〔Abstract〕 Objective To investigate the mechanism by which human hypoxia up-regulation 1(HYOU1) regulates pancreatic cancer development. Methods Bioinformatic and immunohistochemical staining analyses of HYOU1 expression level in pancreatic tumor tissues, normal and paracancerous tissues and its correlation with patients′ survival. Quantitative real-time PCR(qPCR) and Western blot were used to clarify the expression level of HYOU1 in a number of human pancreatic ductal adenocarcinoma cell lines and a normal human pancreatic ductal cell line. Two cell lines with the highest expression levels of HYOU1, BXPC-3 and Panc-1, were selected to knock out HYOU1 by CRISPR-Cas9,and then cell survival, proliferation and migration of these cells were examined by cell counting kit-8(CCK-8),colony formation assay and wound healing experiment separately,as well as apoptosis was detected by flow cytometry. Subsequently,the protein levels of the PI3K/Akt signaling including PI3K,p-PI3K,Akt,and pAkt were detected by Western blot in parental and HYOU1-ablated BXPC-3 and Panc-1 cells. Cell proliferation was also examined in HYOU1-ablated cells after treatment of recilisib,an activator of the PI3K/Akt pathway. Results The expression of HYOU1 in pancreatic tumor tissues was significantly higher than that in normal tissues,and the patients with high expression of HYOU1 had a much shorter survival compared to the patients with low HYOU1(P<0.01). Immunohistochemical staining of pancreatic cancer specimens showed that the expression of HYOU1 was higher in tumor tissues than in paracancerous tissues(P<0.01). The mRNA and protein levels of HYOU1 were higher in all pancreatic cancer cell lines compared to the human normal pancreatic ductal cell(P<0.001,P<0.01). HYOU1 ablation inhibited BXPC-3 and Panc-1 cells survival,proliferation and migration,and promoted early cell apoptosis. In addition,loss of HYOU1 decreased PI3K/Akt signaling activity,whereas the PI3K/Akt activator Recilisib reversed the effects of HYOU1 ablation on cell survival and proliferation. Conclusion HYOU1promotes pancreatic cancer progression by activating the PI3K/Akt signaling pathway.