bims-cepepe 2024-11-24 papers

bims-cepepe

Biomed News

on Cell-penetrating peptides

Issue of 2024–11–24
fourteen papers selected by
Henry Lamb, Queensland University of Technology

De Novo Discovery of a Noncovalent Cell-Penetrating Bicyclic Peptide Inhibitor Targeting SARS-CoV-2 Main Protease.
Cyclic peptides targeting the SARS-CoV-2 programmed ribosomal frameshifting RNA from a multiplexed phage display library.
Rapid prediction of key residues for foldability by machine learning model enables the design of highly functional libraries with hyperstable constrained peptide scaffolds.
Identification of Covalent Cyclic Peptide Inhibitors Targeting Protein-Protein Interactions Using Phage Display.
Molecular dynamics simulations unveil the aggregation patterns and salting out of polyarginines at zwitterionic POPC bilayers in solutions of various ionic strengths.
Cyclic Peptide C5aR1 Antagonist Design Using Solution Conformational Analysis Derived from Residual Dipolar Couplings.
Exploring structure-directed immunogenic cytotoxicity of arginine-rich peptides for cytolysis-induced immunotherapy of cancer.
A Cyclic Peptide Targets Glioblastoma by Binding to Aberrantly Exposed SNAP25.
Rational engineering of an antimalarial peptide with enhanced proteolytic stability and preserved parasite invasion inhibitory activity.
Structural analysis of an Asterias rubens peptide indicates the presence of a disulfide-directed β-hairpin fold.
A quantitative intracellular peptide binding assay reveals recognition determinants and context dependence of short linear motifs.
Recent advances in peptide macrocyclization strategies.
Hybrids of Membrane-Translocating Antimicrobial Peptides Show Enhanced Activity through Membrane Permeabilization.
Advancements in cell-penetrating monoclonal antibody treatment.

J Med Chem. 2024 Nov 18.

De Novo Discovery of a Noncovalent Cell-Penetrating Bicyclic Peptide Inhibitor Targeting SARS-CoV-2 Main Protease.

Yahong Tan, Jinyue Yang, Min Wang, Qi Peng, Yongqi Li, Lifeng Fu, Mengmeng Zhang, Jiang Wu, Guanya Yang, Christopher John Hipolito, Youming Zhang, Jianxun Qi, Yi Shi, Yizhen Yin.

Macrocyclic peptides have garnered significant attention as promising drug candidates. However, they typically face challenges in achieving and enhancing cell permeability for access to intracellular targets. In this study, we focused on the de novo screening of macrocyclic peptide inhibitors against the main protease (Mpro) of SARS-CoV-2 and identified novel noncovalently bound macrocyclic peptides that effectively inhibit proteolytic activity. High-resolution crystal structures further revealed molecular interactions between the macrocyclic peptides and Mpro. Subsequently, a specific macrocyclic peptide lacking cell permeability was further optimized and transformed into a low-toxicity, metabolically stable bicyclic peptide with a cell penetration capacity and therapeutic potential against SARS-CoV-2. The bicyclic peptide was achieved using a novel strategy that involved introducing both a bicyclic structure and a bridging perfluorobiphenyl group. Our study not only provides a lead peptide inhibitor for COVID-19 but also offers valuable insights into achieving cell penetration for macrocyclic peptides through strategic modifications.

DOI: https://doi.org/10.1021/acs.jmedchem.4c01639
Chem Sci. 2024 Oct 17.

Cyclic peptides targeting the SARS-CoV-2 programmed ribosomal frameshifting RNA from a multiplexed phage display library.

Jacob A Iannuzzelli, Rachel Bonn, Andrew S Hong, Abhijith Saseendran Anitha, Jermaine L Jenkins, Joseph E Wedekind, Rudi Fasan.

