Publications by authors named "Sara Pellegrino"

58 Publications

PET-Based Volumetric Biomarkers for Risk Stratification of Non-Small Cell Lung Cancer Patients.

Diagnostics (Basel) 2021 Jan 30;11(2). Epub 2021 Jan 30.

Department of Advanced Biomedical Sciences, University "Federico II", 80131 Naples, Italy.

Despite the recent advances in lung cancer biology, molecular pathology, and treatment, this malignancy remains the leading cause of cancer-related death worldwide and non-small cell lung cancer (NSCLC) is the most common form found at diagnosis. Accurate staging of the disease is a fundamental prognostic factor that correctly predicts progression-free (PFS) and overall survival (OS) of NSCLC patients. However, outcome of patients within each TNM staging group can change widely highlighting the need to identify additional prognostic biomarkers to better stratify patients on the basis of risk. 18F-FDG PET/CT plays an essential role in staging, evaluation of treatment response, and tumoral target delineation in NSCLC patients. Moreover, a number of studies showed the prognostic role of imaging parameters derived from PET images, such as metabolic tumor volume (MTV) and total lesion glycolysis (TLG). These parameters represent three-dimensional PET-based measurements providing information on both tumor volume and metabolic activity and previous studies reported their ability to predict OS and PFS of NSCLC patients. This review will primarily focus on the studies that showed the prognostic and predictive role of MTV and TLG in NSCLC patients, addressing also their potential utility in the new era of immunotherapy of NSCLC.
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http://dx.doi.org/10.3390/diagnostics11020210DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7911597PMC
January 2021

Alternative Strategy to Obtain Artificial Imine Reductase by Exploiting Vancomycin/D-Ala-D-Ala Interactions with an Iridium Metal Complex.

Inorg Chem 2021 Mar 8;60(5):2976-2982. Epub 2021 Feb 8.

Dipartimento di Scienze Farmaceutiche, Università degli Studi di Milano, Via Venezian 21, 20133 Milano, Italy.

Based on the supramolecular interaction between vancomycin (Van), an antibiotic glycopeptide, and D-Ala-D-Ala (DADA) dipeptides, a novel class of artificial metalloenzymes was synthesized and characterized. The presence of an iridium(III) ligand at the -terminus of DADA allowed the use of the metalloenzyme as a catalyst in the asymmetric transfer hydrogenation of cyclic imines. In particular, the type of link between DADA and the metal-chelating moiety was found to be fundamental for inducing asymmetry in the reaction outcome, as highlighted by both computational studies and catalytic results. Using the [IrCp*()Cl]Cl ⊂ Van complex in 0.1 M CHCOONa buffer at pH 5, a significant 70% () e.e. was obtained in the reduction of quinaldine .
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http://dx.doi.org/10.1021/acs.inorgchem.0c02969DOI Listing
March 2021

Nucleobase morpholino β amino acids as molecular chimeras for the preparation of photoluminescent materials from ribonucleosides.

Sci Rep 2020 11 9;10(1):19331. Epub 2020 Nov 9.

DISFARM-Dipartimento Di Scienze Farmaceutiche, Sezione Chimica Generale E Organica "A. Marchesini", Università Degli Studi Di Milano, Via Venezian 21, 20133, Milan, Italy.

Bioinspired smart materials represent a tremendously growing research field and the obtainment of new building blocks is at the molecular basis of this technology progress. In this work, colloidal materials have been prepared in few steps starting from ribonucleosides. Nucleobase morpholino β-amino acids are the chimera key intermediates allowing Phe-Phe dipeptides' functionalization with adenine and thymine. The obtained compounds self-aggregate showing enhanced photoluminescent features, such as deep blue fluorescence and phosphorescence emissions.
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http://dx.doi.org/10.1038/s41598-020-76297-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7652887PMC
November 2020

Exploring the copper binding ability of Mets7 hCtr-1 protein domain and His7 derivative: An insight in Michael addition catalysis.

J Pept Sci 2021 Feb 22;27(2):e3289. Epub 2020 Oct 22.

Dipartimento di Scienze Farmaceutiche, Università degli Studi di Milano, Milan, Italy.

Mets7 is a methionine-rich motif present in hCtr-1 transporter that is involved in copper cellular trafficking. Its ability to bind Cu(I) was recently exploited to develop metallopeptide catalysts for Henry condensation. Here, the catalytic activity of Mets7-Cu(I) complex in Michael addition reactions has been evaluated. Furthermore, His7 peptide, in which Met residues have been substituted with His ones, was also prepared. This substitution allowed His7 to coordinate Cu (II), with the obtainment of a stable turn conformation as evicted by CD experiments. His7-Cu (II) proved also to be a better catalyst than Mets7-Cu(I) in the addition reaction. In particular, when the substrate was the (E)-1-phenyl-3-(pyridin-2-yl)prop-2-en-1-one, a conversion of 71% and a significative 58% of e.e. was observed.
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http://dx.doi.org/10.1002/psc.3289DOI Listing
February 2021

NoPv1: a synthetic antimicrobial peptide aptamer targeting the causal agents of grapevine downy mildew and potato late blight.

Sci Rep 2020 10 16;10(1):17574. Epub 2020 Oct 16.

Department of Biosciences, University of Milan, Milan, Italy.

