Project description:We report a new platform for the rapid phenotypic selection of protein aggregation inhibitors from genetically encoded cyclic peptide libraries in E. coli based on phage-assisted continuous evolution (PACE). Here, we developed a new PACE-compatible selection for protein aggregation inhibition and employed it to identify cyclic peptides that suppress amyloid-β42 (Aβ42) and human islet amyloid polypeptide (hIAPP) aggregation. Additionally, we integrated a negative selection that removes false positives and off-target hits, significantly improving cyclic peptide selectivity. We show that selected inhibitors are active when chemically re-synthesized in in vitro assays. Our platform provides a powerful new approach for the rapid discovery of cyclic peptide inhibitors of protein aggregation and may serve as the basis for the future evolution of cyclic peptides with a broad spectrum of inhibitory activities. Data deposited here are HTS data critical to the conclusions of this study.
Project description:Here we report the discovery of a set of potent de-novo cyclic peptides (CPs) targeting different binding sites on KDM7B. One CP (OC9) bound directly to the KDM7 PHD-finger, as supported by bio-layer interferometry (BLI), isothermal calorimetry (ITC), hydrogen-deuterium exchange mass spectrometry (HDxMS) and NMR studies, and was highly selective for KDM7s over other PHD-fingers. OC9 disrupted PHD-finger binding to H3K4me3, and allosterically modulated KDM7 demethylase activity at H3K9me2 site on peptides and histone extracts demonstrating PHD-finger targeting is as JmjC-domain targeted inhibitors, but more selective for specific KDM7 subfamily member and certain combinatorial histone PTM signatures. Proteomic analysis confirmed OC9 to selectively target KDM7 in nuclear lysates demonstrating its high affinity and selectivity against other H3K4me3 reader domains and KDMs.
Project description:Dengue and Zika are two mosquito-borne diseases of great concern, affecting mainly the tropical and subtropical regions worldwide. The arrival of Zika virus (ZIKV) in dengue virus (DENV) endemic areas imposed challenges for differential diagnosis and the development of candidate vaccines. The use of peptides has shown great potential to achieve these goals. We aimed to identify the linear epitope profile recognized by the serum samples of dengue and Zika patients in the E and NS1 proteins of DENV and ZIKV to select peptides with the potential for the development of diagnostic tests and vaccines. Analysis of a peptide microarray platform with serum samples of dengue and Zika patients demonstrated that the epitopes were evenly distributed across the entire viral proteins, showing no preference for particular regions. However, several epitopes were within epitope hot spots constituted by clusters of peptides recognized in more than 30% of the sub-arrays analyzed with individual or pools of serum samples. The serum samples of dengue and Zika patients showed a high level of cross-reaction for epitopes in the DENV and ZIKV proteins. Analysis of an additional peptide microarray platform containing selected peptides based on the results of the first screening showed that three peptides (DENV: TQGEPSLNEEQDKRF and TQTVGPWHLGKLEID; ZIKV: LELDPPFGDSYIVIG), highly specific for their cognate viruses (p<0.05), were within the epitope hot spots; however, these peptides showed low detection rates (32.5, 35.0, and 28.6%, respectively). We also found two peptides (DENV: WEVEDYGFGVFTTNI and LELDFDLCEGTTVVV) in the epitope hot spots detected by both dengue and Zika patients with similarly high rates (arbitrary detection rate cut-off threshold of ≥40%). The epitope hot spots harbor several immunodominant epitopes recognized by a higher number of individuals when compared to the 15 aa sequence peptides. Therefore, the entire epitope hot spots, spanning up to ~30 aa, would have more potential than peptides of only 15 aa to serve as antigens in diagnostic tests and vaccine developments.
