Acidic Tumor Microenvironment Promotes 5-Fluorouracil Resistance Via Lactate-Driven HIF-1α/MCT4 Activation and...

Tumor acidosis is associated with poor therapeutic response. Yet, it remains unclear whether 5-fluorouracil (5-FU) resistance in gastric cancer is driven by low extracellular pH per se or by lactate as a signaling metabolite.

The human gastric cancer cell line AGS was exposed to extracellular acidosis (pH 6.6; short-term or chronic) or exogenous lactate at neutral pH. 5-FU sensitivity, apoptosis, and lactate production were assessed. Extracellular pH was buffered with 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES); lactate metabolism/ export was disrupted by LDHA inhibition (FX11) or siRNA-mediated MCT4 knockdown. Global histone methylation was analyzed by Western blot, and qPCR quantified H3K4me3 and H3K27me3 occupancy at SLC16A3 (MCT4) and LDHA promoters.

Acidosis reduced 5-FU sensitivity, increased lactate output, suppressed apoptosis, and activated hypoxia- inducible factor 1 alpha (HIF-1α)/monocarboxylate transporter 4 (MCT4) signaling. Buffering pH or inhibiting lactate synthesis/export restored 5-FU sensitivity, indicating a reversible, metabolism-dependent phenotype. Under neutral pH, exogenous lactate alone induced reduced 5-FU sensitivity and activated HIF-1α signaling, supporting a pH-independent role for lactate. Acid exposure increased global H3K27me3 and H3K9me3, which normalized after buffering, whereas SLC16A3 and LDHA promoters showed H3K4me3 enrichment with reduced H3K27me3.

These findings suggest that acidic stress is transduced through lactate signaling to selectively activate chromatin at lactate-metabolism genes, stabilizing an adaptive survival program under chemotherapy.

Lactate signaling, rather than low pH alone, underlies a reversible 5-FU-resistant state in gastric cancer cells and may be mitigated by targeting lactate metabolism and/or extracellular pH control.