Effect of Thalidomide and Revlimid on the Gene Expression That Reveals Molecular Circuitries Involved in T Cell Co-Activation.

Thalidomide (Thal) and its immunomodulatory analog, Revlimid (Rev) have demonstrated anti-myeloma activity in phase II and III studies and part of their activity is immune-mediated. We have previously demonstrated co-stimulatory effects of Thal and Rev on T cells mediated via B7-CD28 activation pathway. To understand the molecular events involved in T cell co-stimulation, we investigated the effects of Thal and Rev on the genome-wide gene expression profile of T cells with or without anti-CD3 activation. Normal PBMC were incubated with Thal or Rev in the presence or absence of anti-CD3 antibodies (200ng/ml) for 3 or 24 hours at 37oC and total RNA was analyzed using U133 GeneChip (Affymetrix). Although Thal and Rev do not show significant effect on T cell proliferation without CD3 stimulation, we observed up-regulation (>2-folds) of 56 genes and down-regulation of 42 genes at 3 hours, and up-regulation of 41genes and down-regulation of 35 genes at 24 hours with Rev. Significant up-regulation of IFN-α (10-folds) was observed at 3 hours. Up-regulation of chemokine CXC5 (9 fold); IFN-β (5 fold), IL-7 (5 fold) and IL-2 (3 fold); and signaling genes like JK 3 (10 folds) and MAPKK5 (5 folds) was observed at 24 hours with Rev. Following anti-CD3 activation, Rev induced significant upregulation of 59 and 48 genes and down-regulation of 126 and 61 genes at 3 and 24 hours respectively. These include down-regulation of Th2 cytokine genes IL-5(5 folds) and TGF-β (2 folds), and apoptotsis related genes, TNFSF12 (4 folds) and TNFRSF 6b (4 folds). Genes involved in B7-CD28 activation pathway and up-regulated include CXC 5 (9 folds), IL-8 (5 folds) and IL-3 (4 folds). Interestingly, p52, which is a key molecule in CD28 signaling, was elevated 18 folds. Additionally less than 2 fold increase was observed in other CD28 signaling genes like IKK, Vov, and RelA (p65). Rev also down-regulated, at 24 hours, the expression of IL-10 (6 folds), IL-17 (3 folds) and TGF-β (3 folds) which allows sustained co-stimulation. Similar changes with lesser magnitude were observed with Thal. These results provide insight into early molecular changes induced by Thal and Rev that is important for co-stimulation and suggest new molecular targets to generate effective immune responses. Moreover, these observations facilitate the pre-clinical rationale for the use of these immunomodulatory compounds to improve clinical outcome in myeloma.