Docetaxel-induced prostate cancer cell death involves concomitant activation of caspase and lysosomal pathways and is attenuated by LEDGF/p75

<p>Abstract</p> <p>Background</p> <p>Hormone-refractory prostate cancer (HRPC) is characterized by poor response to chemotherapy and high mortality, particularly among African American men when compared to other racial/ethnic groups. It is generally accepted that docetaxel, the standard of care for chemotherapy of HRPC, primarily exerts tumor cell death by inducing mitotic catastrophe and caspase-dependent apoptosis following inhibition of microtubule depolymerization. However, there is a gap in our knowledge of mechanistic events underlying docetaxel-induced caspase-independent cell death, and the genes that antagonize this process. This knowledge is important for circumventing HRPC chemoresistance and reducing disparities in prostate cancer mortality.</p> <p>Results</p> <p>We investigated mechanistic events associated with docetaxel-induced death in HRPC cell lines using various approaches that distinguish caspase-dependent from caspase-independent cell death. Docetaxel induced both mitotic catastrophe and caspase-dependent apoptosis at various concentrations. However, caspase activity was not essential for docetaxel-induced cytotoxicity since cell death associated with lysosomal membrane permeabilization still occurred in the presence of caspase inhibitors. Partial inhibition of docetaxel-induced cytotoxicity was observed after inhibition of cathepsin B, but not inhibition of cathepsins D and L, suggesting that docetaxel induces caspase-independent, lysosomal cell death. Simultaneous inhibition of caspases and cathepsin B dramatically reduced docetaxel-induced cell death. Ectopic expression of lens epithelium-derived growth factor p75 (LEDGF/p75), a stress survival autoantigen and transcription co-activator, attenuated docetaxel-induced lysosomal destabilization and cell death. Interestingly, LEDGF/p75 overexpression did not protect cells against DTX-induced mitotic catastrophe, and against apoptosis induced by tumor necrosis factor related apoptosis inducing ligand (TRAIL), suggesting selectivity in its pro-survival activity.</p> <p>Conclusion</p> <p>These results underscore the ability of docetaxel to induce concomitantly caspase-dependent and independent death pathways in prostate cancer cells. The results also point to LEDGF/p75 as a potential contributor to cellular resistance to docetaxel-induced lysosomal destabilization and cell death, and an attractive candidate for molecular targeting in HRPC.</p

Contribution of LEDGF/p75 to Prostate Cancer Chemoresistance

Prostate cancer (PCa) is the second leading cause of male cancer death in the United States, and is considered a health disparity because of its disproportionate incidence and mortality in African American (AA) males. A major challenge in PCa treatment is its recurrence into hormone refractory prostate cancer (HRPC). Docetaxel is the standard of care for first line chemotherapy to HRPC. Unfortunately, most patients ultimately manifest resistance to DTX and succumb to the disease. Circumventing DTX resistance in HRPC will require better understanding of the mechanisms by which this drug kills prostate tumor cells, and the genes that promote this resistance. We investigated the mechanism underlying DTX-induced death and observed that it involved the concomitant activation of caspase and lysosomal pathways. We also investigated the role of the stress response protein and transcription co-activator lens epithelium derived growth factor p75 (LEDGF/p75) in HRPC resistance to DTX. Our studies show that LEDGF/p75 overexpression attenuates DTX-induced lysosomal destabilization and cell death in a cellular model of HRPC, implicating this stress protein in DTX resistance in virto. However, LEDGF/p75 did not protect against cell death induced by the classical apoptosis inducers TRAIL and staurosporine. This selectivity might be related to LEDGF/p75’s ability to protect cells against insults that induce ROS generation, since we observed that DTX but not TRAIL and staurosporine generated ROS. To better understand the mechanism by which LEDGF/p75 confers its chemoresistant phenotype, a protein expression profile was performed in PCa cells stably overexpressing LEDGF/p75. A marked increase in the expression of the thiol-oxidoreductase ERp57 was observed in overexpressing cells. Transactivation of ERp57 by LEDGF/p75 was confirmed using transcription reporter assays, providing evidence that ERp57 might be a target gene of LEDGF/p75. However, there was incomplete correlation in the expression patterns of LEDGF/p75 and ERp57 in PCa cell lines and tissues. These findings suggest that LEDGD/p75 might contribute to cellular resistance to drugs or insults that induce cell death associated with ROS generation. If proven to contribute to DTX resistance in an in vivo model, LEDGF/p75 and its target genes could be attractive targets for novel therapeutic strategies in the treatment of HRPC

