LIM Domain Kinases as Potential Therapeutic Targets for Neurofibromatosis Type 2

Neurofibromatosis Type 2 (NF2) is caused by mutations in the neurofibromatosis 2 (NF2) gene that encodes a tumor suppressor protein called merlin. NF2 is characterized by formation of multiple schwannomas, meningiomas and ependymomas. Merlin loss of function is associated with increased activity of Rac and p21-activated kinases (PAK) and deregulation of cytoskeletal organization. LIM domain kinases (LIMK1 and 2) are substrate for Cdc42/Rac-PAK, and modulate actin dynamics by phosphorylating cofilin at serine-3. This modification inactivates cofilin’s actin severing and depolymerizing activity. LIMKs also translocate into the nucleus and regulate cell cycle progression. Significantly, LIMKs are overexpressed in several tumor types, including skin, breast, lung, liver and prostate. Here we report that mouse Schwann cells (MSCs) in which merlin function is lost as a result of Nf2 exon2 deletion (Nf2(ΔEx2)) exhibited increased levels of LIMK1, LIMK2, and active phospho-Thr508/505-LIMK1/2, as well as phospho-Ser3-cofilin, compared to wild-type normal MSCs. Similarly, levels of LIMK1 and 2 total protein and active phosphorylated forms were elevated in human vestibular schwannomas compared to normal human Schwann cells (SCs). Reintroduction of wild-type NF2 into Nf2(ΔEx2) MSC reduced LIMK1 and LIMK2 levels. We show that pharmacological inhibition of LIMK with BMS-5, decreased the viability of Nf2(ΔEx2) MSCs in a dose-dependent manner, but did not affect viability of control MSCs. Similarly, LIMK knockdown decreased viability of Nf2(ΔEx2) MSCs. The decreased viability of Nf2(ΔEx2) MSCs was not due to caspase-dependent or -independent apoptosis, but rather, to inhibition of cell cycle progression as evidenced by accumulation of cells in G(2)/M phase. Inhibition of LIMKs arrest cells in early mitosis by decreasing Aurora A activation. Our results suggest that LIMKs are potential drug targets for NF2 and tumors associated with merlin deficiency.