Dznep

THP-1 cells (Cell Bank, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences) and monocytes were maintained in RPMI 1640 medium supplemented with 10% (v/v) fetal bovine serum (Gibco), 2 mM L-Glutamine, 1 mM sodium pyruvate, and 10 mM HEPES at 37°C in a humidified atmosphere containing 5% CO2.
PBMCs were isolated using Ficoll-Paque PLUS (Cytiva). Monocytes (>98% purity) were sorted from the PBMCs with a BD FACSAriaTM III cell sorter (BD Biosciences). Monocytes were identified by anti-CD14 antibody (BD Biosciences).
For transient transfection, THP-1 cells (2 × 10⁵/well) were seeded in 24-well plate 24 h before transfection. SiRNAs (combination of four different EZH2 targeting siRNAs, 200 nM) were transfected with LipofectamineTM RNAiMAX (Invitrogen). Twenty-four hours later, the cells were stimulated with 1,000 U/ml of universal IFN-I, then collected for further analysis as shown in the figures. All siRNA sequences (GenePharma) are in Table S2.
For EZH2 inhibitor treatment, THP-1 cells were pretreated with 5 uM GSK126, 5 uM Dznep (Selleck Chemicals), or DMSO for 30 min, then 1,000 U/ml of universal IFN-I was added. Then, the cells were collected for further analysis as shown in the figures.

We used DZNep (Selleck, S7120) , GSK126 (Selleck, S7061) and GSK-J4 (Selleck, S7070) as inhibitors of EZH2 and JMJD3 respectively, which are with DMSO (Dimethyl Sulfoxide) solvent, and used DMSO as negative control. Rat primary HSCs were treated with DZNep (1 μM), GSK126 (5 or 10 μM) or GSK-J4 (5 μM). DZNep and GSK126 treatment were also conducted in JS1 and LX-2 cell lines. Experimentally diseased mice (as below) were administered with DZNep (1 mg/kg) through intraperitoneal injection twice a week. The H3K27me2/3 levels in HSCs and in liver tissue were analyzed by Western blot to ascertain the inhibitory effect.

Pancreatic cancer cells lines CFPAC-1, SW1990, AsPC-1 were all obtained from the Chinese Academy of Sciences Cell Bank (Shanghai, China). AsPC-1 and CFPAC-1cells were cultured in RPMI 1640 (Gibco, USA) supplemented with 10% fetal bovine serum (FBS), grown in 5% CO₂ saturated humidity at 37°C and sub-cultured by harvesting with trypsin-EDTA. SW1990 cells were cultured in L-15 medium (Gibco) supplemented with 10% FBS, and grown in room temperature air.
Paraffin embedded tissue samples were collected from the surgery department, Shanghai General Hospital. Written informed consent was obtained from all subjects, and the study was approved and supervised by the Ethics Committee of the Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine.
Selective EZH2 inhibitors, EPZ-6438 and DZNeP (Selleck), were used to examine differential expression of proteins after EZH2 knockdown.

The PCa cell line LNCaP-FGC was obtained from the American Tissue Culture Collection (Manassas, VA) and the C4-2 cells were obtained from the MD Anderson Cancer Center, University of Texas. Cells were maintained in RPMI 1640 medium supplemented with 10% FBS, 1% penicillin/streptomycin, and 1% glutamine. DHT was obtained from Amersham (Braunschweig, Germany), DZNep was from SELLECK (S71201) and MDV3100 was from Haoyuan Chemexpress (Shanghai, China).

The following drugs were employed in tumor organoid drug studies: cisplatin (Sigma), carboplatin (Sigma), pemetrexed (Sigma), and drug 3-deazaneplanocin A (DZNep, SelleckChem, Houston, TX). cisplatin/pemetrexed and carboplatin/pemetrexed cocktails were reconstituted in DMEM cell culture media with matched platinum agent and pemetrexed concentrations at 0.1 μM and 10 μM or 0.2 μM and 20 μM for administration to organoids. DZNep was reconstituted in DMEM cell culture media at 0.1 μM, 1 μM, and 10 μM for administration to organoids.