RNA provides the genetic blueprint for many pathogenic viruses, including SARS-CoV-2. The propensity of RNA to fold into specific tertiary structures enables the biomolecular recognition of cavities and crevices suited for the binding of drug-like molecules. Despite increasing interest in RNA as a target for chemical biology and therapeutic applications, the development of molecules that recognize RNA with high affinity and specificity represents a significant challenge. Here, we report a strategy for the discovery and selection of RNA-targeted macrocyclic peptides derived from combinatorial libraries of peptide macrocycles displayed by bacteriophages. Specifically, a platform for phage display of macrocyclic organo-peptide hybrids (MOrPH-PhD) was combined with a diverse set of non-canonical amino acid-based cyclization modules to produce large libraries of 107 structurally diverse, genetically encoded peptide macrocycles. These libraries were panned against the -1 programmed ribosomal frameshifting stimulatory sequence (FSS) RNA pseudoknot of SARS-CoV-2, which revealed specific macrocyclic peptide sequences that bind this essential motif with high affinity and selectivity. Peptide binding localizes to the FSS dimerization loop based on chemical modification analysis and binding assays and the cyclic peptides show specificity toward the target RNA over unrelated RNA pseudoknots. This work introduces a novel system for the generation and high-throughput screening of topologically diverse cyclopeptide scaffolds (multiplexed MOrPH-PhD), and it provides a blueprint for the exploration and evolution of genetically encoded macrocyclic peptides that target specific RNAs.

DOI: https://doi.org/10.1039/d4sc04026k
PLoS Comput Biol. 2024 Nov 18. 20(11): e1012609

Rapid prediction of key residues for foldability by machine learning model enables the design of highly functional libraries with hyperstable constrained peptide scaffolds.

Fei Cai, Yuehua Wei, Daniel Kirchhofer, Andrew Chang, Yingnan Zhang.

Peptides are an emerging modality for developing therapeutics that can either agonize or antagonize cellular pathways associated with disease, yet peptides often suffer from poor chemical and physical stability, which limits their potential. However, naturally occurring disulfide-constrained peptides (DCPs) and de novo designed Hyperstable Constrained Peptides (HCPs) exhibiting highly stable and drug-like scaffolds, making them attractive therapeutic modalities. Previously, we established a robust platform for discovering peptide therapeutics by utilizing multiple DCPs as scaffolds. However, we realized that those libraries could be further improved by considering the foldability of peptide scaffolds for library design. We hypothesized that specific sequence patterns within the peptide scaffolds played a crucial role in spontaneous folding into a stable topology, and thus, these sequences should not be subject to randomization in the original library design. Therefore, we developed a method for designing highly diverse DCP libraries while preserving the inherent foldability of each scaffold. To achieve this, we first generated a large-scale dataset from yeast surface display (YSD) combined with shotgun alanine scan experiments to train a machine-learning (ML) model based on techniques used for natural language understanding. Then we validated the ML model with experiments, showing that it is able to not only predict the foldability of peptides with high accuracy across a broad range of sequences but also pinpoint residues critical for foldability. Using the insights gained from the alanine scanning experiment as well as prediction model, we designed a new peptide library based on a de novo-designed HCP, which was optimized for enhanced folding efficiency. Subsequent panning trials using this library yielded promising hits having good folding properties. In summary, this work advances peptide or small protein domain library design practices. These findings could pave the way for the efficient development of peptide-based therapeutics in the future.

DOI: https://doi.org/10.1371/journal.pcbi.1012609
bioRxiv. 2024 Nov 09. pii: 2024.11.08.622749. [Epub ahead of print]

Identification of Covalent Cyclic Peptide Inhibitors Targeting Protein-Protein Interactions Using Phage Display.

Sijie Wang, Franco F Faucher, Matilde Bertolini, Heeyoung Kim, Bingchen Yu, Li Cao, Katharina Roeltgen, Scott Lovell, Varun Shanker, Scott D Boyd, Lei Wang, Ralf Bartenschlager, Matthew Bogyo.

