Abstract 586: Targeting Pim kinases by SGI-1776 in mantle cell lymphoma (2011)

Abstract Proviral integration site for Moloney murine leukemia virus (PIM) kinases are oncoproteins that promote tumor progression. Pim-1, -2 and -3 are highly conserved Ser/Thr kinases that are important for normal B-cell development and are over-expressed in B-cell malignancies such as mantle cell lymphoma (MCL). Pim kinases phosphorylate a number of substrates; including: cMyc and Histone H3, which drive transcription, eukaryotic elongation factor 4E-BP-1 that regulates translation and Bad that activates cell survival. Furthermore, cell cycle protein activation by Pim kinase is involved with proliferation. An imidazo-[1, 2-b]-pyridazine compound, SGI-1776 (SuperGen) has high inhibitory specificity to Pim kinases at nM levels. Based on this knowledge we hypothesized that Pim kinase are druggable targets in MCL and inhibition of Pim kinase-mediated phosphorylation of downstream substrates will disrupt transcriptional and translational processes and trigger apoptosis. To test this hypothesis, Pim kinase expression profile and actions of SGI were studied in MCL cell lines, Jeko and Mino. All three Pim kinase were expressed in both cell lines. SGI was used in concentration (0.1 to 10 µM) and time dependent (up to 24 hrs) fashion. Treatment with SGI resulted in decreased Pim-1 and -3 protein levels in both cell lines; however Pim-2 protein was declined only in Jeko. Pim kinase pathways were investigated to identify mechanism of action of SGI. In both Jeko and Mino, total and phospho cMyc (Ser62) protein expression decreased at 3 and 10 µM, respectively. Interestingly, Histone H3 (Ser10) phosphorylation decreased only in Jeko. Consistent with cMyc data, there was 50% decrease in RNA synthesis (uridine incorporation) at 5 µM SGI in Jeko and 10 µM in Mino. When evaluated in Mino cells, 4E-BP-1 was not affected up to 3 µM of SGI. SGI inhibition on transcription/translation in principle would affect short-lived proteins such as Mcl-1 that are survival factors. Significant decreases of Mcl-1 mRNA (80% and 50%) and protein (50% and 40%) levels were observed in Jeko and Mino at 3 and 10 µM, respectively. In contrast, other anti-apoptotic proteins Bcl-2, Bcl-XL and XIAP were not affected by SGI treatment. In concordance with the survival role of Mcl-1, apoptosis (Annexin V+) was induced at 3, 5 and 10 µM SGI (15%, 30% and 80%) in both cell lines. Similarly, PARP cleavage was detected. Total and phospho Bad (Ser112) protein was decreased at 3 µM in Jeko, suggesting a role of this Pim substrate in apoptosis. Consistent with the observations in cell lines, there was induction of cell death in primary MCL cells from patients (n = 5). Preliminary data suggest inhibition of RNA synthesis in these malignant lymphocytes. Taken together, Pim kinases are potential therapeutic targets in MCL and SGI-1776 is effective to inhibit global RNA synthesis, as well as inducing apoptosis. MCl-1 appears important factor in Pim kinase pathway and warrant more detailed investigation. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 586. doi:10.1158/1538-7445.AM2011-586