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Advances, perspectives and innovation in drug delivery have increased in recent years, however, there is limited information available regarding the actual presence of surfactants, nanomedicines and nanocarriers in investigational medicinal products submitted as part of a request for authorization of clinical trials, particularly for those authorized in the European Economic Area. We retrieve, analyze and report data available at the Clinical Trial Office of the Italian Medicines Agency (AIFA), increasing the transparency and availability of relevant information. An analysis of quality documentation submitted along with clinical trials authorized by the AIFA in 2018 was carried out, focusing on the key terms “surfactant”, “nanomedicine” and “nanocarrier”. Results suggest potential indications and inputs for further reflection and actions for regulators to actively and safely drive innovation from a regulatory perspective and to transpose upcoming evolution of clinical trials within a strong regulatory framework.

The disulfide isomerase ERp57 is a soluble protein mainly located in the endoplasmic reticulum, where it acts in the quality control of newly synthesized glycoproteins, in association with calreticulin and calnexin. It has been also detected in other cell compartments, such as the cytosol, the plasma membrane and the nucleus. In these locations it is implicated in various processes, participating in the rapid response to calcitriol, modulating the activity of STAT3 and being requested for the pre-apoptotic exposure of calreticulin on the plasma membrane. In the present work, the involvement of ERp57 in the activity of the EGF receptor was evaluated for the first time. EGFR is a tyrosine kinase receptor, which is able to activate numerous signaling cascades, leading to cell proliferation and inhibition of apoptosis. In the MDA-MB-468 breast adenocarcinoma cells, which overexpress EGFR, ERp57 expression has been knocked down by siRNA and the effects on EGFR have been studied. ERp57 silencing did not affect EGFR protein expression, cell membrane exposure or EGF binding, whereas the internalization and the phosphorylation of the receptor were impaired. The implication of ERp57 in the activity of EGFR, whose upregulation is known to be associated with tumors, could be relevant for cancer therapy.

Calcitriol, the active form of vitamin D3, can regulate the gene expression through the binding to the nuclear receptor VDR, but it can also display nongenomic actions, acting through a membrane-associated receptor, which has been discovered as the disulfide isomerase ERp57. The aim of our research is to identify the binding sites for calcitriol in ERp57 and to analyze their interaction. We first studied the interaction through bioinformatics and fluorimetric analyses. Subsequently, we focused on two protein mutants containing the predicted interaction domains with calcitriol: abb’-ERp57, containing the first three domains, and a’-ERp57, the fourth domain only. To consolidate the achievements we used the calorimetric approach to the whole protein and its mutants. Our results allow us to hypothesize that the interaction with the a’ domain contributes to a greater extent than the other potential binding sites to the dissociation constant, calculated as a Kd of about 10−9 M.

Chromatin immunoprecipitation in M14 melanoma cells showed that the protein ERp57 (endoplasmic reticulum protein 57) binds to DNA in the proximity of STAT3 in a subset of STAT3-regulated genes. In the same cells, IL-6 induced a significant increase of the expression of one of these genes, i.e. CRP. Upon depletion of ERp57 by RNA interference, the phosphorylation of STAT3 on tyrosine 705 was decreased, and the IL-6-induced activation of CRP expression was completely suppressed. In vitro experiments showed that ERp57 is also required for the binding of STAT3 to its consensus sequence on DNA. Thus ERp57, previously shown to associate with STAT3 in the cytosol and in the nuclear STAT3-containing enhanceosome, is a necessary cofactor for the regulation of at least a subset of STAT3-dependent genes, probably intervening both at the site of STAT3 phosphorylation and at the nuclear level.

International audience; The flavonoid silibinin is known to intervene in many cellular processes involved in a variety of pathol-ogies, thus appearing a promising therapeutic tool. The molecular mechanisms responsible for these activities, however, have not been clearly defined, and although some of its interactions with proteins have been identified, the relative affinities are often too low to appear relevant in vivo. Here we describe the interaction of silibinin with the protein disulfide isomerase ERp57, characterized by a submicromolar dissociation constant. This interaction enhances the formation of a ERp57/REF-1 complex, and furthermore appears to affect the intracellular distribution of ERp57. This protein is involved in signaling pathways which are also affected by silibinin. This suggests that the ERp57–silibinin interaction might explain at least some of the biological effects caused by the flavonoid.

ERp57/PDIA3 is a ubiquitously expressed disulfide isomerase protein, which acts in concert with calreticulin and calnexin in the folding of glycoproteins destined to the plasma membrane or to be secreted. Its canonical compartment is the endoplasmic reticulum, where it acts as a chaperone and redox catalyst, but non canonical locations have been described as well, and ERp57 has been found associated with DNA and nuclear proteins. In previous work performed in HeLa cells, ERp57 has been demonstrated to bind specific DNA sequences involved in the stress response. The direct interaction with the DNA sequences identified as ERp57-targeted regions in HeLa cells has now been confirmed in a melanoma cell line. Furthermore, the ERp57 silencing, achieved by RNA interference, has produced a significant down-regulation of the expression of target genes. The possible involvement of other proteins in complex with ERp57 has been studied by an in vitro biotin–streptavidin based binding assay and the interacting protein APE/Ref-1 has been also assessed for its direct association with the ERp57 target regions. In conclusion, nuclear ERp57 interacts in vivo with DNA fragments in melanoma cells and is potentially involved in the transcriptional regulation of its target genes.

