylalanine (Phe), induced by dietary aspartame (APM), affects tumor-infiltrating immune cell populations during anti–PD-1 immunotherapy. Tumor-infiltrating immune cells were isolated from B16F10 melanoma tumors in C57BL/6 mice assigned to two dietary conditions: Chow diet + anti–PD-1 antibody and Aspartame diet + anti–PD-1 antibody. Mice were treated with anti–PD-1 antibodies to simulate immune checkpoint blockade, and tumors were harvested for analysis. Single-cell suspensions were generated from tumor tissues, and CD45⁺ immune cells were enriched by FACS. This experimental design enabled high-resolution analysis of how dietary Phe elevation reshapes the tumor immune microenvironment, with a particular focus on tumor-associated neutrophils (TANs) and their immunosuppressive programs under anti–PD-1 therapy.