<p>The long-chain acyl-CoA synthetase (ACSL) family comprises key enzymes that are integral to the fatty acid metabolic pathway and play crucial roles in governing fatty acid (FA) metabolism and lipid homeostasis. These enzymes are involved in multiple pathophysiological processes, including cellular metabolism, endoplasmic reticulum stress, lipid peroxidation, and ferroptosis (iron-dependent cell death). The ACSL family consists of five isoforms—ACSL1, ACSL3, ACSL4, ACSL5, and ACSL6—each of which has been implicated in the pathogenesis, treatment, and prognosis of diverse conditions, including metabolic disorders, cancers, cardiovascular and cerebrovascular diseases, and other clinical conditions. By catalyzing the conversion of polyunsaturated fatty acids into fatty acyl-CoA, these enzymes mediate a range of biological activities, including anti-inflammatory, antioxidant, and antitumor responses, as well as the regulation of ferroptosis through lipid metabolism. Over the past five years, however, there has been a notable lack of comprehensive reviews that systematically summarize the relevance of ACSL to clinical diseases and their underlying molecular mechanisms. The present review seeks to fill this gap by summarizing recent advances in understanding the roles of the ACSL family across diverse diseases, with a focus on emerging therapeutic strategies that target these enzymes. This work provides critical insights that may inform future preclinical and clinical investigations of the ACSL family.</p>