Variable responsiveness to zileuton, a leukotriene antagonist used to treat asthma, may be due in part to genetic variation. While individual SNPs were previously associated with zileuton-related lung function changes, specific quantitative trait loci (QTLs) and biological pathways that may contribute have not been identified. In this study, we investigated the hypothesis that genetic variation within biological pathways is associated with zileuton response. We performed an integrative QTL mapping and pathway enrichment study to investigate data from a GWAS of zileuton response, in addition to mRNA expression profiles and leukotriene production data from lymphoblastoid cell lines (LCLs) (derived from asthmatics) that were treated with zileuton or ethanol (control). We identified 1060 QTLs jointly associated with zileuton-related differential LTB4 production in LCLs and lung function change in patients taking zileuton, of which eight QTLs were also significantly associated with persistent LTB4 production in LCLs following zileuton treatment (i.e., 'poor' responders). Four nominally significant trans-eQTLs were predicted to regulate three candidate genes (SELL, MTF2, and GAL), the expression of which was significantly reduced in LCLs following zileuton treatment. Gene and pathway enrichment analyses of QTL associations identified multiple genes and pathways, predominantly related to phosphatidyl inositol signaling via PI3K. We validated the PI3K pathway activation status in a subset of LCLs demonstrating variable zileuton-related LTB4 production, and show that in contrast to LCLs that responded to zileuton, the PI3K pathway was activated in poor responder LCLs. Collectively, these findings demonstrate a role for the PIK3 pathway and its targets as important determinants of differential responsiveness to zileuton.