Additional file 1: of The anti-fibrotic agent pirfenidone synergizes with cisplatin in killing tumor cells and cancer-associated fibroblasts

MTS assay showing the viability of H358 ( A ), H1299 ( B ), H23 ( C ), H157 ( D ), H2122 ( E ), and PC9 ( F ) after 72-h treatment with a low dose of cisplatin (Cis; 10 μM) and a low dose of pirfenidone (Pirf; 0.5 mg/mL). * P < 0.05. (PDF 1253 kb

Additional file 2: of The anti-fibrotic agent pirfenidone synergizes with cisplatin in killing tumor cells and cancer-associated fibroblasts

Immunoblot analysis shows an increase in phosphorylated ERK when A549 cells are treated with low doses of cisplatin (10 μM) and low doses of pirfenidone (0.5 mg/mL) at early time points (15 and 30 min; A ) when compared to each drug alone. B: At 48 h after treatment, there is a decrease in phosphorylated Akt when compared to each drug alone. (PDF 1211 kb

A Novel Antagonist of the Immune Checkpoint Protein Adenosine A2a Receptor Restores Tumor-Infiltrating Lymphocyte Activity in the Context of the Tumor Microenvironment

BACKGROUND: Therapeutic strategies targeting immune checkpoint proteins have led to significant responses in patients with various tumor types. The success of these studies has led to the development of various antibodies/inhibitors for the different checkpoint proteins involved in immune evasion of the tumor. Adenosine present in high concentrations in the tumor microenvironment activates the immune checkpoint adenosine A2a receptor (A2aR), leading to the suppression of antitumor responses. Inhibition of this checkpoint has the potential to enhance antitumor T-cell responsiveness. METHODS: We developed a novel A2aR antagonist (PBF-509) and tested its antitumor response in vitro, in a mouse model, and in non-small cell lung cancer patient samples. RESULTS: Our studies showed that PBF-509 is highly specific to the A2aR as well as inhibitory of A2aR function in an in vitro model. In a mouse model, we found that lung metastasis was decreased after treatment with PBF-509 compared with its control. Furthermore, freshly resected tumor-infiltrating lymphocytes from lung cancer patients showed increased A2aR expression in CD4+ cells and variable expression in CD8+ cells. Ex vivo studies showed an increased responsiveness of human tumor-infiltrating lymphocytes when PBF-509 was combined with anti-PD-1 or anti-PD-L1. CONCLUSIONS: Our studies demonstrate that inhibition of the A2aR using the novel inhibitor PBF-509 could lead to novel immunotherapeutic strategies in non-small cell lung cancer

Transforming growth factor β signaling overcomes dasatinib resistance in lung cancer.

Lung cancer is the second most common cancer and the leading cause of cancer-related deaths. Despite recent advances in the development of targeted therapies, patients with advanced disease remain incurable, mostly because metastatic non-small cell lung carcinomas (NSCLC) eventually become resistant to tyrosine kinase inhibitors (TKIs). Kinase inhibitors have the potential for target promiscuity because the kinase super family is the largest family of druggable genes that binds to a common substrate (ATP). As a result, TKIs often developed for a specific purpose have been found to act on other targets. Drug affinity chromatography has been used to show that dasatinib interacts with the TGFβ type I receptor (TβR-I), a serine-threonine kinase. To determine the potential biological relevance of this association, we studied the combined effects of dasatinib and TGFβ on lung cancer cell lines. We found that dasatinib treatment alone had very little effect; however, when NSCLC cell lines were treated with a combination of TGFβ and dasatinib, apoptosis was induced. Combined TGFβ-1 + dasatinib treatment had no effect on the activity of Smad2 or other non-canonical TGFβ intracellular mediators. Interestingly, combined TGFβ and dasatinib treatment resulted in a transient increase in p-Smad3 (seen after 3 hours). In addition, when NSCLC cells were treated with this combination, the pro-apoptotic protein BIM was up-regulated. Knockdown of the expression of Smad3 using Smad3 siRNA also resulted in a decrease in BIM protein, suggesting that TGFβ-1 + dasatinib-induced apoptosis is mediated by Smad3 regulation of BIM. Dasatinib is only effective in killing EGFR mutant cells, which is shown in only 10% of NSCLCs. Therefore, the observation that wild-type EGFR lung cancers can be manipulated to render them sensitive to killing by dasatinib could have important implications for devising innovative and potentially more efficacious treatment strategies for this disease