Real-time qPCR was used for measuring expression levels of genes of interest in a subset of tumor samples or ESCC lines. The probes or kits used in this study are shown in Table S5. All qPCR reactions were performed in triplicate with an Applied Biosystems StepOnePlus. The relative expression of genes of interest was determined by normalization to GAPDH expression via a standard-curve method with ten serial dilutions according to the manufacturer’s instructions. All real-time PCR experiments included a no-template control and were done in triplicate.