Figure 7.

Targeting TET2-driven epigenetic rewiring to overcome resistance. A, Relative cell proliferation of LNCaP/AR cells transduced with annotated guide RNAs and treated with vehicle (DMSO), enzalutamide (10 µmol/L for 7 days), or Bobcat339 (10 µmol/L for 7 days). B, Dose–response curve of ZNF397-KO and wild-type cells treated with TET2 inhibitor Bobcat339. P values were calculated by non-linear regression with extra sun-of-squares F test. C, Tumor growth curve of xenografted ZNF397-KO LNCaP/AR cells in castrated mice treated with enzalutamide. The mice were randomly separated into two groups when their tumors reached 500 mm³ and treated with vehicle or Bobcat339. D, Tumor growth curve of xenografted ZNF397-KO CWR22Pc cells in intact mice. The mice were randomly separated into two groups and castrated when their tumors reached 500 mm³ and treated with vehicle + enzalutamide or Bobcat339 + enzalutamide. E, Waterfall plot displaying changes in tumor size of xenografted ZNF397-KO LNCaP/AR cells after 5 week of combined treatments. F, Waterfall plot displaying changes in tumor size of xenografted ZNF397-KO LNCaP/AR cells after 20 days of combined treatments. For C–F, Enz denotes enzalutamide treatment at 10 mg/kg. Bobcat 339 denotes Bobcat339 treatment at 10 mg/kg. n = number of independent xenografted tumors in each group. G, Bright field pictures represent the 3D-cultured patient derived organoid models, treated with vehicle (Veh), 5 µmol/L enzalutamide (Enz), 10 µmol/L Bobcat339 or enzalutamide + Bobcat339 for 7 days. H, Bar plots represent the size of the 3D-cultured patient derived organoids, treated with vehicle (Veh), 5 µmol/L enzalutamide (Enz), 10 µmol/L Bobcat339 or enzalutamide + Bobcat339 for 7 days. I, Relative gene expression levels of canonical lineage specific marker genes in ZNF397-KO and wild-type LNCaP/AR cells treated with vehicle (DMSO) or Bobcat339 (10 µmol/L for 5 days), measured by qPCR assay. Data are normalized to vehicle-treated cells. J, A schematic figure illustrates the bifurcated role of ZNF397 as a critical coactivator of AR and a suppressor of TET2-dependent lineage plasticity. ZNF397 deficiency in prostate cancer facilitates the transition of prostate cancer cells from an AR-driven luminal lineage to a TET2-driven, multilineage, and lineage-plastic state, which no longer responded to AR-targeted therapy. A yet-to-be identified TET2 cofactor is depicted as a yellow protein. For all panels unless otherwise noted, n = 3 independently treated cell cultures and mean ± SEM are presented. P values were calculated using two-way ANOVA with Bonferroni multiple-comparison test. Schematic figure was created with BioRender.com. See also Supplementary Fig. S10.