Quantitative analysis of basic fibroblast growth factor and vascular endothelial growth factor in human colorectal cancer.

Tumour growth is angiogenesis dependent. Some authors suggest a prognostic role of microvessel count in colorectal cancer. We tested the role of basic fibroblast growth factor (bFGF) and vascular endothelial growth factor (VEGF) in the switch to the angiogenic phenotype in 35 patients with colorectal cancer at different stages of disease. We evaluated the two angiogenic factors, by enzyme-linked immunosorbent assay (ELISA), in tumour, peritumoral mucosa, pathological mesenteric and peripheral blood. We used ten endoscopic intestinal biopsies and ten peripheral blood samples from healthy subjects as control. bFGF was significantly lower in tumour tissues and in peritumoral mucosas than in healthy mucosas, whereas VEGF was up-regulated in tumours but not in peritumoral mucosa. Both angiogenic factors were greatly increased in mesenteric blood. VEGF tumour and serum levels were significantly correlated with the stage of disease. bFGF tumour and serum concentration were not correlated with the stage of disease. The high levels of bFGF in mesenteric blood suggest that this growth factor might be abnormally released from tumour tissue and peritumoral mucosa and could function as an early effector in the switch to the angiogenic phenotype. In contrast, VEGF, whose levels show a significant correlation with the stage of disease, could act in a following step, supporting tumour progression

Nitric Oxide-Releasing Nanoparticles Prevent Propionibacterium acnes-Induced Inflammation by Both Clearing the Organism and Inhibiting Microbial Stimulation of the Innate Immune Response.

Propionibacterium acnes induction of IL-1 cytokines through the NLRP3 (NLR, nucleotide oligomerization domain-like receptor) inflammasome was recently highlighted as a dominant etiological factor for acne vulgaris. Therefore, therapeutics targeting both the stimulus and the cascade would be ideal. Nitric oxide (NO), a potent biological messenger, has documented broad-spectrum antimicrobial and immunomodulatory properties. To harness these characteristics to target acne, we used an established nanotechnology capable of generating/releasing NO over time (NO-np). P. acnes was found to be highly sensitive to all concentrations of NO-np tested, although human keratinocyte, monocyte, and embryonic zebra fish assays revealed no cytotoxicity. NO-np significantly suppressed IL-1β, tumor necrosis factor-α (TNF-α), IL-8, and IL-6 from human monocytes, and IL-8 and IL-6 from human keratinocytes, respectively. Importantly, silencing of NLRP3 expression by small interfering RNA did not limit NO-np inhibition of IL-1 β secretion from monocytes, and neither TNF-α nor IL-6 secretion, nor inhibition by NO-np was found to be dependent on this pathway. The observed mechanism by which NO-np impacts IL-1β secretion was through inhibition of caspase-1 and IL-1β gene expression. Together, these data suggest that NO-np can effectively prevent P. acnes-induced inflammation by both clearing the organism and inhibiting microbial stimulation of the innate immune response

Translational control in the stress adaptive response of cancer cells: a novel role for the heat shock protein TRAP1

TNF receptor-associated protein 1 (TRAP1), the main mitochondrial member of the heat shock protein (HSP) 90 family, is induced in most tumor types and is involved in the regulation of proteostasis in the mitochondria of tumor cells through the control of folding and stability of selective proteins, such as Cyclophilin D and Sorcin. Notably, we have recently demonstrated that TRAP1 also interacts with the regulatory protein particle TBP7 in the endoplasmic reticulum (ER), where it is involved in a further extra-mitochondrial quality control of nuclear-encoded mitochondrial proteins through the regulation of their ubiquitination/degradation. Here we show that TRAP1 is involved in the translational control of cancer cells through an attenuation of global protein synthesis, as evidenced by an inverse correlation between TRAP1 expression and ubiquitination/degradation of nascent stress-protective client proteins. This study demonstrates for the first time that TRAP1 is associated with ribosomes and with several translation factors in colon carcinoma cells and, remarkably, is found co-upregulated with some components of the translational apparatus (eIF4A, eIF4E, eEF1A and eEF1G) in human colorectal cancers, with potential new opportunities for therapeutic intervention in humans. Moreover, TRAP1 regulates the rate of protein synthesis through the eIF2α pathway either under basal conditions or under stress, favoring the activation of GCN2 and PERK kinases, with consequent phosphorylation of eIF2α and attenuation of cap-dependent translation. This enhances the synthesis of selective stress-responsive proteins, such as the transcription factor ATF4 and its downstream effectors BiP/Grp78, and the cystine antiporter system xCT, thereby providing protection against ER stress, oxidative damage and nutrient deprivation. Accordingly, TRAP1 silencing sensitizes cells to apoptosis induced by novel antitumoral drugs that inhibit cap-dependent translation, such as ribavirin or 4EGI-1, and reduces the ability of cells to migrate through the pores of transwell filters. These new findings target the TRAP1 network in the development of novel anti-cancer strategies

