Au(I) complexes installed on a self-assembled peptide efficiently catalyze intramolecular cyclization reactions.

Self-assembled peptides are used for diverse applications in the biomedical and technological fields. The morphology and function of the assembled systems are dictated by the peptide sequence and length. In this work, a supramolecular catalyst was obtained upon self-assembly of the diphenylalanine peptide conjugated to a triphenylphosphine Au(I) complex in acetonitrile. The assembled molecules were characterized by spectroscopic techniques and by scanning electron microscopy. The activity of the catalyst was tested on two substrates in cyclization reactions. The morphology and the dimensions of the assembled systems vary depending on the presence of a carboxyl versus an amide C-terminal end. The catalyst efficiently promotes intramolecular cyclization reactions. Results obtained encourage the use of self-assembled peptides for the obtainment of new and efficient catalysts.

Microphthalmia-Associated Transcription Factor: A Differentiation Marker in Uveal Melanoma.

Microphthalmia-associated transcription factor (MITF) is an important regulator of melanogenesis and melanocyte development. In cutaneous melanoma, MITF loss has been linked to an increased expression of stem cell markers, a shift in epithelial-to-mesenchymal transition (EMT)-related factors, and increased inflammation. We explored the role of MITF in Uveal Melanoma (UM) using a cohort of 64 patients enucleated at the Leiden University Medical Center. We analysed the relation between MITF expression and clinical, histopathological and genetic features of UM, as well as survival. We performed differential gene expression and gene set enrichment analysis using mRNA microarray data, comparing MITF-low with MITF-high UM. MITF expression was lower in heavily pigmented UM than in lightly pigmented UM (p = 0.003), which we confirmed by immunohistochemistry. Furthermore, MITF was significantly lower in UM with monosomy 3/BAP1 loss than in those with disomy 3/no BAP1 loss (p < 0.001) and with 8q gain/amplification 8q (p = 0.02). Spearman correlation analysis showed that a low MITF expression was associated with an increase in inflammatory markers, hallmark pathways involved in inflammation, and epithelial-mesenchymal transition. Similar to the situation in cutaneous melanoma, we propose that MITF loss in UM is related to de-differentiation to a less favourable EMT profile and inflammation.

Adding the Cancer Genome Atlas Chromosome Classes to American Joint Committee on Cancer System Offers More Precise Prognostication in Uveal Melanoma.

PURPOSE: Uveal melanoma (UM) is a rare disease and the most common primary intraocular malignancy in adults, with a high risk of metastases. Reliable prognostication systems are based on anatomic features, as in the tumor-node-metastasis staging of the American Joint Committee on Cancer (AJCC) system, or on genetic information, as in The Cancer Genome Atlas (TCGA) system. Prior evidence suggests that combining both systems may be beneficial. We evaluated the benefit of combining the TCGA and AJCC systems in a large cohort of patients. DESIGN: Retrospective case series of patients with UM. PARTICIPANTS: Nine hundred seventy-nine patients with a choroidal or ciliary body melanoma treated at the Wills Eye Hospital between 1998 and 2020, 94% of whom received eye-sparing treatment. METHODS: Tumors were classified into 4 TCGA groups based on chromosome copy number: A (disomy 3, normal 8q), B (disomy 3, any 8q gain), C (monosomy 3, 1 extra copy of 8q), and D (monosomy 3, multiple 8q gain). The eighth edition of the AJCC staging manual was used for AJCC staging. Cox regression and the log-rank test were used for survival analysis. MAIN OUTCOME MEASURE: Metastasis-free survival. RESULTS: Combining information of the 2 systems improved prognostication in intermediate groups: in TCGA group C, we saw an increased rate of metastasis in AJCC stage III (28%) compared with stage II (8.9%); the same was seen in AJCC stage II, going from TCGA group C (8.9%) to group D (46%), and in AJCC stage III, going from group C (28%) to group D (49%). In patients with AJCC stage II or III disease, loss of chromosome 3 and gain of 8q (TCGA groups C and D) significantly worsened the prognosis, with multiple 8q gain (TCGA group D) having a greater impact. CONCLUSIONS: Combining information from AJCC stages and TCGA groups yields a better predictive power even in this set of relatively small tumors. We propose that physicians take both systems into account whenever possible, especially in moderate-risk groups.

