MicroRNA-4719 and microRNA-6756-5p Correlate with Castration-Resistant Prostate Cancer Progression through Interleukin-24 Regulation.

Prostate cancer (PCa) is the second leading cause of cancer death in the United States. The five-year survival rate for men diagnosed with localized PCa is nearly 100%, yet for those diagnosed with aggressive PCa, it is less than 30%. The pleiotropic cytokine Interleukin-24 (IL-24) has been shown to specifically kill PCa cells compared to normal cells when overexpressed in both in vitro and in vivo studies. Despite this, the mechanisms regulating IL-24 in PCa are not well understood. Since specific microRNAs (miRNAs) are dysregulated in PCa, we used miRNA target prediction algorithm tools to identify miR-4719 and miR-6556-5p as putative regulators of IL-24. This study elucidates the expression profile and role of miR-4719 and miR-6756-5p as regulators of IL-24 in PCa. qRT-PCR analysis shows miR-4719 and miR-6756-5p overexpression significantly decreases the expression of IL-24 in PCa cells compared to the negative control. Compared to the indolent PCa and normal prostate epithelial cells, miR-4719 and miR-6756-5p are significantly overexpressed in castration-resistant prostate cancer (CRPC) cell lines, indicating that their gain may be an early event in PCa progression. Moreover, miR-4719 and miR-6756-5p are significantly overexpressed in the CRPC cell line of African-American males (E006AA-hT) compared to CRPC cell lines of Caucasian males (PC-3 and DU-145), indicating that miR-4719 and miR-6756-5p may also play a role in racial disparity. Lastly, the inhibition of expression of miR-4719 and miR-6756-5p significantly increases IL-24 expression and inhibits proliferation and migration of CRPC cell lines. Our findings indicate that miR-4719 and miR-6756-5p may regulate CRPC progression through the targeting of IL-24 expression and may be biomarkers that differentiate between indolent and CRPC. Strategies to inhibit miR-4719 and miR-6756-5p expression to increase IL-24 in PCa may have therapeutic efficacy in aggressive PCa.

IL-24 Promotes Apoptosis through cAMP-Dependent PKA Pathways in Human Breast Cancer Cells.

Interleukin 24 (IL-24) is a tumor-suppressing protein, which inhibits angiogenesis and induces cancer cell-specific apoptosis. We have shown that IL-24 regulates apoptosis through phosphorylated eukaryotic initiation factor 2 alpha (eIF2α) during endoplasmic reticulum (ER) stress in cancer. Although multiple stresses converge on eIF2α phosphorylation, the cellular outcome is not always the same. In particular, ER stress-induced apoptosis is primarily regulated through the extent of eIF2α phosphorylation and activating transcription factor 4 (ATF4) action. Our studies show for the first time that cyclic adenosine monophosphate (cAMP)-dependent protein kinase A (PKA) activation is required for IL-24-induced cell death in a variety of breast cancer cell lines and this event increases ATF4 activity. We demonstrate an undocumented role for PKA in regulating IL-24-induced cell death, whereby PKA stimulates phosphorylation of p38 mitogen-activated protein kinase and upregulates extrinsic apoptotic factors of the Fas/FasL signaling pathway and death receptor 4 expression. We also demonstrate that phosphorylation and nuclear import of tumor suppressor TP53 occurs downstream of IL-24-mediated PKA activation. These discoveries provide the first mechanistic insights into the function of PKA as a key regulator of the extrinsic pathway, ER stress, and TP53 activation triggered by IL-24.

Translation Control by p53.

The translation of mRNAs plays a critical role in the regulation of gene expression and therefore, in the regulation of cell proliferation, differentiation and apoptosis. Unrestricted initiation of translation causes malignant transformation and plays a key role in the maintenance and progression of cancers. Translation initiation is regulated by the ternary complex and the eukaryotic initiation factor 4F (eIF4F) complex. The p53 tumor suppressor protein is the most well studied mammalian transcription factor that mediates a variety of anti-proliferative processes. Post-transcriptional mechanisms of gene expression in general and those of translation in particular play a major role in shaping the protein composition of the cell. The p53 protein regulates transcription and controls eIF4F, the ternary complex and the synthesis of ribosomal components, including the down-regulation of rRNA genes. In summary, the induction of p53 regulates protein synthesis and translational control to inhibit cell growth.

Eukaryotic Translation Initiation Factor 4A Down-Regulation Mediates Interleukin-24-Induced Apoptosis through Inhibition of Translation.

Dysregulated activity of helicase eIF4A drives transformation to and maintenance of cancer cell phenotype by reprogramming cellular translation. Interleukin 24 (IL-24) is a tumor-suppressing protein, which has the ability to inhibit angiogenesis, sensitize cancer cells to chemotherapy, and induce cancer cell-specific apoptosis. In this study, we found that eIF4A is inhibited by IL-24. Consequently, selective reduction of translation was observed for mRNAs harboring strong secondary structures in their 5'-untranslated regions (5'UTRs). These mRNAs encode proteins, which function in cell survival and proliferation. Consistently, overexpression of eIF4A conferred cancer cells with resistance to IL-24-induced cell death. It has been established that inhibition of eIF4A triggers mitochondrial-mediated apoptosis. We showed that IL-24 induces eIF4A-dependent mitochondrial depolarization. We also showed that IL-24 induces Sigma 1 Receptor-dependent eIF4A down-regulation and mitochondrial depolarization. Thus, the progress of apoptosis triggered by IL-24 is characterized by a complex program of changes in regulation of several initiation factors, including the eIF4A.

eIF2α Phosphorylation Mediates IL24-Induced Apoptosis through Inhibition of Translation.

