Targeting keratin 6 A overcomes gemcitabine resistance by restoring equilibrative nucleoside transporter 1 and TAM-mediated metabolic compensation in pancreatic cancer.

[BACKGROUND & AIMS] Gemcitabine remains a cornerstone chemotherapy for pancreatic cancer, yet intrinsic and acquired resistance severely limits its efficacy. This study aimed to identify key drivers of gemcitabine resistance and develop a targeted strategy to restore chemosensitivity.

[METHODS] Public datasets of gemcitabine-resistant pancreatic cancer cells were integrated to identify resistance-associated genes, followed by RT-qPCR validation in resistant cell models. Clinical relevance was evaluated using pancreatic cancer cohorts and tissue microarrays with survival and multivariable analyses. Spatial and single-cell transcriptomics revealed keratin 6 A (KRT6A) as a key gene associated with gemcitabine resistance. A cyclic Arg-Gly-Asp (cRGD)-modified lipid nanocarrier with silencing KRT6A (c-Lip@siKRT6A) was engineered for tumor targeting, and therapeutic efficacy was tested in xenograft models with gemcitabine, including rescue experiments using M2-polarized tumor-associated macrophages (TAM-M2) cells and cytidine.

[RESULTS] KRT6A was upregulated in pancreatic cancer compared with adjacent normal tissues and was associated with poor prognosis in public cohorts and tissue microarrays. In a gemcitabine-treated cohort (n = 90), high KRT6A protein expression was correlated with reduced disease control rate and objective response rate, supporting a clinical link between KRT6A and gemcitabine resistance. Functionally, overexpression of KRT6A promoted malignant phenotypes and chemoresistance, whereas KRT6A silencing enhanced gemcitabine sensitivity. Multi-omics analyses indicated that KRT6A-positive tumor regions exhibited decreased equilibrative nucleoside transporter 1 (ENT1) and increased macrophage migration inhibitory factor (MIF), accompanied by enrichment of TAM-M2 subpopulations and activation of the MIF-CD74/CD44 axis. Metabolic analysis suggested a pyrimidine-metabolism biased niche with cytidine dynamics linked to the KRT6A-associated microenvironment. c-Lip@siKRT6A achieved efficient tumor targeting and markedly suppressed tumor growth when it was used in combination with gemcitabine. Mechanistically, KRT6A silencing reduced MIF signaling, restored ENT1, and decreased M2 polarization. TAM-M2 or cytidine rescue partially reversed the antitumor benefit, supporting a TAM-nucleoside-ENT1 competitive axis contributing to gemcitabine resistance.

[CONCLUSIONS] KRT6A drives gemcitabine resistance by suppressing ENT1-mediated drug uptake and promoting a cytidine enriched microenvironment via the MIF-CD74/CD44 axis and TAM-M2 polarization. Tumor-targeted delivery of siKRT6A using c-Lip@siKRT6A sensitizes pancreatic cancer to gemcitabine, which has translational nanotherapeutic implications.