Chromosome 3 and 8q Aberrations in Uveal Melanoma Show Greater Impact on Survival in Patients with Light Iris versus Dark Iris Color.

PURPOSE: Individuals with gray, blue, or green eyes have a higher chance of developing uveal melanoma (UM) than those with brown eyes. We wondered whether iris pigmentation might be related not only to predisposition to UM but also to its behavior; therefore, we compared the clinical, histopathologic, and genetic characteristics of UM between eyes with different colors. DESIGN: We determined iris color in a large cohort of patients enucleated for UM. Clinical and histopathologic tumor characteristics, chromosome status, and survival were compared among 3 groups on the basis of iris color. PARTICIPANTS: A total of 412 patients with choroidal/ciliary body UM, who had undergone primary enucleation at the Leiden University Medical Center, Leiden, The Netherlands, between 1993 and 2019, were divided into 3 groups based on iris color: gray/blue, green/hazel, and brown. The validation cohort included 934 patients with choroidal/ciliary body UM treated at Wills Eye Hospital (WEH). METHODS: Comparison of clinical, histopathologic, and genetic characteristics of UM in patients with different iris colors. MAIN OUTCOME MEASURES: Melanoma-related survival in UM patients, divided over 3 iris color groups, in relation to the tumor's chromosome 3 and 8q status. RESULTS: Moderate and heavy tumor pigmentations were especially seen in eyes with a brown iris (P < 0.001). Survival did not differ between patients with different iris colors (P = 0.27); however, in patients with a light iris, copy number changes in chromosome 3 and 8q had a greater influence on survival than in patients with a dark iris. Likewise, chromosome 3 and chromosome 8q status affected survival more among patients with lightly pigmented tumors than in patients with heavily pigmented tumors. The WEH cohort similarly showed a greater influence of chromosome aberrations in light-eyed individuals. CONCLUSIONS: Although iris color by itself did not relate to UM-related survival, chromosome 3 and 8q aberrations had a larger influence on survival in patients with a light iris than those with a brown iris. This suggests a synergistic effect of iris pigmentation and chromosome status in the regulation of oncogenic behavior of UM. Iris color should be taken into consideration when calculating a patient's risk for developing metastases.

MITF in Normal Melanocytes, Cutaneous and Uveal Melanoma: A Delicate Balance.

Microphthalmia-associated transcription factor (MITF) is an important regulator of melanogenesis and melanocyte development. Although it has been studied extensively in cutaneous melanoma, the role of MITF in uveal melanoma (UM) has not been explored in much detail. We review the literature about the role of MITF in normal melanocytes, in cutaneous melanoma, and in UM. In normal melanocytes, MITF regulates melanocyte development, melanin synthesis, and melanocyte survival. The expression profile and the behaviour of MITF-expressing cells suggest that MITF promotes local proliferation and inhibits invasion, inflammation, and epithelial-to-mesenchymal (EMT) transition. Loss of MITF expression leads to increased invasion and inflammation and is more prevalent in malignant cells. Cutaneous melanoma cells switch between MITF-high and MITF-low states in different phases of tumour development. In UM, MITF loss is associated with loss of BAP1 protein expression, which is a marker of poor prognosis. These data indicate a dual role for MITF in benign and malignant melanocytic cells.

Morpholino-based peptide oligomers: Synthesis and DNA binding properties.

The chemical structure of oligonucleotide analogues dictates the conformation of oligonucleotide analogue oligomers, their ability to hybridize complementary DNA and RNA, their stability to degradation and their pharmacokinetic properties. In a study aimed at investigating new analogues featuring a neutral backbone, we explored the ability of oligomers containing a morpholino-peptide backbone to bind oligonucleotides. Circular Dichroism studies revealed the ability of our oligomers to interact with DNA, molecular modelling studies revealed the interaction responsible for complex stabilization.