Peptide macrocycles are promising therapeutics for a variety of disease indications due to their overall metabolic stability and potential to make highly selective binding interactions with targets. Recent advances in covalent macrocycle peptide discovery, driven by phage and mRNA display methods, have enabled the rapid identification of highly potent and selective molecules from large libraires of diverse macrocycles. However, there are currently limited examples of macrocycles that can be used to disrupt protein-protein interactions and even fewer examples that function by formation of a covalent bond to a target protein. In this work, we describe a directed counter-selection method that enables identification of covalent macrocyclic ligands targeting a protein-protein interaction using a phage display screening platform. This method utilizes binary and ternary screenings of a chemically modified phage display library, employing the stable and weakly reactive aryl fluorosulfate electrophile. We demonstrate the utility of this approach using the SARS-CoV-2 Spike-ACE2 protein-protein interaction and identify multiple covalent macrocyclic inhibitors that disrupt this interaction. The resulting compounds displayed antiviral activity against live virus that was irreversible after washout due to the covalent binding mechanism. These results highlight the potential of this screening platform for developing covalent macrocyclic drugs that disrupt protein-protein interactions with long lasting effects.

DOI: https://doi.org/10.1101/2024.11.08.622749
Comput Struct Biotechnol J. 2024 Dec;23 3897-3905

Molecular dynamics simulations unveil the aggregation patterns and salting out of polyarginines at zwitterionic POPC bilayers in solutions of various ionic strengths.

Man Thi Hong Nguyen, Mario Vazdar.

  This study employs molecular dynamics (MD) simulations to investigate the adsorption and aggregation behavior of simple polyarginine cell-penetrating peptides (CPPs), specifically modeled as R9 peptides, at zwitterionic phosphocholine POPC membranes under varying ionic strengths of two peptide concentrations and two concentrations of NaCl and CaCl2. The results reveal an intriguing phenomenon of R9 aggregation at the membrane, which is dependent on the ionic strength, indicating a salting-out effect. As the peptide concentration and ionic strength increase, peptide aggregation also increases, with aggregate lifetimes and sizes showing a corresponding rise, accompanied by the total decrease of adsorbed peptides at the membrane surface. Notably, in high ionic strength environments, large R9 aggregates, such as octamers, are also observed occasionally. The salting-out, typically uncommon for short positively charged peptides, is attributed to the unique properties of arginine amino acid, specifically by its side chain containing amphiphilic guanidinium (Gdm+) ion which makes both intermolecular hydrophobic like-charge Gdm+ - Gdm+ and salt-bridge Gdm+ - C-terminus interactions, where the former are increased with the ionic strength, and the latter decreased due to electrostatic screening. The aggregation behavior of R9 peptides at membranes can also be linked to their CPP translocation properties, suggesting that aggregation may aid in translocation across cellular membranes.

Keywords:  Ionic strength; Molecular dynamics simulations; Peptide aggregation; Phosphocholine lipid bilayers; Polyarginines; Salting-out

DOI:  https://doi.org/10.1016/j.csbj.2024.11.004
ACS Med Chem Lett. 2024 Nov 14. 15(11): 2060-2066

Cyclic Peptide C5aR1 Antagonist Design Using Solution Conformational Analysis Derived from Residual Dipolar Couplings.

Kathleen A Farley, Ye Che, Ricardo Lira, Peter Jones, Nikolaos Papaioannou, Matthew Hayward, Mark E Flanagan, Jonathan Langille, Sidney Liang, Betsy S Pierce, Gregory Ciszewski, Paul Bonin, Fabien Vincent, Simeon Ramsey, David Hepworth.

To gain further insight into the conformational properties of small cyclic peptides that bind to the G-protein coupled receptor C5aR1, we report here for the first time the elucidation of three peptide solution conformations using residual dipolar couplings and NMR temperature coefficients. Each of these peptides varies by at least one amino acid, adopts a different intramolecular hydrogen bonding pattern, and has a different solution conformation. The solution conformations were used in combination with a homology structure of C5aR1 as a design template for increasing the potency of peptide leads for the C5a receptor. This study provides a framework for using RDC solution conformations to guide the design of peptide mimetics that emulate the target bound state in solution to minimize the strain energy of the bound conformation and improve potency of the peptide for the target.

DOI: https://doi.org/10.1021/acsmedchemlett.4c00316
Bioorg Med Chem. 2024 Nov 08. pii: S0968-0896(24)00398-5. [Epub ahead of print]116 117984

Exploring structure-directed immunogenic cytotoxicity of arginine-rich peptides for cytolysis-induced immunotherapy of cancer.