TRAP1-dependent regulation of p70S6K is involved in the attenuation of protein synthesis and cell migration: Relevance in human colorectal tumors

TNF receptor-associated protein 1 (TRAP1) is an HSP90 chaperone involved in stress protection and apoptosis in mitochondrial and extramitochondrial compartments. Remarkably, aberrant deregulation of TRAP1 function has been observed in several cancer types with potential new opportunities for therapeutic intervention in humans. Although previous studies by our group identified novel roles of TRAP1 in quality control of mitochondria-destined proteins through the attenuation of protein synthesis, molecular mechanisms are still largely unknown. To shed further light on the signaling pathways regulated by TRAP1 in the attenuation of protein synthesis, this study demonstrates that the entire pathway of cap-mediated translation is activated in cells following TRAP1 interference: consistently, expression and consequent phosphorylation of p70S6K and RSK1, two translation activating kinases, are increased upon TRAP1 silencing. Furthermore, we show that these regulatory functions affect the response to translational stress and cell migration in wound healing assays, processes involving both kinases. Notably, the regulatory mechanisms controlled by TRAP1 are conserved in colorectal cancer tissues, since an inverse correlation between TRAP1 and p70S6K expression is found in tumor tissues, thereby supporting the relevant role of TRAP1 translational regulation in vivo. Taken as a whole, these new findings candidate TRAP1 network for new anti-cancer strategies aimed at targeting the translational/quality control machinery of tumor cells

Phosphorylation of mitochondrial proteins in bovine heart Characterization of kinases and substrates

AbstractProtein phosphorylation by [γ-32P]ATP in total extract and subfractions of bovine heart mitochondria has been studied. The results show that, in addition to pyruvate dehydrogenase, three mitochondrial proteins, with molecular weights of 44,000, 39,000 and 31,000 Da, are phosphorylated by a cAMP-independent mitochondrial protein kinase. Three other proteins associated with mitochondria, with molecular weights of 125,000, 19,000 and 6,500 Da, are phosphorylated by the cytoplasmic cAMP-dependent protein kinase (kinase A)

Schedule-dependent activity of 5-fluorouracil and irinotecan combination in the treatment of human colorectal cancer: in vitro evidence and a phase I dose-escalating clinical trial

Several schedules of 5-fluorouracil (FU) and irinotecan (IRI) have been shown to improve overall survival in advanced colorectal cancer (CRC). Preclinical evidence suggests that the sequential administration of IRI and FU produces synergistic activity, although their clinical use has not been fully optimised. We investigated the interaction between short-term exposure to SN-38, the active metabolite of IRI, and prolonged exposure to FU in human CRC HT-29 cells and observed that the synergism of action between the two agents can be increased by extending the time of cell exposure to FU and reducing the interval between administration of the two agents. Based on these findings, we performed a phase I trial in 25 advanced CRC patients using a modified IRI/FU regimen as first-line therapy and evaluated three dose levels of IRI (150–300 mg/m2) and two of continuous infusion of FU (800–1000 mg/m2) in a 3-weekly schedule. The most severe grade III–IV toxicities were neutropoenia in four cycles and diarrhoea in three. One patient achieved complete response (4%), 12 a partial response (48%), the overall response rate was 52% (±20, 95% CI); seven of 25 patients had stable disease (28%), the overall disease control was 80% (±16, 95% CI). This modified IRI/FU schedule is feasible and exhibits potentially interesting clinical activity

What do we have to know about PD-L1 expression in prostate cancer? A systematic literature review. part 4: Experimental treatments in pre-clinical studies (cell lines and mouse models)

In prostate cancer (PC), the PD-1/PD-L1 axis regulates various signaling pathways and it is influenced by extracellular factors. Pre-clinical experimental studies investigating the effects of various treatments (alone or combined) may discover how to overcome the immunotherapyresistance in PC-patients. We performed a systematic literature review (PRISMA guidelines) to delineate the landscape of pre-clinical studies (including cell lines and mouse models) that tested treatments with effects on PD-L1 signaling in PC. NF-kB, MEK, JAK, or STAT inhibitors on human/mouse, primary/metastatic PC-cell lines variably down-modulated PD-L1-expression, reducing chemoresistance and tumor cell migration. If PC-cells were co-cultured with NK, CD8+ Tcells or CAR-T cells, the immune cell cytotoxicity increased when PD-L1 was downregulated (opposite effects for PD-L1 upregulation). In mouse models, radiotherapy, CDK4/6-inhibitors, and RB deletion induced PD-L1-upregulation, causing PC-immune-evasion. Epigenetic drugs may reduce PD-L1 expression. In some PC experimental models, blocking only the PD-1/PD-L1 pathway had limited efficacy in reducing the tumor growth. Anti-tumor effects could be increased by combining the PD-1/PD-L1 blockade with other approaches (inhibitors of tyrosine kinase, PI3K/mTOR or JAK/STAT3 pathways, p300/CBP; anti-RANKL and/or anti-CTLA-4 antibodies; cytokines; nitroxoline; DNA/cell vaccines; radiotherapy/Radium-223)