Exploiting Ultrashort α,ß-Peptides in the Colloidal Stabilization of Gold Nanoparticles.

Colloidal gold nanoparticles (GNPs) have found wide-ranging applications in nanomedicine due to their unique optical properties, ease of preparation, and functionalization. To avoid the formation of GNP aggregates in the physiological environment, molecules such as lipids, polysaccharides, or polymers are employed as GNP coatings. Here, we present the colloidal stabilization of GNPs using ultrashort α,ß-peptides containing the repeating unit of a diaryl ß2,3-amino acid and characterized by an extended conformation. Differently functionalized GNPs have been characterized by ultraviolet, dynamic light scattering, and transmission electron microscopy analysis, allowing us to define the best candidate that inhibits the aggregation of GNPs not only in water but also in mouse serum. In particular, a short tripeptide was found to be able to stabilize GNPs in physiological media over 3 months. This new system has been further capped with albumin, obtaining a material with even more colloidal stability and ability to prevent the formation of a thick protein corona in physiological media.

In vitro angiogenesis inhibition with selective compounds targeting the key glycolytic enzyme PFKFB3.

Abnormal glycolytic metabolism contributes to angiogenic sprouting involved in atherogenesis. We investigated the potential anti-angiogenic properties of specific 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase-3 (PFKFB3) inhibitors in endothelial cells (ECs). ECs were treated with PFKFB3 inhibitors (named PA-1 and PA-2) and their effects on metabolic and functional characteristics of ECs were investigated. The anti-glycolytic compound 3-(pyridinyl)- 1-(4-pyridinyl)- 2-propen-1-one (3PO) was used as reference compound. PFKFB3 expression and activity (IC50 about 3-21 nM) was inhibited upon treatment with both compounds. Glucose uptake and lactate export were measured using commercial assays and showed a partial reduction up to 40%. PFKFB3 inhibition increased intracellular lactate accumulation, and reduced expression of monocarboxylate transporters-1 (MCT1) and MCT4. Furthermore, endothelial cell migration and proliferation assays demonstrated significant reduction upon treatment with both compounds. Matrix- metalloproteinase (MMP) activity, measured by gelatin zymography, and expression was significantly reduced (up to 25%). In addition, PA compounds downregulated the expression of VCAM-1, VE-cadherin, VEGFa, VEGFR2, TGF-ß, and IL-1ß, in inflamed ECs. Finally, PA-1 and PA-2 treatment impaired the formation of angiogenic sprouts measured by both morphogenesis and spheroid-based angiogenesis assays. Our data demonstrate that the anti-glycolytic PA compounds may affect several steps involved in angiogenesis. Targeting the key glycolytic enzyme PFKFB3 might represent an attractive therapeutic strategy to improve the efficacy of cancer treatments, or to be applied in other pathologies where angiogenesis is a detrimental factor.

Efficacy of the DigniCap System in preventing chemotherapy-induced alopecia in breast cancer patients is not related to patient characteristics or side effects of the device.

BACKGROUND: The DigniCap System is an effective scalp cooling device for the prevention of chemotherapy-induced alopecia in early breast cancer patients. AIM: This prospective study was designed to confirm the efficacy and tolerability of the device, to explore potential factors associated with its efficacy and to collect data on patient perceptions and satisfaction. METHODS: Between January 2016 and June 2018, 163 early breast cancer patients eligible for adjuvant chemotherapy were enrolled. Hair loss was assessed using the Dean scale, where a score of 0-2 (hair loss ≤50%) was defined as successful. RESULTS: Hair preservation was successful in 57% of patients in the overall series. The proportion was even higher (81%) in the patient subgroup treated with a paclitaxel and trastuzumab regimen. Side effects (feeling cold, headache, head heaviness, scalp and cervical pain) were mild to moderate and did not correlate with the rate of hair loss. Lifestyle, anthropometric factors and hair characteristics failed to be associated with device efficacy. CONCLUSIONS: The DigniCap System was well tolerated and found to be effective in preventing alopecia in early breast cancer patients. Our study failed to identify factors other than type of chemotherapy regimen associated with hair preservation.

