Crizotinib, a Novel Inhibitor of ALK, Induces Apoptosis and Down Regulation of pSTAT3 in ALK+ Anaplastic Large Cell Lymphoma
Farid Saei Hamedani, Zhicheng Mo, Hesham M Amin, Melisa A Cervania, Serhan Alkan. Cedars-Sinai Medical Center, Los Angeles, CA; MD Anderson Cancer Center, Houston, TX
Background: Approximately, 70% of Anaplastic Large Cell Lymphomas (ALCL) harbor the t(2;5) translocation that leads to the nucleophosmin-anaplastic lymphoma kinase (NPM-ALK). NPM-ALK binds and activates signal transducer and activator of transcription 3 (STAT3) in ALCL cells. Persistent activation of STAT3 appears to play a critical role in the pathogenesis of ALCLs. A novel inhibitor of ALK, Crizotinib, has been recently approved for the treatment of ALK+ non-small cell lung cancer. In the current study, we evaluated the effects of Crizotinib on ALCL cell lines with t(2;5) translocation to provide “proof of principle” data for future clinical therapeutic investigation of this agent.
Design: ALCL tissue specimens (19) were analyzed by immunohistochemistry (IHC) and a rabbit monoclonal antibody against pSTAT3 (Tyr705). The results were categorized as negative and positive (>20% cells expressing moderate to strong pSTAT3). To study, the effects of Crizotinib, ALK+ ALCL cell lines (SU-DHL-1 and Karpas299) were treated with Crizotinib (0.1 µM, 1 µM and 10 uM for up to 72 h). Cell viability, apoptosis, and cell cycle progression were assessed by microscopy, trypan-blue dye staining and quantifying sub-G0/G1 population by flow cytometry. Utilizing a pSTAT3-(Tyr705)-PE conjugated antibody, pSTAT-3 status was determined by flow cytometry.
Results: All of the ALK+ ALCLs showed expression of pSTAT3 while 7/12 of ALK- ALCLs expressed pSTAT3. Our results are in line with previous studies that showed that the phosphorylation/activation of STAT3 is highly associated with ALK expression in ALCL. In order to investigate pSTAT3 status following Crizotinib treatment, ALK+ ALCL cells were briefly treated with Crizotinib at 0.1 uM and 1 uM for 2 h. Analysis of pSTAT3 expression showed decreased pSTAT3 levels following Crizotinib treatment. In addition, apoptosis was markedly increased in Crizotinib treated cells compared with control untreated cells (p<0.01).
Conclusions: Selective inhibition of ALK has the potential to become an effective therapeutic strategy to treat ALK+ALCL patients. Furthermore, our data not only reveal that pSTA3 is present in the vast majority of ALK+ ALCL cases but also support the notion that pSTAT3 is a critical protein that maintains the survival of ALK+ ALCL cells. In light of these observations, we conclude that Crizotinib is a promising small molecule ALK inhibitor that could be successfully utilized for the treatment of this lymphoma.
Tuesday, March 5, 2013 1:00 PM
Poster Session IV # 276, Tuesday Afternoon