e16510 Background: o explore the effects of removing heat and phlegm prescription (RHPP) on the proliferation and autoantigens expression of esophageal carcinoma(EC) cell, so as to provide basis for the molecular pathogenesis and clinical medication of EC. Methods: RHPP was developed by us for treating EC, EC autoantigens CK13, CK16, CaD, ACTG2 were identified in our previous studies. The effects of RHPP and and its ethanol extraction on the proliferation, cell cycle and autoantigen protein expression of Eca109 cell, EC9706 cell and TE-1 cell were investigated by MTT assay, flow cytometry and western blot analysis. Results: RHPP and its removing heat (RH) and removing phlegm (RP) separated prescriptions all have inhibitory effects on the proliferation of EC9706, EC109 and TE-1 cells in dose-dependent and time-dependent manner, changed morphology of four esophageal carcinoma cells, which appeared as round with rough edges, karyopyknosis, and karyorrhexis. Ic50 values of RHPP for Ec9706, Eca109 and TE1 cell were 33.31 ug·ml−1, 20.70 ug·ml−1, 21.93 ug·ml−1 respectively, while Ic50 values of RHPP’s ethanol extraction for Ec9706, Eca109, TE1 were 0.653 ug·ml−1, 0.082 ug·ml−1, 0.172 ug·ml−1 respectively. RHPP and RP induced G2/M phase arrest in EC109 and TE-1 cells, while RH induced G0/G1 phase arrest in EC109 and TE-1 cells; RHPP and RP induced G0/G1 phase arrest in EC9706 cells, while RH induced S phase arrest in EC9706 cells. RHPP and its two separated prescription could downregulate CK16, CaD, ACTG2 expression and upregulate CK13 expression. Conclusions: Autoantigens CK13, CK16, CaD and ACTG2 were expressed in EC cell, RHPP could regulate these four autoantigens expression. This study provides new basis for the EC mlecular mechanism and development of anti-esophageal carcinoma drugs in traditional Chinese medicine.