On-resin multicomponent 1,3-dipolar cycloaddition of cyclopentanone-proline enamines and sulfonylazides as an efficient tool for the synthesis of amidino depsipeptide mimics.

Depsipeptides are biologically active peptide derivatives that possess a high therapeutic interest. The development of depsipeptide mimics characterized by a chemical diversity could lead to compounds with enhanced features and activity. In this work, an on-resin multicomponent procedure for the synthesis of amidino depsipeptide mimics is described. This approach exploits a metal-free 1,3-dipolar cycloaddition of cyclopentanone-proline enamines and sulfonylazides. In this reaction, the obtained primary cycloadduct undergoes a ring opening and molecular rearrangement giving access to a linear sulfonyl amidine functionalized with both a peptide chain and a diazoalkane. The so-obtained diazo function "one pot" reacts with the carboxylic group of N-Fmoc-protected amino acids leading to amidino depsipeptide mimics possessing a C4 aliphatic chain. An important advantage of this procedure is the possibility to easily obtain amidino-functionalized derivatives that are proteolytically stable peptide bond bioisosteres. Moreover, the conformational freedom given by the alkyl chain could promote the obtainment of cyclic depsipeptide with a stabilized secondary structure as demonstrated with both in silico calculations and experimental conformational studies. Finally, labeled depsipeptide mimics can be also synthesized using a fluorescent sulfonylazide in the multicomponent reaction.

New Potential Weapons for Refractory Scleritis in the Era of Targeted Therapy.

OBJECTIVE: To assess the efficacy of biologic drugs, beyond tumor necrosis factor- (TNF-) α inhibitors, in the management of noninfectious refractory scleritis, either idiopathic or associated with systemic immune-mediated disorders. Patients and Methods. This is a retrospective study assessing the efficacy of several biologic agents (rituximab, anakinra, tocilizumab, and abatacept) and the small molecule tofacitinib in the treatment of scleritis through assessment of scleral inflammation and relapses, as well as treatment impact on best-corrected visual acuity (BCVA) and safety profile. RESULTS: Fourteen patients (19 eyes) were enrolled in the study. Scleritis inflammatory grading significantly improved from baseline to 3 months (p = 0.002) and from baseline to the last follow-up visit (p = 0.002). Scleritis relapses significantly decreased between the 12 months preceding and following biologic therapy (p = 0.007). No differences regarding BCVA were observed (p = 0.67). Regarding adverse events, only one patient developed pneumonia and septic shock under rituximab treatment. CONCLUSIONS: Our results, though limited to a low number of patients, highlight the effectiveness of different biologic therapies in the treatment of noninfectious refractory scleritis, showing to control scleral inflammation and allowing a significant reduction in the number of relapses.

Rational Design of a User-Friendly Aptamer/Peptide-Based Device for the Detection of Staphylococcus aureus.

The urgent need to develop a detection system for Staphylococcus aureus, one of the most common causes of infection, is prompting research towards novel approaches and devices, with a particular focus on point-of-care analysis. Biosensors are promising systems to achieve this aim. We coupled the selectivity and affinity of aptamers, short nucleic acids sequences able to recognize specific epitopes on bacterial surface, immobilized at high density on a nanostructured zirconium dioxide surface, with the rational design of specifically interacting fluorescent peptides to assemble an easy-to-use detection device. We show that the displacement of fluorescent peptides upon the competitive binding of S. aureus to immobilized aptamers can be detected and quantified through fluorescence loss. This approach could be also applied to the detection of other bacterial species once aptamers interacting with specific antigens will be identified, allowing the development of a platform for easy detection of a pathogen without requiring access to a healthcare environment.

