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Background A cross-talk between certain oncogenic signaling pathways and PD-L1 expression is emerging. Oncogenic signals often promote cell proliferation, an aggressive cancer hallmark, thus we hypothesized that tumor proliferation rates can influence the levels of PD-L1. Methods We used U251MG and SF767MG glioma cell lines that constitutively express PD-L1 and analyzed the expression level of PD-L1 in condition of serum deprivation and during the phases of cell culture growth. Also, we investigated whether tumor proliferative rates influenced the subcellular distribution of FKBP51s cochaperone, that regulates PD-L1 expression at translational level. Result Our findings suggested that PD-L1 upregulation was associated with DNA synthesis. FKBP51s was detected in ER during DNA replication, but in the nucleus during cell division. A chromatin immunoprecipitation assay using A375-FKBP51 KO melanoma cell line showed that both FKBP51 isoforms can bind the promoter of cyclin D. However, an open chromatin status was found in FKBP51, but FKBP51s enriched chromatin. Conclusion Our study suggests that cell proliferation is an additional factor that drives PD-L1 expression and uncovers an unknown role for its cochaperone that, after assisting PD-L1 production, becomes nuclear in concomitance with the decline of DNA replication and PD-L1 expression.

Melanoma often exploits Treg to avoid immune attack. Treg is a heterogeneous population with respect to immunosuppressive capability. Lymphocytes are particularly rich in FKBP51 (encoded by FKBP5 gene), known as the receptor for FK506. Melanoma aberrantly expresses this protein, which sustains resistance and invasion. Melanoma/immune cell interaction, through PD-L1/PD1, bidirectionally generates FKBP5 splicing inducing a lower molecular weight form termed FKBP51s. In 64 advanced melanoma patient PBMCs, we found that FKBP51s marked a Treg subset that correlated to anti-CTLA4 response. More precisely, a Treg FKBP51s+ count 1.2 and 0.04 and

IL-10 markedly reduces nuclear factor (NF)-kappa B/Rel nuclear activity induced in PBMC by stimulation with the anti-CD3 mAb OKT3. The inhibition is exerted specifically on the NF-kappa B/Rel activation induced by mAb OKT3, and not that produced by PMA. As judged by supershifting the DNA-protein complexes with Abs recognizing specific components of the NF-kappa B/Rel protein family, the p50/p65 (Rel A) heterodimeric form of NF-kappa B is primarily affected. The maximal effect is observed at the IL-10 concentration of 20 U/ml. IL-10 inhibitory activity is exerted on T lymphocytes and is mediated by monocytes. Indeed, monocytes pretreated with IL-10 are able so inhibit NF-kappa B nuclear activity in purified T lymphocytes stimulated with OKT3. Soluble factors do not appear to be involved in the mechanism of inhibition. On the other hand, the up-regulation of CD80 Ag, found on monocytes obtained from PBMC incubated with OKT3, is not detected after addition of IL-10, and the anti-CD28 mAb CLB-CD28/1 restores the NF-kappa B/Rel nuclear activity in IL-10-inhibited lymphocytes. Therefore, the NF-kappa B/Rel inhibition might be ascribed to a lack of cooperation between accessory cells and T lymphocytes, resulting from down-regulation of a costimulatory molecule, such as CD80, produced by IL-10 on activated monocytes. Our results demonstrate that IL-10 can inhibit the induction of NF-kappa B/Rel nuclear activity in CD3-stimulated T lymphocytes. Since inappropriate activation of kappa B-driven genes has a physiopathologic role in a number of diseases, such as HIV infection, our findings support the possibility of using this cytokine to suppress an undesirable activation of these transcription factors.

Recent advances in tumor immunology, fostered by dramatic outcomes with cancer immunotherapy, have opened new scenarios in cancer metastasis. The cancer stemness/mesenchymal phenotype and an excess of immune suppressive signals are emerging as Intertwined aspects of human tumors. This review examines recent studies that explored the mechanistic links between cancer cell stemness and immunoevasion, and the evidence points to these key events in cancer metastasis as two sides of the same coin. This review also covers the mechanisms involved in tumor expression of programmed cell death ligand 1 (PD-L1), a major factor exploited by human neoplasias to suppress immune control. We highlight the convergence of mesenchymal traits and PD-L1 expression and examine the functions of this immune inhibitory molecule, which confers cancer cell resistance and aggressiveness.