Grapevine (Vitis vinifera L.) is a crop of major economic importance. However, grapevine yield is guaranteed by the massive use of pesticides to counteract pathogen infections. Under temperate-humid climate conditions, downy mildew is a primary threat for viticulture. Downy mildew is caused by the biotrophic oomycete Plasmopara viticola Berl. & de Toni, which can attack grapevine green tissues. In lack of treatments and with favourable weather conditions, downy mildew can devastate up to 75% of grape cultivation in one season and weaken newly born shoots, causing serious economic losses. Nevertheless, the repeated and massive use of some fungicides can lead to environmental pollution, negative impact on non-targeted organisms, development of resistance, residual toxicity and can foster human health concerns. In this manuscript, we provide an innovative approach to obtain specific pathogen protection for plants. By using the yeast two-hybrid approach and the P. viticola cellulose synthase 2 (PvCesA2), as target enzyme, we screened a combinatorial 8 amino acid peptide library with the aim to identify interacting peptides, potentially able to inhibit PvCesa2. Here, we demonstrate that the NoPv1 peptide aptamer prevents P. viticola germ tube formation and grapevine leaf infection without affecting the growth of non-target organisms and without being toxic for human cells. Furthermore, NoPv1 is also able to counteract Phytophthora infestans growth, the causal agent of late blight in potato and tomato, possibly as a consequence of the high amino acid sequence similarity between P. viticola and P. infestans cellulose synthase enzymes.
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http://dx.doi.org/10.1038/s41598-020-73027-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7567880PMC
October 2020

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

Sensors (Basel) 2020 Sep 2;20(17). Epub 2020 Sep 2.

Department of Pharmaceutical Sciences, University of Milan, 20133 Milan, Italy.

The urgent need to develop a detection system for , 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 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.
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http://dx.doi.org/10.3390/s20174977DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7506613PMC
September 2020

Brain Metastases Unresponsive to Immunotherapy Detected by 18F-FDG-PET/CT in a Patient with Melanoma.

Diagnostics (Basel) 2020 Jun 17;10(6). Epub 2020 Jun 17.

Department of Advanced Biomedical Sciences, University of Naples Federico II, 80131 Naples, Italy.

Recently, newer therapies such as immunotherapy have been increasingly used in the treatment of several tumors, including advanced melanoma. In particular, several studies showed that the combination of ipilimumab, an anti-Cytotoxic T-lymphocyte Associated Protein 4 (CTLA-4) monoclonal antibody and nivolumab, an anti-Programmed Death 1 (PD-1) monoclonal antibody, leads to improved survival in patients with metastatic melanoma. Despite that, immunotherapeutic agents may not reach therapeutic concentration in the brain due to the blood-brain barrier. We report the case of a 50-year-old man with advanced melanoma who underwent whole-body 18F-FDG-PET/CT before and after treatment with immunotherapy showing resistant brain metastases confirmed by subsequent MRI of the brain. Moreover, 18F-FDG-PET/CT was able to detect an immune-related adverse event such as enterocolitis that contributed to the worsening of patient conditions. This case shows how a whole-body methodology such as 18F-FDG-PET/CT can be useful in identifying melanoma cancer patients unresponsive to immunotherapy that may benefit from traditional palliative therapy in the effort to improve their quality of life.
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http://dx.doi.org/10.3390/diagnostics10060410DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7345060PMC
June 2020

Tuning antiviral CD8 T-cell response via proline-altered peptide ligand vaccination.

PLoS Pathog 2020 05 4;16(5):e1008244. Epub 2020 May 4.

Science for Life Laboratory, Department of Medicine Solna, Karolinska Institute, and Division of Infectious Diseases, Karolinska University Hospital, Solna, Stockholm, Sweden.

Viral escape from CD8+ cytotoxic T lymphocyte responses correlates with disease progression and represents a significant challenge for vaccination. Here, we demonstrate that CD8+ T cell recognition of the naturally occurring MHC-I-restricted LCMV-associated immune escape variant Y4F is restored following vaccination with a proline-altered peptide ligand (APL). The APL increases MHC/peptide (pMHC) complex stability, rigidifies the peptide and facilitates T cell receptor (TCR) recognition through reduced entropy costs. Structural analyses of pMHC complexes before and after TCR binding, combined with biophysical analyses, revealed that although the TCR binds similarly to all complexes, the p3P modification alters the conformations of a very limited amount of specific MHC and peptide residues, facilitating efficient TCR recognition. This approach can be easily introduced in peptides restricted to other MHC alleles, and can be combined with currently available and future vaccination protocols in order to prevent viral immune escape.
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http://dx.doi.org/10.1371/journal.ppat.1008244DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7224568PMC
May 2020

Tuning PFKFB3 Bisphosphatase Activity Through Allosteric Interference.

Sci Rep 2019 12 30;9(1):20333. Epub 2019 Dec 30.

DISFARM- Department of Pharmaceutical sciences, Via Mangiagalli 25, 20133, Milan, Italy.