Eight-week interval in flushing and locking port-a-cath in cancer patients: A single-institution experience and systematic review.

Port-a-cath (PAC) system is one of the most frequently employed venous accesses for administration of chemotherapy and supportive care. To prevent late complications, the latest guidelines recommend flushing/locking procedures every four weeks. In this retrospective study, we evaluate the frequencies of late complications with a eight-week flushing/locking procedure compared to the standard one. This study retrospectively compares the frequency of complications occurred using standard versus delayed flushing schedules. We performed a systematic review of the published studies about PAC complications associated with longer flushing intervals. Three hundred and ninety fully available patients were enrolled. One hundred and six patients had their PAC flushed/locked every month, 347 patients performed the flushing/locking procedures every eight weeks, 63 patients switched from the four to the eight-week schedule. No difference was seen in the number of occlusions, infections and mechanical dysfunctions between the two patient groups. The systematic literature review confirmed, in a total of 1,347 patients, the absence of an increased proportion of complications with delayed schedules. PAC flushing and locking every eight weeks are feasible and safe. This delayed schedule may improve patients' quality of life and decrease both nursing workload and costs for the national health system.

Longitudinal quantitative assessment of macula during therapy with fingolimod in relapsing-remitting multiple sclerosis.

PURPOSE: Fingolimod is the first oral drug approved for treatment of relapsing-remitting multiple sclerosis (RR-MS), and it has potential macular side effects. Despite the qualitative evidence of macular oedema under treatment, longitudinal quantitative assessment is lacking. To address this issue, we measured macular volume and central foveal thickness in a cohort of MS patients on fingolimod over 12 months of treatment. METHODS: Central foveal thickness (CFT) and total macular volume (TMV) were longitudinally recorded with spectral-domain optical coherence tomography in a cohort of 23 RR-MS patients treated with fingolimod at baseline, 3, 6 and 12 months. OCT parameters were analysed considering previous history of optic neuritis (ON). Comparison of means was performed with variance analysis (ANOVA). RESULTS: Macular oedema occurred in none of the patients. Comparing both groups of patients (with and without previous ON), no statistically significant difference was found during the follow-up both for CFT and TMV (p = 0.99 and p = 0.96, respectively) although a slight early but not significant TMV reduction was detected. CONCLUSIONS: In our cohort, therapy with fingolimod did not cause any change in CFT and TMV in MS patients during a 12-month follow-up independent of previous ON.

Tetrahydro-4 H-(pyrrolo[3,4- d]isoxazol-3-yl)methanamine: A Bicyclic Diamino Scaffold Stabilizing Parallel Turn Conformations.

A tetrahydro-4 H-(pyrrolo[3,4- d]isoxazol-3-yl)methanamine scaffold was designed as a diamino derivative to stabilize parallel turn conformations. Its synthesis took advantage of a [1,3]-dipolar cycloaddition reaction between the nitrile oxide derived from the inexpensive enantiopure l -phenylalanine and N-benzyl-3-pyrroline. Two diastereoisomers were formed, whose distribution depends on the selected base. 3a R,6a S-Isomer is favored in organic bases, which formation is driven by π-interactions. However, the above interactions were significantly prevented using an inorganic base due to the chaotropic effect of the cation, decreasing the amount of the above isomer. Finally, we demonstrated that this isomer is able to stabilize parallel turn conformations when inserted in short peptide sequences.

Tandem Tetrahydroisoquinoline-4-carboxylic Acid/ß-Alanine as a New Construct Able To Induce a Flexible Turn.

