Publication Date:
2016-12-02
Description:
The use of the immunomodulatory drugs (IMiDs®), including lenalidomide (LEN), to reverse tumor-mediated immune suppression and amplify multiple myeloma (MM)-specific immunity is currently being explored. Particularly LEN effects on antigen presenting cells are still unknown. In this study we investigated LEN potential effect on Dendritic Cell (DC) differentiation and activity and on the immunosuppressive properties of human mesenchymal stromal cells (hMSCs) on DCs. DCs were obtained either from THP1 (THP1-DC), a human monocytic cell line, or from CD14+ cells, purified by an immunomagnetic method both from peripheral blood (PB) and bone marrow (BM) samples of a total cohort of 19 patients with monoclonal gammopathies including 13 with active MM. Monocyte-derived DC (mo-DC) differentiation was induced by the treatment with GM-CSF and IL-4 for 8 days and TNF-α for the last 24 hours. LEN treatment was performed at concentration ranging from 0.1 to 2 μM. Then, non-adherent cells were analyzed for DC maturation markers (CD83, HLA-DR, CD80, CD86 and CD209) by flow cytometry. The levels of soluble factors involved in the immune response (CCL2, CCL5, CXCL10, IL-6, IL-8, IL-10, IL-12, IFN-γ, TNF-α) were measured in the conditioned media (CM) of mo-DCs by a Bio-Plex® Multiplex System. Moreover, the effect of LEN treatment on DC ability to stimulate T-cell proliferation was investigated through an MTT assay on CD3+ cells, purified from PB of MM patients, co-cultured for 6 days with LEN pretreated autologous mo-DCs, in RPMI medium at 15% AB human serum. Ex vivo studies were also performed in order to evaluate mo-DC differentiation of CD14+ cells purified from PB of 4 MM patients before and after 7 days of LEN treatment at 25 mg/day without Dexamethasone. Moreover, mo-DC differentiation was performed in the presence of CM of human TERT transfected (hTERT) -hMSCs treated for 5 days with LEN (0.1-2 μM). The expression levels of factors with an inhibitory effect on DC development (CCL5, IDO1, IL6, PTGS2, TGFB) were tested in LEN treated hTERT-hMSCs by real time PCR. Finally, the protein levels of Cereblon and its transcription factor targets Aiolos, Ikaros, IRF-4, p62 and Casein kinase (CK) 1 and 2 were evaluated either in THP1-DCs or in hTERT-hMSCs LEN treated cells by Western Blotting. Despite a reduction of both number and % of mature mo-DCs, LEN at the concentration range reached in vivo in MM patients significantly increased the median intensity expression of HLA-DR (LEN vs vehicle, p
Print ISSN:
0006-4971
Electronic ISSN:
1528-0020
Topics:
Biology
,
Medicine
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