Liu Huang, Ang Li, Han-Jie Liu, Shuang-Shuang Ji, Hao Fei.

  The same cells can die with varied immunological consequences. For the purpose of cancer therapy, stronger immunogenic death of cancer cells is considered favorable. Membrane disruptive peptides are cytotoxic agents with tunable structures capable of not just killing heterogeneous cancer cells, but also inducing immunogenic death. However, the chemo-structural principles that underlie their immunogenic cytotoxicity remain elusive. Here we investigated a series of arginine-rich amphipathic peptides with representative structures on the relationship between the mode of cell death and the immunogenic potency. Among several hydrophobic motif-appended cyclic octaarginine peptides, FC-14 was found to induce cell stress and necroptotic death, unlike apoptotic peptide RL2 and membrane-dissolving peptide RL1. Their differing abilities to release immunogenic death markers correlated well with their potential to activate innate immunity and protective vaccinal effect in a prophylactic model, with FC-14 being the most potent. FC-14 can be pre-opsonized with albumin into nanoparticles (PopAN-FC-14) using PopAN technology to improve its pharmacokinetic properties for intravenous injection. In a syngeneic mouse model of subcutaneous breast cancer, PopAN-FC-14 showed superior therapeutic effect and safety profile than the albumin formulated nanomedicine Nab-paclitaxel (Nab-PTX). Boost injections of PopAN-FC-14 significantly enhanced tumor-specific cellular and humoral immunities, acting similarly as in-situ cancer vaccine. Overall, this work demonstrates a novel focus on the immunogenic cytotoxicity of peptides and a practical approach for effective systemic therapy of cancer.

Keywords:  Albumin nanoparticles; Arginine-rich peptide; Cancer medicine; Cytolytic peptides; Immunogenic cell death

DOI:  https://doi.org/10.1016/j.bmc.2024.117984
Mol Pharm. 2024 Nov 22.

A Cyclic Peptide Targets Glioblastoma by Binding to Aberrantly Exposed SNAP25.

Alberto G Arias, Laura Tovar-Martinez, Eliana K Asciutto, Aman Mann, Kristina Põšnograjeva, Lorena Simón Gracia, Miriam Royo, Maarja Haugas, Tambet Teesalu, Cristian Smulski, Erkki Ruoslahti, Pablo Scodeller.

  Disease-specific changes in tumors and other diseased tissues are an important target of research because they provide clues about the pathophysiology of the disease as well as uncover potentially useful markers for diagnosis and treatment. Here, we report a new cyclic peptide, CESPLLSEC (CES), that specifically accumulated (homed) in intracranial U87MG and the WT-GBM model of glioblastoma (GBM) from intravenous (IV) injection, and associated with the vasculature. Affinity chromatography of U87MG tumor extracts on insolubilized CES peptide identified Synaptosomal Associated Protein 25 (SNAP25) as a candidate target molecule (receptor) for CES. Several results supported the identification of SNAP25 as the CES receptor. IV-injected FAM-CES colocalized with SNAP25 in the tumors, and direct binding studies showed specific binding of the CES peptide to recombinant human SNAP25. A CES peptide-drug conjugate designed for photodynamic therapy showed selective cytotoxicity to SNAP25+ glioblastoma cell lines. Specific accumulation of systemically injected anti-SNAP25 antibody in U87MG glioblastoma and labeling of intact U87MG cells with anti-SNAP in flow cytometry showed that SNAP25 is available from the circulation but not in normal tissues and that it is present at the cell surface. Using an array of ECM proteins and surface plasmon resonance revealed that SNAP25 binds moderately to collagen V and strongly to collagen VI. Modeling studies suggested that CES and collagen VI compete for the same binding site on SNAP25. Our results introduce CES as a valuable targeting peptide for drug delivery and its receptor SNAP25 as a possible molecular marker of interest for glioblastoma.

Keywords:  Glioblastoma; Peptide−Drug Conjugate; Photodynamic Therapy; SNAP25; Targeting Peptide

DOI:  https://doi.org/10.1021/acs.molpharmaceut.4c00958
RSC Chem Biol. 2024 Nov 14.