The human inducible phospho-fructokinase bisphosphatase isoform 3, PFKFB3, is a crucial regulatory node in the cellular metabolism. The enzyme is an important modulator regulating the intracellular fructose-2,6-bisphosphate level. PFKFB3 is a bifunctional enzyme with an exceptionally high kinase to phosphatase ratio around 740:1. Its kinase activity can be directly inhibited by small molecules acting directly on the kinase active site. On the other hand, here we propose an innovative and indirect strategy for the modulation of PFKFB3 activity, achieved through allosteric bisphosphatase activation. A library of small peptides targeting an allosteric site was discovered and synthesized. The binding affinity was evaluated by microscale thermophoresis (MST). Furthermore, a LC-MS/MS analytical method for assessing the bisphosphatase activity of PFKFB3 was developed. The new method was applied for measuring the activation on bisphosphatase activity with the PFKFB3-binding peptides. The molecular mechanical connection between the newly discovered allosteric site to the bisphosphatase activity was also investigated using both experimental and computational methods.
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http://dx.doi.org/10.1038/s41598-019-56708-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6937325PMC
December 2019

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

Amino Acids 2020 Jan 28;52(1):15-24. Epub 2019 Nov 28.

DISFARM-Dipartimento di Scienze Farmaceutiche, Sezione Chimica Generale e Organica "A. Marchesini", Università degli Studi di Milano, Via Venezian 21, 20133, Milan, Italy.

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.
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http://dx.doi.org/10.1007/s00726-019-02805-3DOI Listing
January 2020

Visual and volumetric parameters by 18F-FDG-PET/CT: a head to head comparison for the prediction of outcome in patients with multiple myeloma.

Ann Hematol 2020 Jan 27;99(1):127-135. Epub 2019 Nov 27.

Department of Medicine, Surgery and Dentistry, University of Salerno, Salerno, Italy.

In multiple myeloma (MM) patients, 18F-FDG-PET/CT allows either the detection of disease spread by using visual parameters based on the Italian Myeloma criteria for PET Use (IMPeTUs) or the direct measurement of metabolic tumor burden by volume-based parameters such as metabolic tumor volume (MTV). The purpose is to evaluate the contribution of visual and volumetric parameters in the prediction of progression-free survival (PFS) and overall survival (OS) in MM patients. Forty-seven patients in stage IIIA who had undergone whole-body 18F-FDG-PET/CT were retrospectively evaluated. In each patient, visual parameters were determined and compared with volumetric parameters for PFS and OS prediction after a mean follow-up period of 53 months. Among the visual and volumetric parameters tested, a statistically significant difference was found between maximum standardized uptake value, MTV, total lesion glycolysis, and number of lytic lesions of patients with (n = 26) or without (n = 21) progression (p = 0.0400, p = 0.0065, p = 0.015, and p = 0.0220, respectively) and of dead (n = 24) vs survivors (n = 23) (p = 0.0171, p = 0.0037, p = 0.0060, and p = 0.0270, respectively). At univariate and multivariate analysis, MTV and hemoglobin were predictive of both PFS (p = 0.008) and OS (p = 0.0026). The best MTV discriminative value assessed by receiver operating characteristic curve analysis for predicting both PFS and OS was 39.4 ml. By Kaplan-Meier analysis and log-rank test, PFS and OS were significantly better in patients with MTV ≤ 39.4 ml (p = 0.0004 and p = 0.0001, respectively) as compared with those having an MTV higher than the cutoff. The volume-based parameter MTV determined by 18F-FDG-PET/CT may be used in the prediction of PFS and OS in myeloma patients.
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http://dx.doi.org/10.1007/s00277-019-03852-2DOI Listing
January 2020

Total metabolic tumor volume by 18F-FDG PET/CT for the prediction of outcome in patients with non-small cell lung cancer.

Ann Nucl Med 2019 Dec 14;33(12):937-944. Epub 2019 Oct 14.

Department of Advanced Biomedical Sciences, University "Federico II", Via Pansini 5, Edificio 10, 80131, Naples, Italy.

Objective: Metabolic tumor volume (MTV) and total lesion glycolysis (TLG) are imaging parameters derived from 18F-FDG PET/CT that have been proposed for risk stratification of cancer patients. The aim of our study was to test whether these whole-body volumetric imaging parameters may predict outcome in patients with non-small cell lung cancer (NSCLC).

Methods: Sixty-five patients (45 men, 20 women; mean age ± SD, 65 ± 12 years), with histologically proven NSCLC who had undergone 18F-FDG PET/CT scan before any therapy, were included in the study. Imaging parameters including SUV, SUV, total MTV (MTV) and whole-body TLG (TLG) were determined. Univariate and multivariate analyses of clinical and imaging variables were performed using Cox proportional hazards regression. Survival analysis was performed using Kaplan-Meier method and log-rank tests.

Results: A total of 298 lesions were analyzed including 65 primary tumors, 114 metastatic lymph nodes and 119 distant metastases. MTV and TLG could be determined in 276 lesions. Mean value of MTV was 81.83 ml ± 14.63 ml (SE) whereas mean value of TLG was 459.88 g ± 77.02 g (SE). Univariate analysis showed that, among the variables tested, primary tumor diameter (p = 0.0470), MTV of primary tumor (p = 0.0299), stage (p < 0.0001), treatment (p < 0.0001), MTV (p = 0.0003) and TLG (p = 0.0002) predicted progression-free survival in NSCLC patients, while age (p = 0.0550), MTV of primary tumor (p = 0.0375), stage (p < 0.0001), treatment (p < 0.0001), MTV (p = 0.0001) and TLG (p = 0.0008) predicted overall survival. At multivariate analysis age, TLG and stage were retained in the model for prediction of progression-free survival (p < 0.0001), while age, MTV and stage were retained in the model for prediction of overall survival (p < 0.0001). Survival analysis showed that patients with TLG ≤ 54.7 g had a significantly prolonged progression-free survival as compared to patients with TLG > 54.7 g (p < 0.0001). Moreover, overall survival was significantly better in patients showing a MTV ≤ 9.5 ml as compared to those having MTV > 9.5 ml (p < 0.0001). Similar results were obtained in a subgroup of 43 patients with advanced disease (stages III and IV).

