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Besides its function in normal cellular growth, the molecular chaperone heat shock protein 90 (Hsp90) binds to a large number of client proteins required for promoting cancer cell growth and/or survival. In an effort to discover new small molecules able to inhibit the Hsp90 ATPase and chaperoning activities, we screened, by a surface plasmon resonance assay, a small library including different plant polyphenols. The ellagitannin geraniin, was identified as the most promising molecule, showing a binding affinity to Hsp90α similar to that of 17-(allylamino)-17-demethoxygeldanamycin (17AGG). Geraniin was able to inhibit in vitro the Hsp90α ATPase activity in a dose-dependent manner, with an inhibitory efficiency comparable to that measured for 17-AAG. In addition, this compound compromised the chaperone activity of Hsp90α, monitored by the citrate synthase thermal induced aggregation assay. Geraniin decreased the viability of HeLa and Jurkat cell lines and caused an arrest in G2/M phase. We also proved that following exposure to different concentrations of geraniin, the level of expression of the client proteins c-Raf, pAkt, and EGFR was strongly down-regulated in both the cell lines. These results, along with the finding that geraniin did not exert any appreciable cytotoxicity on normal cells, encourage further studies on this compound as a promising chemical scaffold for the design of new Hsp90 inhibitors.

Annexin A1 (ANXA1, lipocortin-1) is a glucocorticoid-regulated 37-kDa protein, so called since its main property is to bind (i.e. to annex) to cellular membranes in a Ca(2+)-dependent manner. Although ANXA1 has predominantly been studied in the context of immune responses and cancer, the protein can affect a larger variety of biological phenomena, including cell proliferation and migration. Our previous results show that endogenous ANXA1 positively modulates myoblast cell differentiation by promoting migration of satellite cells and, consequently, skeletal muscle differentiation. In this work, we have evaluated the hypothesis that ANXA1 is able to exert effects on myoblast cell migration acting through formyl peptide receptors (FPRs) following changes in its subcellular localization as in other cell types and tissues. The analysis of the subcellular localization of ANXA1 in C2C12 myoblasts during myogenic differentiation showed an interesting increase of extracellular ANXA1 starting from the initial phases of skeletal muscle cell differentiation. The investigation of intracellular Ca(2+) perturbation following exogenous administration of the ANXA1 N-terminal derived peptide Ac2-26 established the engagement of the FPRs which expression in C2C12 cells was assessed by qualitative PCR. Wound healing assay experiments showed that Ac2-26 peptide is able to increase migration of C2C12 skeletal muscle cells and to induce cell surface translocation and secretion of ANXA1. Our results suggest a role for ANXA1 as a highly versatile component in the signaling chains triggered by the proper calcium perturbation that takes place during active migration and differentiation or membrane repair since the protein is strongly redistributed onto the plasma membranes after an rapid increase of intracellular levels of Ca(2+). These properties indicate that ANXA1 may be involved in a novel repair mechanism for skeletal muscle and may have therapeutic implications with respect to the development of ANXA1 mimetics.

Heat shock protein 90 (Hsp90) is a significant target in the development of rational cancer therapy due to its role at the crossroads of multiple signaling pathways associated with cell proliferation and cell viability. The relevance of Hsp90 as a therapeutic target for numerous diseases states has prompted the identification and optimization of novel Hsp90 inhibitors as an emerging therapeutic strategy. We performed a screening aimed to identify novel Hsp90 inhibitors among several natural compounds and we focused on the iminosugar (+)-lentiginosine, a natural amyloglucosidases inhibitor, for its peculiar bioactivity profile. Characterization of Hsp90 inhibition was performed using a panel of chemical and biological approaches, including limited proteolysis, biochemical and cellular assays. Our result suggested that the middle domain of Hsp90, as opposed to its ATP-binding pocket, is a promising binding site for new classes of Hsp90 inhibitors with multi-target anti-cancer potential.

Improved diagnosis of psoriasis, by new biomarkers, is required for evaluating the progression rate of the disease and the response to treatment. Haptoglobin (Hpt), a glycoprotein secreted by hepatocytes and other types of cells including keratinocytes, was found with glycan changes in psoriasis and other diseases. We previously reported that Hpt isolated from plasma of psoriatic patients is more fucosylated than Hpt of healthy subjects. The aim of this study was to compare the glycosylation pattern of Hpt isolated from skin scales or plasma of patients with psoriasis with that of Hpt from cornified epidermal layer or plasma of healthy subjects. High performance liquid chromatography analysis of the glycans isolated from the protein backbone revealed that glycan patterns from skin and plasma of patients were similar, and mostly displayed quantitative rather than qualitative differences from normal pattern. Biotin-labeled lectins were used to evaluate quantitative differences in the glycoforms of Hpt from plasma and psoriatic skin scales. Hpt from skin and plasma of patients showed more fucosylated and branched glycans than Hpt from plasma of healthy subjects. Tryptic glycopeptides of Hpt were also analyzed by mass spectrometry, and a decreased amount of sialylated glycan chains was found in glycopeptides of skin Hpt, as compared with Hpt from plasma. High levels of glycans with fucosylated and tetra-antennary chains were detected on the peptide NLFLNHSENATAK from Hpt of psoriatic patients. Our data demonstrate that specific changes in glycan structures of Hpt, such as enhanced glycan branching and fucose content, are associated with psoriasis, and that differences between circulating and skin Hpt do exist. A lower extent of glycan fucosylation and branching was found in Hpt from plasma of patients in disease remission. Altered glycoforms might reflect changes of Hpt function in the skin, and could be used as markers of the disease.