Fig. 4: MEKi reduces GATA6 expression that is restored by HDAC inhibitors with augmented MHCI expression.

a Patient-derived xenografts (PDXs) from 15 PDAC patients were transplanted subcutaneously into athymic nude mice, and the mice of third generation were used for MEKi treatment. When tumor volume reached 200 mm³, the mice were treated with vehicle control (Ctrl) or MEKi. Tumors were harvested after 4 weeks of treatment. Created with BioRender.com. b IHC staining of GATA6 and MHCI of the xenograft tumors was performed, and the proportion of marker-positive cells out of total cells was quantified by Definiens software. Xenografts were dichotomized into GATA6^high (GATA6^hi, n = 7 independent mice) and GATA_low (GATA_lo, n = 8 independent mice) expression groups using median GATA6⁺ cells (% total) as cutoff. MHCI⁺ cells (% total) were quantified by Definiens and compared between vehicle control and corresponding MEKi-treated mice in GATA6^high and GATA_low expression groups, respectively. Statistical significance was calculated by two-tailed Wilcoxon matched-pairs signed rank test. c IHC staining showing reduction of GATA6 expression in MEKi-treated tumors in GATA6^high expression group. Signal intensities were quantified by Definiens and compared between vehicle control and MEKi-treated mice groups (n = 7 independent mice for each group). Statistical significance was calculated by two-tailed Wilcoxon matched-pairs signed rank test. d Flow cytometry analysis of GATA6 expression in 110299 cells treated with MEKi, class I HDAC inhibitors (mocetinostat, quisinostat, domatinostat), class II HDAC inhibitors (tasquinimod, LMK235, ricolinostat), or their combinations (n = 4 independent experiments for each cell line). Data are presented as mean values ± SD. Individual dots represent independent biological replicates. Statistical significance was calculated by One-way ANOVA, Kruskal–Wallis test. e Western blot analysis of GATA6 protein levels in four GATA6^high (orange: 110299, 511950, 2838c3, 60400) and four GATA6_low (blue: 60531, 511892, 60590, 6694c2) murine PDAC cell lines following treatment with MEKi and/or domatinostat; β-actin served as a loading control (n = 1 independent experiment). f Surface MHCI (H-2Db) expression in the four GATA6^high (orange) and four GATA6_low (blue) murine PDAC cell lines treated with MEKi and/or domatinostat, assessed by flow cytometry (n = 4 independent experiments for cell line 511950, 60400, 511892, 60590, and 5 independent experiments for cell line 110299, 2838c3, 60531, 6694C2). Data are presented as box plots: the center line indicates the median, the bounds of the box indicate the 25th and 75th percentiles, and whiskers extend to minima and maxima. One-way ANOVA and Kruskal–Wallis test were used. Ctrl: vehicle control; MEKi: trametinib; Moce: mocetinostat; Quis: quisinostat; Doma: domatinostat; Tas: tasquinimod; LMK: LMK235; Rico: ricolinostat. MFI mean fluorescence intensity. Scale bar: μm.