Conclusions: Whole-body PET-based volumetric imaging parameters are able to predict outcome in NSCLC patients.
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http://dx.doi.org/10.1007/s12149-019-01407-zDOI Listing
December 2019

Vancomycin-Iridium (III) Interaction: An Unexplored Route for Enantioselective Imine Reduction.

Molecules 2019 Jul 30;24(15). Epub 2019 Jul 30.

Dipartimento di Scienze Farmaceutiche, Università degli Studi di Milano, Via Venezian 21, 20133 Milano, Italy.

The chiral structure of antibiotic vancomycin (Van) was exploited as an innovative coordination sphere for the preparation of an IrCp* based hybrid catalysts. We found that Van is able to coordinate iridium (Ir(III)) and the complexation was demonstrated by several analytical techniques such as MALDI-TOF, UV, Circular dichroism (CD), Raman IR, and NMR. The hybrid system so obtained was employed in the Asymmetric Transfer Hydrogenation (ATH) of cyclic imines allowing to obtain a valuable 61% () in the asymmetric reduction of quinaldine . The catalytic system exhibited a saturation kinetics with a calculated efficiency of K/K = 0.688 hmM.
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http://dx.doi.org/10.3390/molecules24152771DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6695689PMC
July 2019

The selective disruption of presynaptic JNK2/STX1a interaction reduces NMDA receptor-dependent glutamate release.

Sci Rep 2019 05 9;9(1):7146. Epub 2019 May 9.

Laboratory of Neuronal Cell Signaling, EBRI Rita Levi-Montalcini Foundation, Rome, 00161, Italy.

The neuronal loss caused by excessive glutamate release, or 'excitotoxicity', leads to several pathological conditions, including cerebral ischemia, epilepsy, and neurodegenerative diseases. Over-stimulation of presynaptic N-methyl-D-aspartate (NMDA) receptors is known to trigger and support glutamate spillover, while postsynaptic NMDA receptors are responsible for the subsequent apoptotic cascade. Almost all molecules developed so far are unable to selectively block presynaptic or postsynaptic NMDA receptors, therefore a deeper knowledge about intracellular NMDA pathways is required to design more specific inhibitors. Our previous work showed that presynaptic c-Jun N-terminal kinase 2 (JNK2) specifically regulates NMDA-evoked glutamate release and here we demonstrate that an interaction between Syntaxin-1a and JNK2 is fundamental to this mechanism. Based on this evidence, a new cell permeable peptide (CPP), "JGRi1", has been developed to disrupt the JNK2/STX1a interaction to indirectly, but specifically, inhibit presynaptic NMDA receptor signaling. JGRi1 reduces the NMDA-evoked release of glutamate both in in-vitro and ex-vivo experiments while also being able to widely diffuse throughout brain tissue via intraperitoneal administration. In conclusion, the JNK2/STX1 interaction is involved in presynaptic NMDA-evoked glutamate release and the novel CPP, JGRi1, acts as a pharmacological tool that promotes neuroprotection.
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http://dx.doi.org/10.1038/s41598-019-43709-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6509125PMC
May 2019

Fluoro-Aryl Substituted α,β-Peptides in the Development of Foldameric Antiparallel β-Sheets: A Conformational Study.

Front Chem 2019 2;7:192. Epub 2019 Apr 2.

Department of Pharmaceutical Sciences (DISFARM), University of Milan, Milan, Italy.

α,β-Disteroisomeric foldamers of general formula Boc(Ala-β-2,3Fpg)OMe or Boc(Ala-β-2,3Fpg)OMe were prepared from both enantiomers of H-2-(2-F-Phe)-h-PheGly-OH (named β-Fpg) and alanine. Our peptides show two appealing features for biomedical applications: the presence of fluorine, attractive for non-covalent interactions, and aryl groups, crucial for π-stacking. A conformational study was performed, using IR, NMR and computational studies of diastereoisomeric tetra- and hexapeptides containing the β-amino acid in the - and -stereochemistry, respectively. We found that the stability of peptide conformation is dependent on the stereochemistry of the β-amino acid. Combining -Ala with β-2-Fpg, a stable extended β-strand conformation was obtained. Furthermore, β-2-Fpg containing hexapeptide self-assembles to form antiparallel β-sheet structure stabilized by intermolecular H-bonds and π,π-interactions. These features make peptides containing the β-fluoro amino acid very appealing for the development of bioactive proteolytically stable foldameric β-sheets as modulators of protein-protein interaction (PPI).
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http://dx.doi.org/10.3389/fchem.2019.00192DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6454073PMC
April 2019

Bicyclic Pyrrolidine-Isoxazoline γ Amino Acid: A Constrained Scaffold for Stabilizing α-Turn Conformation in Isolated Peptides.