Rational engineering of an antimalarial peptide with enhanced proteolytic stability and preserved parasite invasion inhibitory activity.

Abhisek Kar, Akash Narayan, Vishal Malik, Kalyaneswar Mandal.

We describe rational chemical engineering to enhance the proteolytic stability of a chimeric peptide using a combination of unique strategies that involve the incorporation of a series of d-amino acids into the parent l-peptide sequence and restricting the conformational freedom of the peptide by covalent stitching. We hypothesize that replacing a stretch of sequence of an unstructured peptide motif with d-amino acids would increase its proteolytic stability without significantly affecting its affinity to the target protein. Also, considering the Cβ-Cβ distances, replacing an appropriate pair of residues with cysteine to form an additional disulfide bond in the molecule would provide additional stability to the engineered peptide. To verify this hypothesis, we have implemented these strategies to a previously reported peptidic inhibitor RR, against P. falciparum invasion into red blood cells (RBCs) and designed two novel heterochiral chimeric peptides, RR-I and RR-II. We have demonstrated that these peptides exhibit remarkable inhibitory activity with dramatically enhanced proteolytic stability. Finally, we have designed a cyclic analog, RR-III, to enhance the stability of the peptide against endopeptidases. The RR-III peptide exhibits the same inhibitory activity as RR-II while demonstrating impressive resistance to enzymatic degradation and prolonged stability in human plasma. These developments hold promise for a new generation of peptide-based therapeutics, showcasing the potential of residue selection for tailored modifications, as demonstrated in this work.

DOI: https://doi.org/10.1039/d4cb00229f
FEBS Open Bio. 2024 Nov 19.

Structural analysis of an Asterias rubens peptide indicates the presence of a disulfide-directed β-hairpin fold.

Rozita Takjoo, David T Wilson, Justine Le Quilliec, Casey A Schmidt, Guangzu Zhao, Michael J Liddell, Naeem Y Shaikh, Kartik Sunagar, Alex Loukas, Michael J Smout, Norelle L Daly.

  Sea stars are an abundant group of marine invertebrates that display remarkably robust regenerative capabilities throughout all life stages. Numerous proteins and peptides have been identified in a proteome study on the coelomic fluid (biofluid) of the common sea star Asterias rubens, which appear to be involved with the wound-healing response in the organism. However, the three-dimensional structure and function of several of these injury-responsive peptides, including the peptide KASH2, are yet to be investigated. Here, we show that the KASH2 peptide adopts a disulfide-directed β-hairpin fold (DDH). The DDH motif appears to be evolutionarily related to the inhibitor cystine knot motif, which is one of the most widespread disulfide-rich peptide folds. The DDH motif was originally thought to be restricted to arachnids, but our study suggests that as a result of convergent evolution it could also have originated in sea stars. Although the widely conserved DDH fold has potential cross-phyla wound-healing capacity, we have shown that KASH2 does not enhance the proliferation of human fibroblasts, a simple method for wound-healing re-epithelialisation screening. Therefore, additional research is necessary to determine the role of KASH2 in the sea stars.

Keywords:  NMR spectroscopy; cell proliferation assay; disulfide‐directed β‐hairpin; molecular evolution; peptide synthesis

DOI:  https://doi.org/10.1002/2211-5463.13931
bioRxiv. 2024 Nov 01. pii: 2024.10.30.621084. [Epub ahead of print]

A quantitative intracellular peptide binding assay reveals recognition determinants and context dependence of short linear motifs.

Mythili S Subbanna, Matthew J Winters, Mihkel Örd, Norman E Davey, Peter M Pryciak.