The disulfide isomerase ERp57 is a soluble protein mainly located in the endoplasmic reticulum, where it acts in the quality control of newly synthesized glycoproteins, in association with calreticulin and calnexin. It has been also detected in other cell compartments, such as the cytosol, the plasma membrane and the nucleus. In these locations it is implicated in various processes, participating in the rapid response to calcitriol, modulating the activity of STAT3 and being requested for the pre-apoptotic exposure of calreticulin on the plasma membrane. In the present work, the involvement of ERp57 in the activity of the EGF receptor was evaluated for the first time. EGFR is a tyrosine kinase receptor, which is able to activate numerous signaling cascades, leading to cell proliferation and inhibition of apoptosis. In the MDA-MB-468 breast adenocarcinoma cells, which overexpress EGFR, ERp57 expression has been knocked down by siRNA and the effects on EGFR have been studied. ERp57 silencing did not affect EGFR protein expression, cell membrane exposure or EGF binding, whereas the internalization and the phosphorylation of the receptor were impaired. The implication of ERp57 in the activity of EGFR, whose upregulation is known to be associated with tumors, could be relevant for cancer therapy.

The protein ERp57 (also known as PDIA3) is a widely distributed protein, mainly localized in the endoplasmic reticulum, where it acts as disulfide isomerase, oxidoreductase and chaperone, in concert with the lectins calreticulin (CRT) and calnexin. The ERp57/CRT complex has been detected on the cell surface and previous studies have suggested its involvement in programmed cell death. Although the ERp57-CRT complex has been characterized, little is known about its role in different cellular compartments as well as inhibitors of this interaction. We focused on the kinetic, extent and stability of the ERp57-CRT complex, using the surface plasmon resonance spectroscopy, investigating the possible role as inhibitor of the antibiotic vancomycin. Equilibrium thermodynamic data suggested that vancomycin may hinder the interaction between the two proteins and could interfere with the ERp57 conformational changes that stabilize the complex. Furthermore, by means of confocal microscopy, we evaluated the effect of the in vivo administration of vancomycin on the ERp57/CRT complex on the surface of HeLa cells. The model presented here could be used for the search of other specific inhibitors/interactors of ERp57, which can be extremely helpful to understand the biological pathways where the protein is involved and to modulate its activity.

Self-assembled nanohydrogels based on hydrophobically modified polysaccharides have been extensively studied due to their wide potential applications as drug delivery systems. In the present study, we developed and characterized self-assembling nanohydrogels based on sonicated gellan gum chains. Prednisolone (Pred), a poorly water soluble anti-inflammatory drug, was chemically conjugated to the carboxylic groups of gellan (Ge–Pred) and it was the hydrophobic moiety responsible for the self-assembling process. Ge–Pred was characterized by proton nuclear magnetic resonance spectra (1H-NMR) and the cellular cytotoxicity was assessed by the MTS assay. The self-aggregation behavior in aqueous media of Ge–Pred was evaluated by the pyrene fluorescence technique and the nanohydrogels (NHs), prepared by bath sonication in water, were analyzed by dynamic light scattering (DLS) and ζ-potential. The average size of the nanohydrogels was about 300 nm and their ζ-potential values were negative. Our results showed that Ge–Pred NHs are cytocompatible, that the drug is bioavailable and, consequently, they represent an interesting and innovative carrier for prednisolone.

The protein ERp57/GRP58 is a stress-responsive protein and a component of the protein disulfide isomerase family. Its functions in the endoplasmic reticulum are well known, concerning mainly the proper folding and quality control of glycoproteins, and participation in the assembly of the major histocompatibility complex class 1. However, ERp57 is present in many other subcellular locations, where it is involved in a variety of functions, primarily suggested by its participation in complexes with other proteins and even with DNA. While in some instances these roles need to be confirmed by further studies, a great number of observations support the participation of ERp57 in signal transduction from the cell surface, in regulatory processes taking place in the nucleus, and in multimeric protein complexes involved in DNA repair.

In the melanoma M14 cell line, we found that the antimetastatic protein NM23/nucleoside diphosphate kinase binds to the promoters of the oncogene cMYC and of P53, a gene often mutated in human cancer (Cervoni et al. [2006] J. Cell. Biochem. 98:421-428). In a further study, we find now that IFI16, a transcriptional repressor, in both promoters binds to the G-rich fragment that also binds NM23/NDPK. These fragments possess non-B DNA structures. Moreover, by sequential chromatin immunoprecipitation (re-ChIP) we show that the two proteins (IFI16 and NM23/NDPK) are simultaneously bound in vivo to the same DNA fragments. Since P53 stimulates apoptosis and inhibits cellular growth, and cMYC promotes cell growth and, in several instances, also apoptosis, the presence of NM23 and IFI16 on the same DNA fragments suggests their common involvement in the reduced development of some tumors.

The effects of five antibiotics, previously described as ligands of protein disulfide isomerase PDI, have now been studied on the homologous protein ERp57. They bind to this protein with much higher affinity than to PDI, and some of them inhibit the reductase and the DNA-binding activities of ERp57. In view of the high affinity of vancomycin, erythromycin and streptomycin, some effects of their interaction with this protein might be expected in vivo.