Front Chem 2019 18;7:133. Epub 2019 Mar 18.

Department of Pharmaceutical Sciences, University of Milan, Milan, Italy.

Unnatural amino acids have tremendously expanded the folding possibilities of peptides and peptide mimics. While α,α-disubstituted and β-amino acids are widely studied, γ-derivatives have been less exploited. Here we report the conformational study on the bicyclic unnatural γ amino acid, 4,5,6,6a-tetrahydro-3a-pyrrolo[3,4-d]isoxazole-3-carboxylic acid . In model peptides, the (+)-(3a6a)-enantiomer is able to stabilize α-turn conformation when associated to glycine, as showed by H-NMR, FT-IR, and circular dichroism experiments, and molecular modeling studies. α-turn is a structural motif occurring in many biologically active protein sites, although its stabilization on isolated peptides is quite uncommon. Our results make the unnatural γ-amino acid of particular interest for the development of bioactive peptidomimetics.
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http://dx.doi.org/10.3389/fchem.2019.00133DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6431668PMC
March 2019

Identification of the first enantiopure Rac1-Tiam1 protein-protein interaction inhibitor and its optimized synthesis phosphine free remote group directed hydroarylation.

Medchemcomm 2019 Feb 26;10(2):310-314. Epub 2018 Dec 26.

Department of Pharmaceutical Sciences , General and Organic Chemistry Section "Alessandro Marchesini" , Via Venezian 21 , 20133 Milano , Italy . Email:

A phospine free hydroarylation reaction applied to norbornene derivatives is described for the first time and was exploited for the regioselective gram scale synthesis of , a known Rac1-Tiam1 PPI inhibitor. Umpolung conversion of the nitro group into free amine allowed the regiocontrol of the key arylation step a long range effect. The effect of in comparison with its enantiomers on Rac1 activation of has been evaluated and (-) has been identified as the first enantiomerically pure inhibitor of Rac1-Tiam1 PPI.
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http://dx.doi.org/10.1039/c8md00477cDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6396393PMC
February 2019

Successive crystal structure snapshots suggest the basis for MHC class I peptide loading and editing by tapasin.

Proc Natl Acad Sci U S A 2019 03 26;116(11):5055-5060. Epub 2019 Feb 26.

Science for Life Laboratory, Department of Medicine Solna, Karolinska Institute, and Division of Infectious Diseases, Karolinska University Hospital, Solna, SE-17176 Stockholm, Sweden;

MHC-I epitope presentation to CD8 T cells is directly dependent on peptide loading and selection during antigen processing. However, the exact molecular bases underlying peptide selection and binding by MHC-I remain largely unknown. Within the peptide-loading complex, the peptide editor tapasin is key to the selection of MHC-I-bound peptides. Here, we have determined an ensemble of crystal structures of MHC-I in complex with the peptide exchange-associated dipeptide GL, as well as the tapasin-associated scoop loop, alone or in combination with candidate epitopes. These results combined with mutation analyses allow us to propose a molecular model underlying MHC-I peptide selection by tapasin. The N termini of bound peptides most probably bind first in the N-terminal and middle region of the MHC-I peptide binding cleft, upon which the peptide C termini are tested for their capacity to dislodge the tapasin scoop loop from the F pocket of the MHC-I cleft. Our results also indicate important differences in peptide selection between different MHC-I alleles.
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http://dx.doi.org/10.1073/pnas.1807656116DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6421438PMC
March 2019

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

J Org Chem 2018 10 18;83(19):11493-11501. Epub 2018 Sep 18.

DISFARM, Sezione di Chimica Generale e Organica "A. Marchesini" , Università degli Studi Milano Via Venezian 21 , Milano 20133 , Italy.

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.
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http://dx.doi.org/10.1021/acs.joc.8b01299DOI Listing
October 2018

Memory T cells specific to citrullinated α-enolase are enriched in the rheumatic joint.

J Autoimmun 2018 08 28;92:47-56. Epub 2018 May 28.

Rheumatology Unit, Department of Medicine Solna, Center for Molecular Medicine, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden. Electronic address:

ACPA-positive rheumatoid arthritis (RA) is associated with distinct HLA-DR alleles and immune responses to many citrullinated self-antigens. Herein we investigated the T cell epitope confined within α-enolase in the context of HLA-DRB1*04:01 and assessed the corresponding CD4 T cells in both the circulation and in the rheumatic joint. Comparative crystallographic analyses were performed for the native and citrullinated α-enolase peptides in complex with HLA-DRB1*04:01. HLA-tetramers assembled with either the native or citrullinated peptide were used for ex vivo and in vitro assessment of α-enolase-specific T cells in peripheral blood, synovial fluid and synovial tissue by flow cytometry. The native and modified peptides take a completely conserved structural conformation within the peptide-binding cleft of HLA-DRB1*04:01. The citrulline residue-327 was located N-terminally, protruding towards TCRs. The frequencies of T cells recognizing native eno were similar in synovial fluid and peripheral blood, while in contrast, the frequency of T cells recognizing cit-eno was significantly elevated in synovial fluid compared to peripheral blood (3.6-fold, p = 0.0150). Additionally, citrulline-specific T cells with a memory phenotype were also significantly increased (1.6-fold, p = 0.0052) in synovial fluid compared to peripheral blood. The native T cell epitope confined within α-enolase does not appear to lead to complete negative selection of cognate CD4 T cells. In RA patient samples, only T cells recognizing the citrullinated version of α-enolase were found at elevated frequencies implicating that neo-antigen formation is critical for breach of tolerance.
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http://dx.doi.org/10.1016/j.jaut.2018.04.004DOI Listing
August 2018

Performance of FDG-PET/CT in solitary pulmonary nodule based on pre-test likelihood of malignancy: results from the ITALIAN retrospective multicenter trial.

