p21-Associated Kinase 4 (PAK4) Addiction in Multiple Myeloma: A Potential Therapeutic Application with Enhanced Activity in FGFR3 Expressing Myeloma

Dysregulated oncogenic serine/threonine kinases (STKs) play a pathological role in diverse forms of malignancies, including multiple myeloma (MM), and thus represent potential therapeutic targets. Here, we evaluated the biological and functional role of p21-activated kinase 4 (PAK4), and its potential as a new therapeutic target in MM. The PAK4 gene lies within chromosome region 19q13.2 commonly amplified in 30% of MM patients. We confirmed a correlation between copy number amplification and increased expression of PAK4 in two large myeloma patient datasets. High expression of total and phosphorylated PAK4 was also observed in the premalignant MGUS stage, suggesting that its overexpression may be an early event in the pathogenesis of myeloma. Myeloma cells with high PAK4 expression display sensitivity to conditional PAK4 knockdown, implying that an oncogene-addicted state exists in such cells. Moreover, we show that PAK4 promotes myeloma cell proliferation in vitro as well as in vivo in murine models of human myeloma through activation of the MEK/ERK pathway. This is further highlighted by the correlation between the basal levels of ERK activity and sensitivity to a novel small molecule inhibitor of PAK4, KPT-9274, a lead clinical agent. KPT-9274 effectively inhibits MM cell growth and survival via ERK inhibition in a large panel of MM cell lines and primary MM cells, also in the presence of bone marrow microenvironment, with no significant effect on normal PBMCs suggesting a potentially favorable therapeutic index. Inhibition of ERK by KPT-9274 also correlates with decreased DNA binding activity of ERK-dependent transcription factors AP1, ETS, CREB and EGR and decreased expression of ERK target genes such as CCND2, CCR1 and MYC. Finally, using a protein binding array, we have identified FGFR3, a commonly disrupted tyrosine kinase receptor, and Grb2, an adaptor protein involved in RAS activation, as novel PAK4 binding partners in myeloma and report disruption of this binding by KPT-9274. As a results, t(4;14)-positive MM cells expressing FGFR3 show greater sensitivity to PAK4 inhibition compared to MM cell lines without FGFR3 aberrations (e.g. U266 and ANBL6). FGFR3 may therefore serve as a predictive marker of KPT-9274 sensitivity in MM patients, with the potential for broader application to other malignancies associated with dysregulation of FGFR3 such as human bladder and cervical carcinomas.

In conclusions, our study sheds light on the oncogenic role of the serine/threonine kinase PAK4 as survival and anti-apoptotic factor in myeloma and its inhibition with a new allosteric modulator, KPT-9274, as a potential novel therapeutic intervention in MM especially in high risk t(4:14)-positive patients.