Anti-ITGB5 (aa 421-516) polyclonal antibody

Overexpression of the oncogene ERG (ETS-related gene) is an adverse prognostic factor in acute myeloid and T-cell lymphoblastic leukemia (AML and T-ALL). We hypothesize that ERG overexpression is associated with primary drug resistance thereby influencing the outcome in leukemia. We previously reported a cell-line based model of ERG overexpression which induced a potentially chemo-resistant spindle shape cell type. Herein, we report a specific transcriptional gene signature for the observed spindle shaped morphology. Genes significantly over-expressed after ERG induction strongly resembled adhesive mesenchymal-like genes that included integrins (ITGA10, ITGB5, ITGB3, ITGA2B), CD44, and CD24. Interestingly, the mesenchymal-like signature was accompanied by the repression of DNA chromatin remodeling and DNA repair genes, such as CHEK1, EZH2, SUZ12, and DNMT3a. The ERG-induced mesenchymal-like signature positively correlated with TMPRSS2-ERG prostate tissues and invasive breast cancer mRNA expression datasets reflecting a general ERG-driven pattern of malignancy. Furthermore, inhibitors modulating ERG druggable pathways WNT, PKC, and AKT, and chemotherapeutic agent cytarabine revealed ERG-induced drug resistance. In particular, PKC412 treatment enhanced proliferative rates and promoted spindle shape formation in ERG-induced cells. Nilotinib and dasatinib were effective at abolishing ERG-induced cells. Moreover, ERG overexpression also led to an increase in double strand breaks. This report provides mechanistic clues into ERG-driven drug resistance in the poor prognostic group of high ERG expressers, provides insight to improved drug targeted therapies, and provides novel markers for a mesenchymal-like state in acute leukemia.

Introduction The pathophysiology of systemic sclerosis (SSc) is closely linked to overactive TGF beta signaling. TGF beta is produced and circulates in latent form, making its activation crucial for signaling. This activation can be mediated via integrins. We investigated the balance between active and latent TGF beta in serum of SSc patients and investigated if this correlates with integrin expression on monocytes. Methods A TGF beta/SMAD3- or BMP/SMAD1/5-luciferase reporter construct was expressed in primary human skin fibroblasts. Both acidified and non-acidified sera of ten SSc patients and ten healthy controls were tested on these cells to determine total and active TGF beta and BMP levels respectively. A pan-specific TGF beta 1/2/3 neutralizing antibody was used to confirm TGF beta signaling. Monocytes of 20 SSc patients were isolated using CD14+ positive selection, and integrin gene expression was measured using qPCR. Integrin expression was modulated using rhTGF beta 1 or a small molecule inhibitor of TGFBR1: SB-505124. Results SSc sera induced 50% less SMAD3-reporter activity than control sera. Serum acidification increased reporter activity, but a difference between healthy control and SSc serum was no longer observed, indicating that total TGF beta levels were not different. Addition of a pan-specific TGF beta 1/2/3 neutralizing antibody fully inhibited SMAD3-reporter activity of both acidified and not-acidified control and SSc sera. Both HC and SSc sera induced similar SMAD1/5-reporter activity, and acidification increased this, but not differently between groups. Interestingly, expression of two integrin alpha subunits ITGA5 and ITGAV was significantly reduced in monocytes obtained from SSc patients. Furthermore, ITGB3, ITGB5, and ITGB8 expression was also reduced in SSc monocytes. Stimulation of monocytes with TGF beta 1 induced ITGA5 and ITGAV but lowered ITGB8 expression, whereas the use of the TGF beta receptor inhibitor SB-505124 had the opposite effect. Conclusion Total TGF beta serum levels are not different between SSc patients and controls, but TGF beta activity is. This coincides with a reduced expression of TGF beta-activating integrins in monocytes of SSc patients. Because TGF beta regulates expression of these integrins in monocytes, a negative feedback mechanism possibly underlies these observations.