Malignant pleural mesothelioma (MPM) is a disease with few therapeutic options. Protein kinase C beta (PKCβ) is involved in important cellular functions. Enzastaurin (LY317615.HCl) is a novel inhibitor of PKC in clinical development. MPM cell lines (7) and patient tumor tissues (24) were evaluated for expression of PKCβ by immunoblotting and immunohistochemistry, respectively. In-vitro cell growth assays were performed with enzastaurin with or without cisplatin. Cell migration was evaluated with the wound healing assay. Downstream signaling (survival and focal adhesion pathways) was studied by immunoblotting for related molecules in the presence of phorbol ester with or without enzastaurin. Expression for PKCβ1 was seen in all cases, with a mean integrated optical density of 152.5 (standard deviation=95.47, n=24), whereas PKCβ2 expression was less intense, with a mean integrated optical density of 11.45 (standard deviation=16.27, n=21). There was a trend toward lower overall survival among patients expressing above-median PKCβ1 (P=0.064), but not PKCβ2. Robust expression of PKCβ1 and low expression of PKCβ2 were observed in MPM cell lines. Treatment of MPM cell lines with enzastaurin revealed an IC50 of 5 μmol/l, and strong synergism was observed when combined with cisplatin. Wound healing assay revealed that treatment of H2461 cells with enzastaurin reduced migration by 59.2%. Enzastaurin treatment led to disruption of F-actin architecture. Downstream signaling showed reduced phosphorylation of AKT, FAK (focal adhesion kinase), p130Cas, S6 ribosomal protein, and paxillin. PKCβ1 was expressed in the majority of MPM samples. Enzastaurin has preclinical activity against MPM, and exhibited synergism with cisplatin. PKCβ inhibition in MPM might be able to reduce the invasiveness of MPM by affecting cytoskeletal function.