Esophageal squamous cell carcinoma (ESCC) is the most common subtype of esophageal cancer (ESCA). ESCC is one of the malignancies with high incidence and mortality rates. Studies have found that laminin subunit alpha 3 (LAMA3) functions as an oncogene in a variety of cancers. SPI1 is highly expressed in ESCC, but whether LAMA3 and SPI1 regulate the development of ESCC is still unclear. In this study, bioinformatics analysis tools were used to predict the expression of LAMA3 and SPI1 in ESCA. Subsequently, the levels of mRNA and protein were respectively detected by RT-qPCR and WB. Then, the cell biological behaviors were measured by CCK-8, colony formation, EdU, and tube formation assays. To investigate the in vivo effects of LAMA3 knockdown on ESCC, a xenograft tumor model was established, followed by IHC analysis. Additionally, glucose consumption, lactate production, ROS, and Fe2+ levels were determined by the corresponding kits. Besides, the interaction of LAMA3 and SPI1 was examined by ChIP and dual luciferase reporter assays. LAMA3 was highly expressed in ESCC and silencing it could curb the viability and proliferation of ESCC cells, tumor growth in vivo, tube formation of HUVECs, and induce oxidative stress and ferroptosis of ESCC cells. SPI1 was highly expressed in ESCC and could bind to the promoter of LAMA3 to jointly regulate the progression of ESCC. This study elucidated that SPI1 aggravated ESCC by binding to the promoter of LAMA3, thereby stimulating the growth and proliferation of ESCC cells and suppressing oxidative stress and ferroptosis.