Role of downstream signaling events of interleukin-6 triggered JAK/STAT3 and PI3K/AKT signaling pathways in the development of resistance to dexamethasone by 7TD1 B-cell murine hybridoma

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Multiple myeloma (MM), a B-cell malignancy with poor prognosis, utilizes interleukin-6 (IL6) as an important growth factor for proliferation and survival, and IL6 may play a role in patient relapse and resistance. In this report we have utilized the IL6-dependent 7TD1 murine B-cell hybridoma as an in vitro model to study the role of IL6-triggered signaling pathways, JAK/STAT3 and PI3K/AKT in the development of resistance to dexamethasone (Dxm). Initially, 7TD1 cell growth is IL6-dependent and dexamethasone promotes growth arrest and apoptosis. However, chronic exposure led to a population of cells (7TD1-Dxm) which were resistant to dexamethasone and grew independent of exogenous IL6. Aim of our study was to investigate the role of inhibition of downstream signaling events in IL6-triggered JAK/STAT3 and PI3K/AKT signaling pathways in development of resistance to dexamethasone by 7TD1-Dxm cells. Previous studies in this laboratory have shown that STAT3 was constitutively phosphorylated in resistant cells and its phosphorylation was resistant to inhibition by AG490. Treatment of resistant cells with a membrane-permeable STAT3 inhibitor peptide induced apoptosis but did not alter their insensitivity to dexamethasone. Also, treatment with the PI3K inhibitor, LY294002, inhibited proliferation of 7TD1-Dxm cells suggesting the role of PI3K/AKT signaling pathway in promoting proliferation of the cells. Current results employing HA-14, a Bcl-2 specific inhibitor, at a concentration of 20 µM, and rapamycin, inhibitor of mTOR, at a concentration of 5 nM, suggested that they inhibited proliferation of both 7TD1 and 7TD1-Dxm cells. Results from TUNEL assay showed that treatment with HA-14 at 20 µM alone induced spontaneous apoptosis but did not alter insensitivity of 7TD1-Dxm cells to dexamethasone and treatment with rapamycin at 5 nM alone did not induce apoptosis in either cell line. Also, treatment of 7TD1-Dxm cells with HA-14 at a concentration lesser than that needed for induction of spontaneous apoptosis did not alter insensitivity to dexamethasone. In conclusion, our results suggest that there are multiple mechanisms involved in the development of resistance to dexamethasone by 7TD1-Dxm cells. Further investigations focusing on Bcl-2 protein expression and mitochondrial involvement may give an idea on the mechanisms leading to the development of resistance to dexamethasone by 7TD1-Dxm cells.
 This study was partially supported by NIH Grant # P20 RR016454 from the INBRE Program of the National Center for Research Resources.