The FLT3 and Pim kinases inhibitor SGI-1776 preferentially target FLT3-ITD AML cells

To the editor:

The Pim (for provirus integration site for Moloney murine leukemia virus) family of serine/threonine kinases are strongly involved in oncogenic processes and efforts are ongoing to develop specific Pim kinase inhibitors as anti-cancer therapy.¹  Interestingly, recent reports from Gandhi's group emphasized the SGI-1776 compound (Astex Pharmaceuticals) as a potent Pim inhibitor with potential interest in chronic lymphocytic leukemia²  and in acute myeloid leukemia (AML).³  In this latter work, the SGI-1776 compound was showed to induce the apoptosis of both AML cell lines and primary AML samples, and to transiently reduce the size of tumors generated by subcutaneous injection of the MV4-11 human leukemic cell line without evidence for toxicity in mice.³  To explain the anti-leukemic activity of SGI-1776, the authors demonstrated an inhibition of Pim target phosphorylation such as c-Myc S⁶² and BAD S¹¹². However, as SGI-1776 also inhibits the FLT3 kinase in vitro,²  major attention should be given to the FLT3 mutational status of AML cells, that is, the presence or not of FLT3 alleles harbouring internal tandem duplication (ITD). Chen and coworkers considered that SGI-1776 was effective against primary AML cells and AML cell lines regardless of their FLT3 mutational status; although a superior effect of this molecule was found in FLT3-ITD AML cells in some experiments.³  In contrast, we made slightly different observations. Although we also found an inhibition of Pim substrate phosphorylation in SGI-1776 treated AML cells (data not shown), we observed an inhibition of FLT3 autophosphorylation on tyrosine (Tyr) residues in the MOLM-14 FLT3-ITD AML cell line, which was similar to that observed after treatment with the specific FLT3 inhibitor AC-220^4,5  (Figure 1A). Moreover, SGI-1776 inhibited the phosphorylation of well-known signaling relays downstream of FLT3, such as Akt S⁴⁷³, ERK T²⁰²/Y²⁰⁴ and STAT5 Y⁶⁹⁴, similarly to AC-220 (Figure 1A). Our results highly suggest that SGI-1776 directly inhibits the FLT3 kinase activity in AML. This conclusion could also explain the observation made by Chen and colleagues that SGI-1776 treatment results in an increased maturation of the FLT3-ITD in MV4-11 cells, assessed by an increased accumulation of the mature 150 kDa form compared with the 130 kDa endoplasmic reticulum form in SGI-1776–treated cells.³  Indeed, it has been previously demonstrated by Schmidt-Arras and coworkers that inhibition of FLT3-ITD catalytic activity promotes the maturation of the receptor with acquisition of complex glycosylation, leading to an increase of FLT3-ITD molecular mass from 130 to 150 kDa.⁶  We further showed that both AC-220 and SGI-1776 increased annexin V binding (Figure 1B) and induced the cleavage of caspase-3 (Figure 1C) in the MOLM-14 (FLT3-ITD) but not in the OCI-AML3 (FLT3-WT) AML cell lines. Although Chen and colleagues observed apoptosis in SGI-1776–treated OCI-AML3 cells, higher doses of SGI-1776 were required in their experiments to induce apoptosis in these cells expressing FLT3-WT than in MV4-11 or MOLM-13 cells that express FLT3-ITD receptors. Similarly, higher doses of SGI-1776 were required to inhibit Pim target phosphorylation or RNA synthesis.³  Thus, our explanation is that the inhibitory effects of SGI-1776 reported by Chen and colleagues are mostly because of the inhibition of FLT3 catalytic activity. Overall, we assume that no definitive conclusion on the role of the Pim kinases in the survival of AML cells could be established as yet.