Het 1a

Human ESCC cell lines (ECa-109, EC9706, KYSE30 and KYSE450) and normal cell line (Het-1A) were utilized for this study and all of them were deposited with 5% CO₂ at 37 °C. Het-1A cells were purchased from ATCC (Manassas, VA, USA). ECa-109 cells were purchased from Cell Bank of Chinese Academy of Sciences (Shanghai, China). EC9706 cells were purchased from Fuxiang Biotechnology Co., Ltd (Shanghai, China). KYSE30 cells were purchased from Procell Life Science & Technology Co., Ltd. (Wuhan, China). KYSE450 cells were purchased from Cobioer Biosciences Co., Ltd (Nanjing, China). Het-1A cells were cultured in BEGM medium (Gibco, Grant Island, NY, USA). ESCC cell lines were all cultivated in RPMI-1640 medium with 10% FBS and 1% p/s (Gibco). All cell lines have been certified using STR profiling.

Human EC cell lines (KYSE30 and KYSE150) and a healthy esophageal cell line (Het-1A) were obtained from the American Type Culture Collection. Het-1A, KYSE30 and KYSE150 cells were cultured in DMEM (Gibco; Thermo Fisher Scientific, Inc.), supplemented with 10% FBS (Beyotime Institute of Biotehcnology) and 1% streptomycin-penicillin. All cell lines were maintained at 37˚C in a 5% CO₂ atmosphere incubator.

Three esophageal cell lines (KYSE150, EC9706 and TE-9) and human normal esophageal epithelial cell line Het-1A were purchased from the Cell Bank of Shanghai Institute of Cell Biology (Chinese Academy of Medical Sciences, Shanghai, China). KYSE150, EC9706 and TE-9 cells were cultured in RPMI-1640 medium (Gibco, Carlsbad, CA, USA) supplemented with 10% fetal bovine serum (FBS; Gibco) and a 1% penicillin (Invitrogen, Shanghai, China). Het-1A cells were cultured in Dulbeccos modified Eagles medium (DMEM; Gibco) supplemented with 10% FBS and 1% penicillin. All the cell lines were grown at 37°C in an incubator.

Human ESCC cell lines EC109, KYSE140, KYSE30, KYSE150, KYSE450, KYSE510, and CaES17 were obtained from the Cell Bank of the Chinese Academy of Sciences (Shanghai, China). The human normal esophageal epithelial cell line Het‐1A was obtained from the American Tissue Culture Collection (ATCC). The human embryonic kidney cell line 293T and the luciferase‐labeled KYSE30 cell line (KYSE30‐fLuc) were kindly provided by colleagues in other departments. All cancer cells were cultured in RPMI 1640 (Gibco, Carlsbad, CA, USA) supplemented with 10% FBS (BI, Carlsbad, CA, USA). Het‐1A and 293T cells were cultured in DMEM (Gibco, Carlsbad, CA, USA) containing 10% FBS. Cells were kept in an humidified incubator at 37°C and 5% CO₂. Hypoxia (<1% O₂) experiments were performed in a hypoxia incubator (Billups‐Rothenberg Inc, USA). The CXCR4 antagonist MSX‐122 was purchased from MedChemExpress (Shanghai, China).
ESCC cell lines were searched on Cellosaurus (https://web.expasy.org/cellosaurus/).¹⁹ Keyword C4024 (NCI Thesaurus Code for ESCC) was used for searching, and the data of ESCC cell lines used in our study are shown in Table S1. The Cancer Cell Line Encyclopedia (CCLE) is a database that contains multiomic data of over 1000 cancer cell lines, 28 ESCC cell lines included. Before analysis in this paper, cell lines from CCLE were checked to see if any problematic cell lines existed.

ESCC cell lines Eca-109, TE-1, KYSE-150, and TE-10, as well as the human normal
esophageal epithelial cell line Het-1A, were purchased from the Cell Bank of
Shanghai Institute of Cell Biology (Shanghai, China). All ESCC cells were
cultured in RPMI-1640 medium (Gibco, Carlsbad, CA, USA) containing 10% FBS
(Gibco, Carlsbad, CA, USA), and Het-1A was cultured in DMEM (Gibco, Carlsbad,
CA, USA) supplemented with 10% FBS (Gibco). All cells were incubated at 37°C in
an incubator with 5% CO₂.
The miR-1269 mimic (5′-CUGGACUGAGCCGUGCUACUGG-3′), mimic negative control (NC;
5′-UUCUCCGAACGUGUCACGUTT-3′), miR-1269 inhibitor (5′-CCAGUAGCACGGCUCAGUCCAG-3′),
and inhibitor NC (5′-CAGUACUUUUGUGUAGUACAA-3′) were synthesized from GenePharma
Co., Ltd (Shanghai, China). ESCC cells were seeded in 6-well plates and cultured
at 37°C overnight. Cells were subsequently transfected with miR-1269 mimic,
mimic NC, miR-1269 inhibitor, or inhibitor NC using Lipofectamine 3000
(Invitrogen; Thermo Fisher Scientific, Inc.) according to the manufacturer’s
instructions. After 48 h transfection, cells were collected for subsequent
assays. Untreated cells were used as control.