In the 70s, after a decade from the purification of cyclosporine, a selective immunosuppressant agent and potent tool in transplantation medicine, a novel molecule was purified from bacteria Streptomyces tsukubaensis. This molecule, called FK506, showed the same selective immunosuppressant action as cyclosporine but was 10 to 100 fold more potent. In an attempt to clarify the molecular mechanism through which the new drug exerted such a selective effect on T-cells activation, two laboratories identified the cytosolic receptor for FK506. This so-called FK506 binding protein (FKBP) was purified from bovine thymus, human spleen, and Jurkat T-cell line. The isolated FKBP had an approximate molecular mass of 14 kDa and showed an isomerase activity similar to the recently purified cyclosporine-binding protein, cyclophilin, but, it was inhibited by FK506 and rapamycin but not cyclosporine. The subsequent cloning of FKBP gene revealed that FKBP and cyclophilin had dissimilar sequences in spite of their common enzymatic activity. The identified FKBP gene encoded for a protein of 108 aminoacids with a relative molecular mass of 11,819. For this reason, the progenitor of this nascent class of proteins was later known as FKBP12. The subsequent studies showed that FKBP12 was just a member of a ubiquitous and evolutionarily conserved sub-family of proteins which differ from each other in their molecular weight and structure. All FKBPs share a highly conserved domain, termed “FK-12 like domain”, capable of binding to FK506 and exerting isomerase properties, i.e. interconversion from cis-to-trans and trans-to-cis of peptide bonds involving proline, on protein substrates. A schematic historical background of the 17 FKBPs so far identified is shown. A general overview of FKBP structure, function and eventually associated disease is given in this monograph, with the order of proteins following the chronology of discovery.

Background: The use of inhibitors of glycoprotein IIb/IIIa (GPIIb/IIIa) has provided dramatic results in terms of the prevention of acute stent thrombosis and a reduction in major adverse coronary events in patients subjected to percutaneous coronary intervention. GPIIb/IIIa or αIIbβ3 is a member of the β3 subfamily of integrins, which also includes αVβ3. GPIIb/IIIa functions as a receptor for fibrinogen and several adhesion proteins sharing an arginine-glycine-aspartic acid (RGD) sequence. GPIIb/IIIa antagonists, through blockade of the receptor, prevent platelet aggregation. Among the three GPIIb/IIIa antagonists used in therapy, abciximab is an anti-β3 monoclonal antibo dy, while tirofiban and eptifibatide mimic the binding sequence of the fibrinogen ligand. Although antiplatelet aggregation represents the central function of GPIIb/IIIa inhibitors, further actions have been documented for these compounds. Objective: The aim of the present article is to review the structures and functions of GPIIb/IIIa antagonists and to highlight the clinical outcomes and results of randomized trials with these compounds. Hypotheses on the unexplored potential of GPIIb/IIIa antagonists will be put forward. Conclusion: GPIIb/IIIa inhibitors were developed to prevent platelet aggregation, however, these compounds can exert further biological functions, both platelet- and non-platelet-related. Large-scale studies comparing the efficacy and safety of GPIIb/IIIa antagonists are lacking. More insights into the functions of these compounds may lead to generation of novel small molecules able to antagonize platelet aggregation while promoting vascular repair.

