Growth inhibition of a human myeloma cell line by all-trans retinoic acid is not mediated through downregulation of interleukin-6 receptors but through upregulation of p21(WAF1).
All-trans retinoic acid (ATRA) has previously been shown to inhibit the growth of OPM-2 human myeloma cells. The growth inhibition was postulated to result from a transcriptional downregulation of interleukin-6 receptor alpha (IL-6Ralpha) with IL-6Rbeta (gp130) unaffected. To formally test this hypothesis, an expression vector designed for constitutive IL-6Ralpha expression was constructed and used for transfection of OPM-2 cells. Six stable transfectants were cloned. The expression of IL-6Ralpha was shown by immunofluorescence with anti-IL-6Ralpha antibody and 125I-IL-6 binding. In five of six transfectant clones, cellular IL-6Ralpha was 1.5- to 6-fold higher than the parental cells, with the ligand binding affinity unchanged. While ATRA reduced IL-6Ralpha expression in the parental OPM-2 cells, it enhanced its expression in these five transfectants. The clonogenic growth of these transfectants, however, remained strongly inhibited by ATRA. Further analysis, comparing the parental OPM-2 cells and a representative transfectant, clone C5, showed that IL-6 caused rapid tyrosine phosphorylation of gp130 in both OPM-2 and C5 clones. Pretreatment with ATRA greatly reduced IL-6-induced gp130 phosphorylation in OPM-2 cells, reflecting a reduction in cellular IL-6Ralpha. In contrast, IL-6-induced gp130 phosphorylation was not reduced by ATRA pretreatment in C5 cells, indicating that the expressed IL-6Ralpha was functional. Similar to OPM-2 cells, C5 cells were sensitive to growth inhibition by dexamethasone, which was entirely reversed by exogenous IL-6, suggesting that the IL-6 postreceptor signal transduction remained intact. ATRA was further shown to upregulate p21(WAF1) expression and cause dephosphorylation of the retinoblastoma protein (pRB) in both OPM-2 and C5 cells. Exogenous IL-6 also failed to reverse these effects of ATRA. Thus, the growth inhibitory activity of ATRA is not mediated through cellular IL-6Ralpha downregulation and is likely to result from a direct upregulation of p21(WAF1) and consequent dephosphorylation of pRB.
Chen
Y H
YH
Department of Medicine, University of Illinois College of Medicine and VA West Side Medical Center, Chicago, IL 60612, USA.
Lavelle
D
D
DeSimone
J
J
Uddin
S
S
Platanias
L C
LC
Hankewych
M
M
D000970:Antineoplastic Agents / Q000494:pharmacology* / Q000627:therapeutic use; D002455:Cell Division / Q000187:drug effects; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins / Q000378:metabolism*; D015536:Down-Regulation; D006801:Humans; D009101:Multiple Myeloma / Q000188:drug therapy* / Q000378:metabolism* / Q000473:pathology; D019947:Receptors, Interleukin-6 / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects; D014162:Transfection; D014212:Tretinoin / Q000494:pharmacology* / Q000627:therapeutic use; D014407:Tumor Cells, Cultured; D015854:Up-Regulation
D000970:Antineoplastic Agents; C493956:CDKN1A protein, human; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D019947:Receptors, Interleukin-6; D014212:Tretinoin