Tetrahydroisoquinoline-4-carboxylic acid, a constrained ß2 -amino acid named ß-TIC, was synthesised for the first time in enantiopure form. The biocatalytic route applied herein represents one of the few successful examples of enzymatic resolution of ß2 -amino acids. Model tetrapeptides, namely, Fmoc-l-Ala-ß-TIC-ß-Ala-l-Val-OBn (Fmoc=fluorenylmethyloxycarbonyl, Bn=benzyl), containing both isomers of ß-TIC, were prepared. Both computational and NMR spectroscopy studies were performed. A reverse-turn conformation was observed in the case of (R)-ß-TIC enantiomer that was obtained in 99 % enantiomeric excess by enzymatic resolution. The ß-TIC/ß-Ala construct represents the first example of a flexible turn mimetic containing a cyclic and an acyclic ß-amino acid. Furthermore, the presence of an aromatic ring of ß-TIC could facilitate non-covalent interactions to increase the potential of this scaffold for the preparation of protein-protein interaction modulators.

Peptide modulators of Rac1/Tiam1 protein-protein interaction: An alternative approach for cardiovascular diseases.

Rac1 GTPase interaction with guanine nucleotide exchange factor Tiam1 is involved in several cancer types and cardiovascular diseases. Although small molecules interfering with their protein-protein interaction (PPI) were identified and studied, the ability of small peptides and peptide mimics acting as Rac1/Tiam1 PPI inhibitors has not been yet explored. Using computational alanine scanning (CAS), the "hot" interfacial residues have been determined allowing the design of a small library of putative PPI inhibitors. In particular, the insertion of an unnatural alpha, alpha disubstituted amino acid, that is norbornane amino acid, and the side chain stapling have been evaluated regarding both conformational stability and biological activity. REMD calculations and CD studies have indicated that one single norbornane amino acid at the N-terminus is not sufficient to stabilize the helix structure, while the side-chain stapling is a more efficient strategy. Furthermore, both engineered peptides have been found able to reduce Rac1-GTP levels in cultured human smooth muscle cells, while wild type sequence is not active.

Self-assembly of an amphipathic ααß-tripeptide into cationic spherical particles for intracellular delivery.

The development of molecular carriers able to carry molecules directly into the cell is an area of intensive research. Cationic nanoparticles are effective delivery systems for several classes of molecules, such as anticancer agents, oligonucleotides and antibodies. Indeed, a cationic charge on the outer surface allows a rapid cellular uptake together with the possibility of carrying negatively charged molecules. In this work, we studied the self-assembly of an ultra-short ααß-tripeptide containing an l-Arg-l-Ala sequence and an unnatural fluorine substituted ß2,3-diaryl-amino acid. The presence of the unnatural ß2,3-diaryl-amino acid allowed us to obtain a protease stable sequence. Furthermore, an arginine guanidinium group triggered the formation of spherical assemblies that were able to load small molecules and enter cells. These spherical architectures, thus, represent interesting candidates for the delivery of exogenous entities directly into cells.

Molecular insights into dimerization inhibition of c-Maf transcription factor.

The Maf protein family belongs to the activator protein 1 (AP-1) superfamily of transcription factors that bind specific DNA target sequences through a basic region and exploit a leucine zipper (LZ) motif for protein-protein interactions leading to homo- or hetero-dimerization. Mafs unique DNA-binding domain contains a highly conserved extended homology region (EHR) that allows to recognize longer DNA sequences than other basic leucine zipper (bZIP) transcription factors. Inspired by the fact that overexpression of Mafs is observed in about 50% of cases of multiple myeloma, a hematological malignant disorder, we undertook a peptide inhibitor approach. The LZ domain of c-Maf, one of large Mafs, was produced by solid phase peptide synthesis. We characterized its secondary structure and dimerization properties, and found that dimerization and folding events are strictly coupled. Moreover, potential peptidic c-Maf dimerization inhibitors were computationally designed and synthesized. These compounds were demonstrated by circular dichroism (CD) spectroscopy and MALDI-TOF mass spectrometry to bind to c-Maf LZ monomers, to drive folding of their partially disordered structure and to efficiently compete with dimerization, suggesting a way for interfering with the function of c-Maf and, more generally, of intrinsically disordered proteins, till now considered undruggable targets.