Eur J Nucl Med Mol Imaging 2018 10 7;45(11):1898-1907. Epub 2018 May 7.

Struttura Complessa di Medicina Nucleare, Ospedale San Giuseppe Moscati, Avellino, Italy.

Purpose: The aim of this study was to determine the performance of F-FDG-PET/CT in patients with solitary pulmonary nodule (SPN), stratifying the risk according to the likelihood of pulmonary malignancy.

Methods: FDG-PET/CT of 502 patients, stratified for pre-test cancer risk, were retrospectively analyzed. FDG uptake in SPN was assessed by a 4-point scoring system and semiquantitative analysis using the ratio between SUVmax in SPN and SUVmean in mediastinal blood pool (BP) and between SUVmax in SPN and SUVmean in liver (L). Histopathology and/or follow-up data were used as standard of reference.

Results: SPN was malignant in 180 (36%) patients, benign in 175 (35%), and indeterminate in 147 (29%). The 355 patients with a definitive SPN nature (malignant or benign) were considered for the analysis. Considering FDG uptake ≥ 2, sensitivity, specificity, positive (PPV) and negative (NPV) predictive values, and accuracy were 85.6%, 85.7%, 86%, 85.2%, and 85.6% respectively. Sensitivity and PPV were higher (P < 0.05) in intermediate and high-risk patients, while specificity and NPV were higher (P < 0.05) in low-risk patients. On receiver operating characteristic curve analysis, the cut-offs for better discrimination between benign and malignant SPN were 1.56 (sensitivity 81% and specificity 87%) and 1.12 (sensitivity 81% and specificity 86%) for SUVmax/SUVmeanBP and SUVmax/SUVmeanL respectively. In intermediate and high-risk patients, including the SUVmax/SUVmeanBP, the specificity shifted from 85% and 50% to 100%.

Conclusion: Visual FDG-PET/CT has an acceptable performance in patients with SPN, but accuracy improves when SUVratios are considered, particularly in patients with intermediate and high risk of malignancy.
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http://dx.doi.org/10.1007/s00259-018-4016-1DOI Listing
October 2018

2-deoxy-2-[18F]fluoro-D-glucose positron emission tomography/computed tomography in primary extranodal lymphomas: treatment response evaluation and prognosis.

Q J Nucl Med Mol Imaging 2020 Jun 24;64(2):219-225. Epub 2018 Apr 24.

Scuola Medica Salernitana Department of Medicine, Surgery, and Dentistry, University of Salerno, Salerno, Italy.

Background: We evaluated the role of [18F]FDG PET/CT in tumor response assessment and prognosis of primary extranodal lymphoma (PEL) patients.

Methods: We examined retrospectively, 56 PEL patients: 31 with aggressive diffuse large B cell lymphoma (DLBCL) and 25 with indolent lymphoma (20 mucosa-associated lymphoid tissue lymphoma and five follicular lymphoma). All patients had undergone [18F]FDG PET/CT at diagnosis (PET-I) and 50 of them also after therapy (PET-II). Moreover, 52 patients were subjected to a mean follow-up period of 76 months.

Results: PET-I was positive in 50 (89%) patients (mean SUVmax 10.3±6.7). In the assessment of tumor response, according to Lugano classification, 45 patients showed complete metabolic response (CMR), four patients had partial metabolic response (PMR) and one had progressive metabolic disease (PMD). Based on 66% ΔSUVmax cut-off, among CMR patients, 41 showed a ΔSUVmax>66% whereas among non-responders, four patients showed a ΔSUVmax<66%. At follow-up, univariate analysis showed that age, performance status, prognostic index, ΔSUVmax and Lugano classification predicted progression-free survival (PFS) (P<0.05), while, performance status, prognostic index, ΔSUVmax and Lugano classification predicted overall survival (OS) (P<0.05). At multivariate analysis only Lugano classification was retained in the model for prediction of both PFS (P<0.05) and OS (P<0.05). By Kaplan-Meier analysis and log-rank testing both PFS and OS were significantly better in patients in CMR as compared to patients in PMR or PMD according to Lugano classification (P<0.01).

Conclusions: [18F]FDG PET/CT represents a useful tool in the detection of disease response and in the evaluation of outcome in PEL patients.
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http://dx.doi.org/10.23736/S1824-4785.18.03043-1DOI Listing
June 2020

The Immunogenicity of a Proline-Substituted Altered Peptide Ligand toward the Cancer-Associated TEIPP Neoepitope Trh4 Is Unrelated to Complex Stability.

J Immunol 2018 04 5;200(8):2860-2868. Epub 2018 Mar 5.

