Metformin, the most commonly used drug for type 2 diabetes, has recently been shown to have beneficial effects in patients with cancer. Despite growing evidence that metformin can inhibit tumor cell proliferation, invasion, and metastasis, studies on drug resistance and its side effects are lacking. Here, we aimed to establish metformin-resistant A549 human lung cancer cells (A549-R) to determine the side effects of metformin resistance. Toward this, we established A549-R by way of prolonged treatment with metformin and examined the changes in gene expression, cell migration, cell cycle, and mitochondrial fragmentation. Metformin resistance is associated with increased G1-phase cell cycle arrest and impaired mitochondrial fragmentation in A549 cells. We demonstrated that metformin resistance highly increased the expression of proinflammatory and invasive genes, includingBMP5,CXCL3,VCAM1, andPOSTN, using RNA-seq analysis. A549-R exhibited increased cell migration and focal adhesion formation, suggesting that metformin resistance may potentially lead to metastasis during anti-cancer therapy with metformin. Taken together, our findings indicate that metformin resistance may lead to invasion in lung cancer cells.