BackgroundThe CXCL12/CXCR4 axis transactivates HER2 and promotes intraosseous tumor growth. To further explore the transactivation of HER2 by CXCL12, we investigated the role of small GTP protein Gαi2 in Src and HER2 phosphorylation in lipid raft membrane microdomains and the significance of CXCR4 in prostate cancer bone tumor growth.MethodsWe used a variety of methods such as lipid raft isolation, invasion assays, an in vivo model of intratibial tumor growth, bone histomorphometry, and immunohistochemistry to determine the role of CXCR4 signaling in lipid raft membrane microdomains and effects of targeting of CXCR4 for bone tumor growth.ResultsWe determined that (a) CXCL12/CXCR4 transactivation of EGFR and HER2 is confined to lipid raft membrane microdomains, (b) CXCL12 activation of HER2 and Src is mediated by small GTP proteins in lipid rafts, (c) inhibition of the CXCL12/CXCR4 axis through plerixafor abrogates the initial establishment of tumor growth without affecting the growth of established bone tumors, and (d) inhibition of EGFR signaling through gefitinib leads to inhibition of established bone tumor growth.ConclusionsThese data suggest that lipid raft membrane microdomains are key sites for CXCL12/CXCR4 transactivation of HER2 via small GTP binding protein Gαi2 and Src kinase. The initial establishment of prostate cancer is supported by the endosteal niche, and blocking the CXCL12/CXCR4 axis of this niche along with its downstream signaling severely compromises initial establishment of tumors in the bone microenvironment, whereas expanding bone tumors are sensitive only to the members of growth factor receptor inhibition.Electronic supplementary materialThe online version of this article (doi:10.1186/s12943-016-0552-0) contains supplementary material, which is available to authorized users.

Purpose Intratumoral androgen synthesis in prostate cancer (PCa) contributes to the development of castration-resistant prostate cancer (CRPC). Several enzymes responsible for androgen biosynthesis have been shown to be overexpressed in CRPC, thus contributing to CRPC in a castrated environment. The TMPRSS2-ERG transcription factor has been shown to be present in primary PCa tumors as well as CRPC tumors. We hypothesize that TMPRSS2-ERG fusions regulate androgen biosynthetic enzyme (ABE) gene expression and the production of androgens, which contributes to the development of CRPC. Experimental design We used a panel of assays including lentivirus transduction, gene expression, chromatin immunoprecipitation and sequencing, Liquid chromatography-Mass spectrometric quantitation, immunocytochemistry, immunohistochemistry and bio-informatics analysis of gene microarray data bases to determine ERG regulation of androgen synthesis. Results We found that ERG regulated the expression of the ABE AKR1C3 in PCa cells via direct binding to the AKR1C3 gene. Knockdown of ERG resulted in reduced AKR1C3 expression, which caused a reduction in both DHT synthesis and PSA expression in VCaP PCa cells treated with 5α-androstanedione, a DHT precursor metabolite. Immunohistochemical staining revealed that ERG was co-expressed with AKR1C3 in PCa tissue samples. Conclusions These data suggest that AKR1C3 catalyzes the biochemical reduction of 5α-Androstanedione to DHT in PCa cells, and that ERG regulates this step through upregulation of AKR1C3 expression. Elucidation of ERG regulation of ABEs in CRPC may help to stratify TMPRSS2-ERG fusion-positive PCa patients in the clinic for anti-AR driven therapies; and AKR1C3 may serve as a valuable therapeutic target in the treatment of CRPC.

We have shown that overexpression of SMS1, an enzyme that converts de novo ceramide into sphingomyelin, is accompanied by attenuated ceramide response and apoptotic resistance after photodamage with the photosensitizer Pc 4 (photodynamic therapy; PDT). To test whether SMS1 overexpression-related effects after PDT can be reversed, in this study SMS1 was downregulated in Jurkat T lymphoma/leukemia cells using small inhibitory RNA (siRNA) for SMS1. Compared to scrambled (control) siRNA-transfectants, in SMS1 siRNA-transfected cells the activity of SMS at rest was downregulated with concomitant decrease in sphingomyelin mass. In SMS1 siRNAtransfected cells increases in ceramides were higher than in control siRNA-transfectants after PDT. Similar findings were obtained for dihydroceramides suggesting the involvement of de novo ceramide pathway. PDT-induced DEVDase (caspase-3-like) activation was enhanced in SMS1 siRNA-transfected cells compared to their control counterparts. The data show that RNA interference-dependent downregulation of SMS1 is associated with increased accumulation of ceramide and dihydroceramide with concomitant sensitization of cells to apoptosis after photodamage. Similarly, in SMS2 siRNA-transfected cells, downregulation of SMS activity was accompanied by potentiated DEVDase activation post-photodamage. These findings suggest that SMS is a potential novel molecular target that can augment therapeutic efficacy of PDT.

CXCR4 is a chemokine receptor that mediates invasion and metastasis. CXCR4 expression is transcriptionally regulated in cancer cells and is associated with aggressive phenotypes of prostate cancer. Previously, we and others have shown that the ERG transcription factor regulates CXCR4 expression in prostate cancer cells. We further showed that androgens regulate CXCR4 expression via increasing ERG transcription factor expression. Herein, we investigated molecular mechanisms of ERG-mediated CXCR4 promoter activation, phosphorylation of ERG by intracellular kinases and subsequent CXCR4 expression, as well as expression of ERG and CXCR4 in human prostate tumor tissues. Using multiple molecular strategies, we demonstrate that: (a) ERG expressed in TMPRSS2-ERG fusion positive VCaP cells selectively binds with specific ERG/Ets bindings sites in the CXCR4 promoter; (b) distal binding sites mediate promoter activation; (c) exogenously expressed ERG promotes CXCR4 expression; (d) ERG is phosphorylated at Serine 81 and 215, both IKK and Akt kinases induce serine phosphorylation, and Akt mediates CXCR4 expression; (e) ERG-induced CXCR4 drives CXCL12-dependent adhesion to fibronectin; (f) ERG and CXCR4 were co-expressed in human prostate tumor tissues, consistent with ERG-mediated transcriptional activation of CXCR4. These data demonstrates that ERG factor activates CXCR4 expression by binding to the specific ERG/Ets responsive elements and intracellular kinases phosphorylate at ERG at serine residues to induce CXCR4 expression. These findings may provide a mechanistic link between TMPRSS2-ERG translocations and intracellular kinase mediated phosphorylation of ERG on enhanced metastasis of tumor cells via CXCR4 expression and function in prostate cancer cells.