Secretory autophagy mediates SLC16A3/MCT4-dependent lactate secretion to drive metastatic progression in triple-negative breast cancer.

Triple-negative breast cancer (TNBC) exhibits hyperactive EGF (epidermal growth factor) signaling that drives metabolic plasticity and metastasis. Here, we identify secretory macroautophagy/autophagy as a key downstream effector linking EGF signaling to metabolic reprogramming that fuels TNBC metastatic progression. In TNBC cells, EGF stimulation redirected autophagosomes toward the plasma membrane through a SEC22B-dependent route, signifying activation of secretory autophagy. Proteomic profiling of purified autophagosomes revealed enrichment of the lactate transporter SLC16A3/MCT4 and its chaperone BSG/CD147 on autophagosomal membranes. Mechanistically, EGF promoted MAP1LC3/LC3-SLC16A3 interaction, facilitating SLC16A3 trafficking to the plasma membrane and enhancing lactate efflux. Genetic or pharmacological blockade of autophagy abrogated SLC16A3 surface localization, reduced extracellular lactate accumulation, and markedly suppressed lung metastasis originating from orthotopic TNBC tumors in mice. Although pharmacological inhibition of SLC16A3 effectively blocks its transporter activity and reduces lactate secretion, targeting autophagy provides a more precise approach to suppress EGF-driven SLC16A3 expression and the consequent rise in lactate secretion. Clinically, multiplex immunofluorescence of patient tumors demonstrated strong co-expression of EGFR, LC3, and SLC16A3, which correlated with poor disease-free survival. Our study reveals a previously unrecognized EGF-secretory autophagy axis that orchestrates metabolic remodeling in TNBC and highlights the therapeutic potential of targeting the secretory autophagy- SLC16A3-lactate pathway to restrain metastasis.: 3-MA: 3-methyladenine; ATG5: autophagy related 5; ATG7: autophagy related 7; APf: autophagosome fraction; CQ: chloroquine; CRISPR-Cas9: clustered regularly interspaced short palindromic repeats-CRISPR-associated protein 9; EGF: epidermal growth factor; EGFR: epidermal growth factor receptor; ER: endoplasmic reticulum; ERBB2/HER2: erb-b2 receptor tyrosine kinase 2; GOBP: gene ontology biological process; imBI: induced metabolic bioluminescence imaging; i.p.: intraperitoneal injection; IVIS: in vivo imaging system; LAMP2: lysosomal associated membrane protein 2; LIR: LC3-interacting region; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MAPK/ERK: mitogen-activated protein kinase; PLA: proximity ligation assay; PNS: postnuclear supernatant; SEC22B: SEC22 homolog B, vesicle trafficking protein; shRNA: short hairpin RNA; SLC16A3/MCT4: solute carrier family 16 member 3; SNARE: soluble N-ethylmaleimide-sensitive-factor attachment protein receptor; TIRF: total internal reflection fluorescence; TME: tumor microenvironment; TNBC: triple-negative breast cancer; ULK1/Atg1: unc-51 like autophagy activating kinase 1.