Science for Life Laboratory, Department of Medicine Solna, Karolinska Institutet, 17165 Stockholm, Sweden;

Human cancers frequently display defects in Ag processing and presentation allowing for immune evasion, and they therefore constitute a significant challenge for T cell-based immunotherapy. We have previously demonstrated that the antigenicity of tumor-associated Ags can be significantly enhanced through unconventional residue modifications as a novel tool for MHC class I (MHC-I)-based immunotherapy approaches. We have also previously identified a novel category of cancer neo-epitopes, that is, T cell epitopes associated with impaired peptide processing (TEIPP), that are selectively presented by MHC-I on cells lacking the peptide transporter TAP. In this study, we demonstrate that substitution of the nonanchoring position 3 into a proline residue of the first identified TEIPP peptide, the murine Trh4, results in significantly enhanced recognition by antitumor CTLs toward the wild-type epitope. Although higher immunogenicity has in most cases been associated with increased MHC/peptide complex stability, our results demonstrate that the overall stability of H-2D in complex with the highly immunogenic altered peptide ligand Trh4-p3P is significantly reduced compared with wild-type H-2D/Trh4. Comparison of the crystal structures of the H-2D/Trh4-p3P and H-2D/Trh4 complexes revealed that the conformation of the nonconventional methionine anchor residue p5M is altered, deleting its capacity to form adequate sulfur-π interactions with H-2D residues, thus reducing the overall longevity of the complex. Collectively, our results indicate that vaccination with Thr4-p3P significantly enhances T cell recognition of targets presenting the wild-type TEIPP epitope and that higher immunogenicity is not necessarily directly related to MHC/peptide complex stability, opening for the possibility to design novel peptide vaccines with reduced MHC/peptide complex stability.
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http://dx.doi.org/10.4049/jimmunol.1700228DOI Listing
April 2018

Crystal structures of H-2Db in complex with the LCMV-derived peptides GP92 and GP392 explain pleiotropic effects of glycosylation on antigen presentation and immunogenicity.

PLoS One 2017 18;12(12):e0189584. Epub 2017 Dec 18.

Science for Life Laboratory, Department of Medicine Solna, Karolinska Institutet, and Department of Infectious Diseases, Karolinska University Hospital, Solna, Stockholm, Sweden.

Post-translational modifications significantly broaden the epitope repertoire for major histocompatibility class I complexes (MHC-I) and may allow viruses to escape immune recognition. Lymphocytic choriomeningitis virus (LCMV) infection of H-2b mice generates CD8+ CTL responses directed towards several MHC-I-restricted epitopes including the peptides GP92 (CSANNSHHYI) and GP392 (WLVTNGSYL), both with a N-glycosylation site. Interestingly, glycosylation has different effects on the immunogenicity and association capacity of these two epitopes to H-2Db. To assess the structural bases underlying these functional results, we determined the crystal structures of H-2Db in complex with GP92 (CSANNSHHYI) and GP392 (WLVTNGSYL) to 2.4 and 2.5 Å resolution, respectively. The structures reveal that while glycosylation of GP392 most probably impairs binding, the glycosylation of the asparagine residue in GP92, which protrudes towards the solvent, possibly allows for immune escape and/or forms a neo-epitope that may select for a different set of CD8 T cells. Altogether, the presented results provide a structural platform underlying the effects of post-translational modifications on epitope binding and/or immunogenicity, resulting in viral immune escape.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0189584PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5734757PMC
January 2018

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

Biopolymers 2017 Nov 27. Epub 2017 Nov 27.

Department of Pharmaceutical Sciences, University of Milano, Milano, Italy.

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.
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http://dx.doi.org/10.1002/bip.23089DOI Listing
November 2017

Fruit and vegetable intake among participants in a District of Columbia farmers' market incentive programme.

Public Health Nutr 2018 02 6;21(3):601-606. Epub 2017 Nov 6.

1Department of Exercise and Nutrition Sciences,Milken Institute School of Public Health,George Washington University,950 New Hampshire Avenue,NW,2nd Floor,Washington,DC 20052,USA.

Objective: Limited research is available on whether participation in healthy food incentive programmes is associated with higher fruit and vegetable intake. The objective of the present study was to determine fruit and vegetable intake among participants in the Produce Plus Program, a farmers' market-based healthy food incentive programme in Washington, DC, and identify demographic and behavioural factors associated with higher fruit and vegetable intake.

Design: Using a cross-sectional survey, programme participants were interviewed at markets across DC between June and September 2015. Questions included the Behavioral Risk Factor Surveillance System (BRFSS) fruit and vegetable module. Fruit and vegetable intake among 2013 DC BRFSS participants reporting annual household incomes of ≤$US 35 000 was calculated for context.

Setting: Washington, DC, USA.

Subjects: Participants (n 288) in the Produce Plus Program.

Results: On average, participants reported consuming both fruits (interquartile range: 1·0-3·0) and vegetables (interquartile range: 1·3-3·5) two times/d. Participants who reported eating home-cooked meals ≥3 times/week also reported higher median fruit (2·0 v. 0·8) and vegetable (2·3 v. 1·3) intake compared with those eating home-cooked meals less frequently. No statistically significant differences in reported median fruit or vegetable intake were observed over the course of the farmers' market (June v. August/September) season.