Project description:Dengue and Zika are two mosquito-borne diseases of great concern, affecting mainly the tropical and subtropical regions worldwide. The arrival of Zika virus (ZIKV) in dengue virus (DENV) endemic areas imposed challenges for differential diagnosis and the development of candidate vaccines. The use of peptides has shown great potential to achieve these goals. We aimed to identify the linear epitope profile recognized by the serum samples of dengue and Zika patients in the E and NS1 proteins of DENV and ZIKV to select peptides with the potential for the development of diagnostic tests and vaccines. Analysis of a peptide microarray platform with serum samples of dengue and Zika patients demonstrated that the epitopes were evenly distributed across the entire viral proteins, showing no preference for particular regions. However, several epitopes were within epitope hot spots constituted by clusters of peptides recognized in more than 30% of the sub-arrays analyzed with individual or pools of serum samples. The serum samples of dengue and Zika patients showed a high level of cross-reaction for epitopes in the DENV and ZIKV proteins. Analysis of an additional peptide microarray platform containing selected peptides based on the results of the first screening showed that three peptides (DENV: TQGEPSLNEEQDKRF and TQTVGPWHLGKLEID; ZIKV: LELDPPFGDSYIVIG), highly specific for their cognate viruses (p<0.05), were within the epitope hot spots; however, these peptides showed low detection rates (32.5, 35.0, and 28.6%, respectively). We also found two peptides (DENV: WEVEDYGFGVFTTNI and LELDFDLCEGTTVVV) in the epitope hot spots detected by both dengue and Zika patients with similarly high rates (arbitrary detection rate cut-off threshold of ≥40%). The epitope hot spots harbor several immunodominant epitopes recognized by a higher number of individuals when compared to the 15 aa sequence peptides. Therefore, the entire epitope hot spots, spanning up to ~30 aa, would have more potential than peptides of only 15 aa to serve as antigens in diagnostic tests and vaccine developments.
Project description:Oxidative damage contributes significantly to the pathogenesis of psoriasis. We recently developed antioxidative peptides (UPF peptides) activating the endogenous glutathione system (GSH). In the present study, we analyzed gene expression profiles in the samples of psoriasis patients to find if these peptides could reduce oxidative damage during psoriasis. Peripheral blood mononuclear cells (PBMCs) from patients with psoriasis and from healthy controls were collected and cultivated. Cultured PBMCs were incubated with two different UPF peptides for 12 hours. Gene expression analysis was performed with Affymetrix Human Gene 1.0 ST arrays and with quantitative real-time PCR. Gene expression profile in the PBMCs of patients with psoriasis indicated significant up-regulation of the immune response pathway. Treatment with UPF peptides normalized the gene expression pattern in psoriasis samples. Therefore, treatment with antioxidative drugs have potential anti-psoriatic activity.
Project description:Oxidative damage contributes significantly to the pathogenesis of psoriasis. We recently developed antioxidative peptides (UPF peptides) activating the endogenous glutathione system (GSH). In the present study, we analyzed gene expression profiles in the samples of psoriasis patients to find if these peptides could reduce oxidative damage during psoriasis. Peripheral blood mononuclear cells (PBMCs) from patients with psoriasis and from healthy controls were collected and cultivated. Cultured PBMCs were incubated with two different UPF peptides for 12 hours. Gene expression analysis was performed with Affymetrix Human Gene 1.0 ST arrays and with quantitative real-time PCR. Gene expression profile in the PBMCs of patients with psoriasis indicated significant up-regulation of the immune response pathway. Treatment with UPF peptides normalized the gene expression pattern in psoriasis samples. Therefore, treatment with antioxidative drugs have potential anti-psoriatic activity. 5 healthy controls (C1-5), 5 psoriasis patients (P1-5), 2 different drugs (upf17 peptide, upf1 peptide) and a control drug. Overall, 30 samples and six groups: PSOR, PSORUPF1, PSORUPF17, CON, CONUPF1, CONUPF17.
Project description:Tumor hypoxia is not a stable phenomenon but cycles between periods of deep hypoxia and reoxygenation. Cyclic hypoxia originates from heterogeneities in red blood cell flux and from the permanent remodelling of the angiogenic vascular network. Endothelial cells lining tumor blood vessels are therefore also influenced by cyclic hypoxia. The gene expression pattern promoted by cyclic hypoxia differs from those observed under normoxia and even continuous hypoxia. PTGS2 is one gene exquisitely up-regulated in endothelial cells (and tumor cells) in response to cyclic hypoxia. Elevated COX-2 (the PTGS2 gene product) expression and activity account for cyclic hypoxia-driven increase in endothelial cell survival and angiogenesis.
Project description:The acidic tumor microenvironment in melanoma drives immune evasion by up-regulating cyclic adenosine monophosphate (cAMP) in tumor-infiltrating monocytes..Melanoma growth can be suppressed by releasing non-toxic concentrations of an adenylate cyclase (AC) inhibitor from polypept(o)id micelles at the tumor site in an immune cell-dependent manner.