Transient protein-protein interactions play key roles in controlling dynamic cellular responses. Many examples involve globular protein domains that bind to peptide sequences known as Short Linear Motifs (SLiMs), which are enriched in intrinsically disordered regions of proteins. Here we describe a novel functional assay for measuring SLiM binding, called Systematic Intracellular Motif Binding Analysis (SIMBA). In this method, binding of a foreign globular domain to its cognate SLiM peptide allows yeast cells to proliferate by blocking a growth arrest signal. A high-throughput application of the SIMBA method involving competitive growth and deep sequencing provides rapid quantification of the relative binding strength for thousands of SLiM sequence variants, and a comprehensive interrogation of SLiM sequence features that control their recognition and potency. We show that multiple distinct classes of SLiM-binding domains can be analyzed by this method, and that the relative binding strength of peptides in vivo correlates with their biochemical affinities measured in vitro. Deep mutational scanning provides high-resolution definitions of motif recognition determinants and reveals how sequence variations at non-core positions can modulate binding strength. Furthermore, mutational scanning of multiple parent peptides that bind human tankyrase ARC or YAP WW domains identifies distinct binding modes and uncovers context effects in which the preferred residues at one position depend on residues elsewhere. The findings establish SIMBA as a fast and incisive approach for interrogating SLiM recognition via massively parallel quantification of protein-peptide binding strength in vivo.

DOI: https://doi.org/10.1101/2024.10.30.621084
Chem Soc Rev. 2024 Nov 19.

Recent advances in peptide macrocyclization strategies.

Pengyuan Fang, Wing-Ka Pang, Shouhu Xuan, Wai-Lun Chan, Ken Cham-Fai Leung.

Recently, owing to their special spatial structures, peptide-based macrocycles have shown tremendous promise and aroused great interest in multidisciplinary research ranging from potent antibiotics against resistant strains to functional biomaterials with novel properties. Besides traditional monocyclic peptides, many fascinating polycyclic and remarkable higher-order cyclic, spherical and cylindric peptidic systems have come into the limelight owing to breakthroughs in various chemical (e.g., native chemical ligation and transition metal catalysis), biological (e.g., post-translational enzymatic modification and genetic code reprogramming), and supramolecular (e.g., mechanically interlocked, metal-directed folding and self-assembly via noncovalent interactions) macrocyclization strategies developed in recent decades. In this tutorial review, diverse state-of-the-art macrocyclization methodologies and techniques for peptides and peptidomimetics are surveyed and discussed, with insights into their practical advantages and intrinsic limitations. Finally, the synthetic-technical aspects, current unresolved challenges, and outlook of this field are discussed.

DOI: https://doi.org/10.1039/d3cs01066j
ACS Med Chem Lett. 2024 Nov 14. 15(11): 1918-1924

Hybrids of Membrane-Translocating Antimicrobial Peptides Show Enhanced Activity through Membrane Permeabilization.

Giulia F Trevellin, JuYoung Kwag, Michelle L Shui, Hannah Klim, Valentina Alvarez, Louise E O Darling, Donald E Elmore.

Antimicrobial peptides (AMPs) hold promise as useful tools to combat bacterial infection. Hybrid peptides, made by linking two independent AMPs together through peptide bonds, have the potential for enhancing antimicrobial activity. Here we explore hybrids created by combining two histone-derived antimicrobial peptides (HDAPs), BF2 and DesHDAP1, that each translocate across bacterial membranes. Our work represents the first systematic approach considering the activity and mechanism of hybrids made from two translocating AMPs. BF2/DesHDAP1 hybrids showed increased antimicrobial activity against both Gram-positive and Gram-negative bacteria compared with the parent peptides and no cytotoxicity against eukaryotic cells. Introducing amino acid linkers between the parent peptides did not further enhance the antibacterial activity. The increased antimicrobial activity comes from a mechanistic shift, as hybrid peptides show decreased translocation across bacterial cell membranes but increased membrane permeabilization compared to BF2 and DesHDAP1. These observations lay the groundwork for the further design of hybrid AMPs made from translocating peptides.

DOI: https://doi.org/10.1021/acsmedchemlett.4c00375
Oncotarget. 2024 Nov 22. 15 815-816

Advancements in cell-penetrating monoclonal antibody treatment.

Sai Pallavi Pradeep, Raman Bahal.



Keywords:  3E10; cell penetration; monoclonal anti-bodies; nucleic acid delivery

DOI:  https://doi.org/10.18632/oncotarget.28674