Conclusions: Produce Plus Program participants reported higher median fruit and vegetable intake compared with DC BRFSS respondents with similar incomes, but still below recommended levels. More frequent home-cooked meals were associated with higher fruit and vegetable intake. Thus, efforts to increase home cooking may represent an opportunity to increase fruit and vegetable intake among healthy food incentive participants.
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http://dx.doi.org/10.1017/S1368980017003020DOI Listing
February 2018

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

Org Biomol Chem 2017 Aug;15(32):6773-6779

University of Milano, Department of Pharmaceutical Sciences, Milano, Italy.

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 ααβ-tripeptide containing an l-Arg-l-Ala sequence and an unnatural fluorine substituted β-diaryl-amino acid. The presence of the unnatural β-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.
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http://dx.doi.org/10.1039/c7ob01693jDOI Listing
August 2017

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

Chemistry 2017 Aug 18;23(45):10822-10831. Epub 2017 Jul 18.

DISFARM, Sezione di Chimica Generale e Organica "A. Marchesini", Università degli Studi Milano, Via Venezian 21, 20133, Milano, Italy.

Tetrahydroisoquinoline-4-carboxylic acid, a constrained β -amino acid named β-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 β -amino acids. Model tetrapeptides, namely, Fmoc-l-Ala-β-TIC-β-Ala-l-Val-OBn (Fmoc=fluorenylmethyloxycarbonyl, Bn=benzyl), containing both isomers of β-TIC, were prepared. Both computational and NMR spectroscopy studies were performed. A reverse-turn conformation was observed in the case of (R)-β-TIC enantiomer that was obtained in 99 % enantiomeric excess by enzymatic resolution. The β-TIC/β-Ala construct represents the first example of a flexible turn mimetic containing a cyclic and an acyclic β-amino acid. Furthermore, the presence of an aromatic ring of β-TIC could facilitate non-covalent interactions to increase the potential of this scaffold for the preparation of protein-protein interaction modulators.
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http://dx.doi.org/10.1002/chem.201701045DOI Listing
August 2017

Evaluation of metabolic response with F-FDG PET-CT in patients with advanced or recurrent thymic epithelial tumors.

Cancer Imaging 2017 Mar 7;17(1):10. Epub 2017 Mar 7.

Department of Advanced Biomedical Sciences, University of Naples Federico II, Via Pansini 5, Edificio 10, 80131, Naples, Italy.

Background: Patients with advanced or recurrent thymic epithelial tumors (TETs) often need several consecutive lines of chemotherapy. The aim of this retrospective monocentric study was to test whether F-Fluorodeoxyglucose positron emission tomography-computed tomography (F-FDG PET-CT) is able to monitor standard chemotherapy efficacy in those patients and whether metabolic response correlates with morphovolumetric response as assessed by Response Evaluation Criteria in Solid Tumor (RECIST).

Methods: We evaluated 27 consecutive patients with advanced (16 patients) or recurrent (11 patients) TETs. All patients underwent F-FDG PET-CT before and after at least 3 cycles of chemotherapy. Maximum standardized uptake value (SUV) of all detected lesions was recorded and the most F-FDG avid lesion in each patient was selected for determination of percentage change of SUV (ΔSUV) in pre- and post-treatment scans. Tumor response was assessed by contrast-enhanced computed tomography (CE-CT) using RECIST criteria. Receiver operating characteristic (ROC) curve analysis was performed to define the optimal threshold of ΔSUV discriminating responders from non-responders.

Results: Metabolic response expressed as ΔSUV was significantly correlated with morphovolumetric response (Spearman's rank correlation, r = 0.64, p = 0.001). ROC curve analysis showed that a ΔSUV value of -25% could discriminate responders from non-responders with a sensitivity of 88% and a specificity of 80%. Conversely, basal SUV values were not predictive of morphovolumetric tumor response.

Conclusions: Our findings indicate that metabolic response assessed by F-FDG PET-CT, through evaluation of ΔSUV, may allow identification of responders and non-responders thus guiding adaptation of therapy in patients with advanced or recurrent TETs.
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http://dx.doi.org/10.1186/s40644-017-0112-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5339950PMC
March 2017

Development of poly(lactide-co-glycolide) nanoparticles functionalized with a mitochondria penetrating peptide.

J Pept Sci 2017 Feb 17;23(2):182-188. Epub 2017 Jan 17.

Department of Pharmaceutical Sciences, Università degli Studi di Milano, via Venezian, 21, 20133, Milano, Italy.

The development of mitochondria-targeting cell permeable vectors represents a promising therapeutic approach for several diseases, such as cancer and oxidative pathologies. Nevertheless, access to mitochondria can be difficult. A new hybrid material composed by poly(lactide-co-glycolide) (PLGA) functionalized with a 6-mer mitochondria penetrating peptide (MPP), consisting in alternating arginine and unnatural cyclohexylalanine, was developed. Circular dichroism, FT-IR and DSC studies indicated that the conjugation of the peptide with the polymer led to the obtainment of a more rigid material with respect to both PLGA and MPP as such. In particular, a conformational rearrangement to a helical structure was observed for MPP. MPP-PLGA conjugates were used for the preparation of nanoparticles that showed no cytotoxicity in MTT assay, suggesting their putative use for future studies on mitochondria targeting. Copyright © 2017 European Peptide Society and John Wiley & Sons, Ltd.
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http://dx.doi.org/10.1002/psc.2952DOI Listing
February 2017