Mascolo M, Ilardi G, Romano M F, Celetti A, Siano M, Romano S, Luise C, Merolla F, Rocco A, Vecchione M L, De Rosa G & Staibano S (2012) Histopathology Overexpression of chromatin assembly factor-1 p60, poly(ADP-ribose) polymerase 1 and nestin predicts metastasizing behaviour of oral cancer Aims: The natural history of oral squamous cell carcinomas (OSCCs) is variable and difficult to predict. This study aimed to assess the value of the expression of poly(ADP-ribose) polymerase 1 (PARP-1), chromatin assembly factor-1 (CAF-1)/p60 and the stem cell markers CD133, CD166, CD44, CD44v6 and nestin as markers of outcome and progression-free survival in OSCC patients. Methods: Clinical data were collected from 66 patients (41 male and 25 female, aged 29–92 years) who underwent surgery for OSCC of the tongue, floor, lips, and palate. During follow-up (range: 12–131 months), 14 patients experienced relapse/metastasis and/or death. The study was performed by immunohistochemistry on paraffin-embedded tumour tissues, western blot analysis of tumour protein lysates and human cell lines, and RNA silencing assays. In addition, the human papillomavirus (HPV) status of primary tumours was evaluated by immunohistochemistry and viral subtyping. Univariate and multivariate analyses were performed to determine the correlation between these parameters and the clinical and pathological variables of the study population. Results and conclusions: We found that a PARP-1high/CAF-1 p60high/nestinhigh phenotype characterized the OSCCs with the worst prognosis (all HPV-negative). This may be of benefit in clinical management, since radio-enhancing anti-PARP-1 and/or anti-CAF-1/p60 agents may allow radioresistance to be bypassed in the nestin-overexpressing, metastasizing OSCC cells.

Cell death by apoptosis is largely responsible for control of tissue homeostastis, differentiative and immune processes. Among apoptosis regulators are NF-κB/Rel transcription factors. Indeed, overexpression of p65- or c-Rel- containing dimers can impair apoptosis, while inhibition of NF-κ8/Rel activity can enhance death induced by TNF-a, ionizing radiations or chemotherapeutic agents, in several cell types. A number of NF-κB/Rel - regulated genes, that influence cell apoptosis, have been identified; however, a body of evidence indicate that other genes remain to be recognized. Such NF-κB/Rel regulated novel genes are expected to represent innovative tools for both the understanding, monitoring and specific modulation of apoptosis in physiologic and pathologic settings. Our group has been involved in studying NF-κB/Rel activity and apoptosis modulation in normal and pathological cell types, identifying a regulatory role of NF-κB/Rel factors in B-CLL and AML cell apoptotic response to, respectively, fludarabine and cytosine arabinoside, and in human primary CD34+ haematopoietic progenitor cell survival and response to cytoprotectants. Here we comment results from our laboratory concerning NF-κB/Rel anti- apoptotic activity in human primary monocytes

CD40-CD154 interactions play a key role in regulating immune response and are involved in the development of some autoimmune diseases. We analysed the expression of CD154 antigen in CD3-activated PBMC from 10 systemic sclerosis (SSc) patients and 10 control subjects by immunofluorescence. PBMC from SSc patients showed an increased expression of this molecule, since, 6 h following CD3 stimulation, the percentage of CD154(+)cells was of 17. 53+/-2.0 (mean+/-SE) in control and 25.33+/-2.93 in patient cells (P0.03). The higher expression of CD154 antigen was ascribible to CD4(+)cells. The enhanced induction of CD154 following CD3 stimulation depended on protein synthesis, since was abolished when the cells were stimulated via CD3 in the presence of cycloheximide. By analysing the expression of the CD40-induced antigen CD80, we verified that a blocking anti-CD40 antibody inhibited CD80 appearance in SSc activated monocytes, indicating that CD154 molecule was functional. These results show an enhanced expression of a functional CD154 molecule in SSc CD4(+)activated T lymphocytes.

Apoptosis is a form of cell suicide that is essential for the control of cell growth in multicellular organisms. Apoptosis has a very important biological significance in mammals, both during embryonic life and adulthood. Defects in the apoptosis machinery cause several human diseases, including cancer. The induction of apoptosis is now recognized as a major strategy for the treatment and prevention of cancer. Colorectal cancer is a leading cause of cancer death, with the highest prevalence in North America, Europe, Australia, and New Zealand. Aim of the present article is to review the mechanism and biological significance of apoptosis and highlight the efficacy of natural agents that target apoptosis in colo-rectal cancer prevention.