HIFA-AS1, an antisense transcript of HIF1α gene, is a 652-bp long non-coding RNA (lncRNA) which globally expressed in multiple tissues of animals. Recent evidence indicated that the HIFA-AS1 was involved in tumorigenesis of several types of cancer, but there were no reports on pancreatic cancer (PC). In order to investigate whether the HIFA-AS1 could mediate the PC or not, it was overexpressed in a PC cell line (PANC-1), and a series of experiments including cell viability detection, flow cytometry, and transwell migration were performed. Functionally, the results indicated that overexpression (OE) of HIFA-AS1 could inhibit proliferation and shift, and promote apoptosis of PC cells. Moreover, to explore underlying molecular mechanism of HIFA-AS1 in PC cells, the iTRAQ (isobaric tags for relative and absolute quantification) quantitative proteomics analysis was implemented. Mechanistically, it was found that OE of HIFA-AS1 globally affected the expression levels of multiple protein associated with metabolism of cancer, suggesting that HIFA-AS1 might regulate the tumorigenesis of PC by up- and down-regulating the gene expression.