Fig. 3: GATA6 is essential for MEKi-induced tumor control and anti-tumor cytotoxicity in vivo.

a Schematic diagram showing the experimental workflow. Orthotopic tumors derived from GATA6_low (60590) or GATA6^high (110299) cell lines were monitored weekly by 3D ultrasound imaging. MEKi treatment was initiated when tumor volume (TV) reached 200–400 mm³, and experiments were terminated when tumor volume (TV) reached approximately 1000 mm³. Created with BioRender.com. b IHC staining of GATA6 in 60590 and 110299 tumors without treatment. The percentage of GATA6⁺ cells out of total cells was quantified by HALO software (n = 5 independent mice per group). Data are presented as mean values ± SD. Individual data points represent independent mice. Statistical significance was calculated by unpaired two-tailed Mann–Whitney test. c Tumor volume (TV) dynamics following MEKi treatment. Left panel: representative 3D ultrasound images of 110299 tumors at post 1 week with or without MEKi (Ctrl: vehicle control) treatment. Right panel: Changes in tumor volume from pre- to post 1 week treatment with or without MEKi in both models (n = 4 independent mice for the 60590 model, and n = 5 independent mice for the 110299 model). Data are presented as mean values ± SD. Individual data points represent independent mice. Statistical significance was calculated by One-way ANOVA and Kruskal–Wallis test. d IHC staining of MHCI, CD8, GzmB, cleaved caspase-3 (Cl casp3), and phosphorylated STAT1 (pSTAT1) in 110299 tumors with or without MEKi treatment. The proportion of marker-positive cells out of total cells quantified by HALO software is shown below (n = 4 independent mice for the 60590 model, and n = 5 independent mice for the 110299 model). Data are presented as mean values ± SD. Individual data points represent independent mice. Statistical significance was calculated by unpaired two-tailed Mann–Whitney test. e Schematic diagram of the in vivo MEKi treatment in targeted GATA6 degradation model. Created with BioRender.com. Orthotopic tumors were established using 110299^AID-GATA6 cells or 110299^WT cells. GATA6 degradation was induced by daily administration of 5-Ph-IAA (auxin) after tumor formation. MEKi treatment was started 3 days after auxin administration. f Tumor growth curves showing the ablation of MEKi-induced tumor inhibition of 110299 orthotopic tumors upon GATA6 degradation (n = 5 independent mice for each group). g IHC staining confirmed GATA6 degradation after auxin treatment in 110299^AID-GATA6 tumors. Right panel: Quantification of GATA6⁺ cells out of total cells by HALO software (n = 5 independent mice for each group). Data are presented as mean values ± SD. Individual data points represent independent mice. Statistical significance was calculated by unpaired two-tailed Mann–Whitney test. h IHC analysis in GATA6-depleted tumors showing abolished MEKi-induced increases in MHCI, CD8, Cl casp3, and pSTAT1. Right panel: Quantification of marker-positive cells out of total cells by HALO software (n = 5 independent mice for each group). Data are presented as mean values ± SD. Individual data points represent independent mice. Statistical significance was calculated by unpaired two-tailed Mann–Whitney test. Scale bar: μm.