Immunomodulatory agents lenalidomide and pomalidomide co-stimulate T cells by inducing degradation of T cell repressors Ikaros and Aiolos via modulation of the E3 ubiquitin ligase complex CRL4(CRBN.).
Bottom Line: We confirm that Aiolos and Ikaros are transcriptional repressors of interleukin-2 expression.The findings link lenalidomide- or pomalidomide-induced degradation of these transcriptional suppressors to well documented T cell activation.In conclusion, we present a molecular model in which drug binding to cereblon results in the interaction of Ikaros and Aiolos to CRL4(CRBN) , leading to their ubiquitination, subsequent proteasomal degradation and T cell activation.
Affiliation: Celgene Corporation, Summit, NJ, USA.Show MeSH
Mentions: Aiolos and Ikaros are known negative regulators of the IL2 promoter in T cells (Bandyopadhyay et al, 2007; Gandhi et al, 2010; Quintana et al, 2012). To investigate whether they are direct mediators of the known lenalidomide- and pomalidomide-induced upregulation of IL2, we knocked down Aiolos in T cells alone or in combination with drug treatment. T cells transfected with Aiolos siRNA for 24 h displayed a 61% decrease in IKZF3 RNA and a 67% decrease in Aiolos protein (Fig 6A). Reduction of Aiolos resulted in a corresponding 2·5 fold increase in IL2 mRNA (P < 0·0001; Fig 6B), confirming that Aiolos is a negative transcriptional regulator of the IL2 gene. There was a concomitant twofold increase in IL2 protein expression (P < 0·01; Fig 6C, compare black bars within each graph). Control siRNA-treated T cells resulted in the expected enhancement of IL2 protein production as previously reported (Fig 6C, left half of graphs; Lopez-Girona et al, 2012). The Aiolos siRNA-mediated reduction in Aiolos protein expression was further augmented by the addition of drug (Fig 6D). This correlated with the finding that the combination of lenalidomide or pomalidomide treatment and Aiolos siRNA expression further enhanced IL2 protein expression beyond that of Aiolos siRNA or drug treatment alone (Fig 6C, right half of graphs). Even a partial loss of Aiolos resulted in the induction of IL2 and stimulation of T cells. Additional cytokines upregulated by Aiolos siRNA and further enhanced by drug treatment include IL4, IL6, IL10, IL13, granulocyte macrophage colony-stimulating factor (GM-CSF) and interferon γ (IFN-γ; Fig S1). However, cytokine levels of IL8 and IL12 remained unchanged by Aiolos siRNA and/or drug treatment (Fig S2). The Aiolos expression levels after drug treatment in the presence or absence of Aiolos siRNA indicate a reverse correlation between Aiolos expression and IL2 production (Fig 6D). Therefore, Aiolos knockdown mimics the pharmacological effects of lenalidomide or pomalidomide on T cells.