IL24 is an immunomodulatory cytokine that also displays broad cancer-specific suppressor effects. The tumor-suppressor activities of IL24 include inhibition of angiogenesis, sensitization to chemotherapy, and cancer-specific apoptosis. Supra-physiologic activation and/or overexpression of translation initiation factors are implicated in the initiation and progression of cancer animal models as well as a subset of human cancers. Activation and/or overexpression of translation initiation factors correlate with aggressiveness of cancer and poor prognosis. Two rate-limiting translation initiation complexes, the ternary complex and the eIF4F complex, are regulated by eIF2α and 4E-BP1 phosphorylation, respectively. The work reported here provides direct evidence that IL24 induces inhibition of translation initiation leading to apoptosis in squamous cell carcinoma. A dominant constitutively active mutant of eIF2α, which is resistant to phosphorylation, was used to determine the involvement of eIF2α in IL24-induced apoptosis. Treatment with IL24 resulted in inhibition of protein synthesis, expression of downstream biomarkers of ternary complex depletion such as CHOP, and induction of apoptosis in cancer cells. The constitutively active nonphosphorylatable mutant of eIF2α, eIF2α-S51A, reversed both the IL24-mediated translational block and IL24-induced apoptosis. Intriguingly, IL24 treatment also caused hypophosphorylation of 4E-BP1, which binds to eIF4E with high affinity, thus preventing its association with eIF4G and therefore preventing elF4F complex assembly.Implications: These results demonstrate a previously unrecognized role of IL24 in inhibition of translation, mediated through both phosphorylation of eIF2α and dephosphorylation of 4E-BP1, and provide the first direct evidence for translation control of gene-specific expression by IL24. Mol Cancer Res; 15(8); 1117-24. ©2017 AACR.

Mechanism of Action and Applications of Interleukin 24 in Immunotherapy.

Interleukin 24 (IL-24) is an important pleiotropic immunoregulatory cytokine, whose gene is located in human chromosome 1q32-33. IL-24's signaling pathways have diverse biological functions related to cell differentiation, proliferation, development, apoptosis, and inflammation, placing it at the center of an active area of research. IL-24 is well known for its apoptotic effect in cancer cells while having no such effect on normal cells. IL-24 can also be secreted by both immune and non-immune cells. Downstream effects of IL-24, after binding to the IL-20 receptor, can occur dependently or independently of the JAK/STAT signal transduction pathway, which is classically involved in cytokine-mediated activities. After exogenous addition of IL-24, apoptosis is induced in tumor cells independently of the JAK/STAT pathway. We have shown that IL-24 binds to Sigma 1 Receptor and this event induces endoplasmic reticulum stress, calcium mobilization, reactive oxygen species generation, p38MAPK activity, and ceramide production. Here we review IL-24's role in autoimmunity, infectious disease response, wound repair, and vascular disease. Detailed understanding of the pleiotropic roles of IL-24 signaling can assist in the selection of more accurate therapeutic approaches, as well as targeting of appropriate cell types in treatment strategy development, and ultimately achieve desired therapeutic effects.

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) activates caspases in human prostate cancer cells through sigma 1 receptor.

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-based therapy is currently evaluated in clinical studies as a tumor cell-selective pro-apoptotic approach. Unfortunately, many clinical studies have shown that cancer cells acquire TRAIL resistance and finally avoid TRAIL-induced apoptosis. Therefore, defining the mechanisms that permit TRAIL to activate apoptosis is critical for the development of strategies that maximize the potential effectiveness of TRAIL in clinical applications. This study aims at understanding the molecular mechanisms underlying TRAIL-induced apoptosis and unraveling signaling pathways that could revert sensitivity to apoptosis stimuli. Our current study demonstrates for the first time that Sigma 1 Receptor (Sig1R), a ligand-regulated protein chaperone, contributes to TRAIL induction of apoptosis. We show that Sig1R agonist (+)-SKF10047 action or increasing Sig1R expression, significantly reduced apoptosis by TRAIL in prostate cell lines, indicating the importance of Sig1R and signifying that higher levels of Sig1R in prostate cancer cells make them more resistant to TRAIL treatment. Here we show that Sig1R is critically involved in TRAIL-induced caspase activation. Furthermore, we show that Sig1R protein is degraded upon TRAIL treatment. Knockdown of Sig1R, by siRNA transfection increased the sensitivity of breast cancer cells to TRAIL. These results indicate that Sig1R could represent a promising molecule to sensitize human breast cancers to TRAIL. Collectively, these studies define Sig1R as a key mediator of TRAIL induction of cancer-specific killing.