Lyn and Fyn inhibition as a potential novel treatment for glioblastoma multiforme

Abstract

Background: Glioblastoma multiforme (GBM) is the most malignant and most aggressive of the adult brain tumors. The current therapies including surgical resection, radiation therapy and chemotherapy remain palliative with median survival of 14 months. Two of the Src family kinases, Lyn and Fyn, have been reported to be involved in the pathogenesis and aggressiveness of GBM in clinical and laboratory research studies. Specific Aims: We evaluated the effect of Lyn and Fyn kinase inhibition on GBM cell motility and survival by utilizing a clinically relevant Lyn and Fyn kinase inhibitor, bafetinib or formerly INNO-406 (CytRx Corporation, Los Angeles, CA). Bafetinib, a dual Bcr-Abl and Lyn tyrosine kinase inhibitor has been shown to inhibit 4 of the 79 tyrosine kinases, namely Abl, Abl-related gene (Arg), and two of the Src family kinases Lyn and Fyn but not Src. Methodology: We used Lyn- expressing T98G and Fyn-expressing U87MG human GBM cell lines. The effect of bafetinib on the motile activity was evaluated by the wound healing, single cell trajectory, and the trans-well assays. The cell survival and mitochondrial membrane potential were evaluated by Guava Easycyte *HT flowcytometer (Millipore) in cells treated with bafetinib for 24 h. Further, we examined the effect of bafetinib on cell survival in combination with temozolomide (TMZ), a widely used chemotherapy agent for the management of GBM, in U87MG cells using microplate MTT assay. Results: At 16 h post-wounding, bafetinib at 5 µM concentration inhibited the motility of T98G and U87MG cells by 33% (P < 0.01) and 24% (P < 0.01), respectively. This significant inhibition in the wound healing was accompanied by the inhibition of total length of cell trajectory, total length of the final cell displacement and average speed of cell locomotion. Bafetinib induced 62% mitochondrial membrane depolarization in U87MG cells and 20% depolarization in T98G cells. Higher depolarization levels in U87MG cells correlated with higher apoptosis (49.4%). Treatment of U87MG cells for 4 days with a combination of bafetinib and TMZ reduced the IC50 values as much as 60% and 80%, respectively, compared to the values obtained with each of the drugs, suggesting that bafetinib has the potential to improve the therapeutic effect of TMZ in GBM cells. Conclusion: Our preliminary findings suggest that targeting Lyn and Fyn kinase pathways with bafetinib may have a therapeutic potential for GBM. Our in vivo studies in mouse models with bafetinib will further establish the feasibility of targeting Lyn and Fyn kinases as possible therapeutic targets in GBM.