The synthetic triterpenoid CDDO-imidazolide induces monocytic differentiation by activating the Smad and ERK signaling pathways in HL60 leukemia cells

Synthetic triterpenoids, CDDO (2-cyano-3,12-dioxooleana-1,9-dien-28-oic acid) or CDDO-imidazolide [2-cyano-3,12-dioxooleana-1,9-dien-28-oic acid imidazolide (CDDO-lm)], induce cell differentiation in myeloid leukemia cells but their mechanism of action is not known. CDDO-im induces monocytic differentiation markers, CD14, and nonspecific esterase in HL60 leukemia cells. We show that CDDO-lm activates the extracellular signal-regulated kinase [ERK) signaling pathway and up-regulates CCAAT/enhancer-binding protein 13, a transcription factor critical for monocytic differentiation. The monocytic differentiation induced by CDDO-lm was partially blocked by the mitogen-activated protein kinase/ERK kinase 1 inhibitor PD98059, suggesting that the mitogen-activated protein kinase-ERK1/2 pathway plays a role in the differentiation induced by CDDO-lm. Furthermore, CDDO-lm activates the transforming growth factor beta (TGF-beta)/Smad signaling pathway. CDDO-lm enhanced the phosphorylation of the receptor-regulated Smads, phospho-Smad3, and phospho-Smad1/5, but not phospho-Smad2, and induced the expression of Smad4. Monocytic differentiation induced by CDDO-lm was blocked by both TGF-beta antibody and the bone morphogenetic protein (BMP) antagonist Noggin. This indicates that activation of the Smad signaling pathway by triterpenoids is an important mechanism of monocytic differentiation. CDDO-lm induced the synthesis of mRNA for TGF-beta 2, BMP6, TGF-beta type 11 receptor, and BMP type 11 receptor. CDDO-lm synergized with members of the TGF-beta superfamily or with 1 alpha,25(OH)(2)Vitamin D-3 (D3) in monocytic differentiation, and the synergistic effect was particularly striking in combination with D3. The combination of triterpenoids and D3 may have a practical use in differentiation therapy of myeloid leukemia as well as for promoting the formation of bone and cartilage.