bims-cepepe Biomed News
on Cell-penetrating peptides
Issue of 2024–05–19
eleven papers selected by
Henry Lamb, Queensland University of Technology
  1. DexDesign: an OSPREY-based algorithm for designing de novo D-peptide inhibitors.
  2. Macrocyclization strategy for improving candidate profiles in medicinal chemistry.
  3. Synthesis of Membrane-Permeable Macrocyclic Peptides via Imidazopyridinium Grafting.
  4. Isolation and Characterization of Insecticidal Cyclotides from Viola communis.
  5. An Amphiphilic Cell-Penetrating Macrocycle for Efficient Cytosolic Delivery of Proteins, DNA and CRISPR Cas9.
  6. Bioinformatics leading to conveniently accessible, helix enforcing, bicyclic ASX motif mimics (BAMMs).
  7. An In Vitro Model of Blood-Brain Barrier for Studies on HIV Neuroinflammation and CNS Antibody Penetration.
  8. Merging Flow Synthesis and Enzymatic Maturation to Expand the Chemical Space of Lasso Peptides.
  9. Protocol to generate scaffold-free, multicomponent 3D melanoma spheroid models for preclinical drug testing.
  10. Radiopharmaceuticals for Cancer Diagnosis and Therapy: New Targets, New Therapies-Alpha-Emitters, Novel Targets.
  11. Synthesis and Preclinical Evaluation of Urea-Based Prostate-Specific Membrane Antigen-Targeted Conjugates Labeled with 177Lu.
  1. Protein Eng Des Sel. 2024 Jan 29. pii: gzae007. [Epub ahead of print]37
    DexDesign: an OSPREY-based algorithm for designing de novo D-peptide inhibitors.
    Nathan Guerin, Henry Childs, Pei Zhou, Bruce R Donald.
      With over 270 unique occurrences in the human genome, peptide-recognizing PDZ domains play a central role in modulating polarization, signaling, and trafficking pathways. Mutations in PDZ domains lead to diseases such as cancer and cystic fibrosis, making PDZ domains attractive targets for therapeutic intervention. D-peptide inhibitors offer unique advantages as therapeutics, including increased metabolic stability and low immunogenicity. Here, we introduce DexDesign, a novel OSPREY-based algorithm for computationally designing de novo D-peptide inhibitors. DexDesign leverages three novel techniques that are broadly applicable to computational protein design: the Minimum Flexible Set, K*-based Mutational Scan, and Inverse Alanine Scan. We apply these techniques and DexDesign to generate novel D-peptide inhibitors of two biomedically important PDZ domain targets: CAL and MAST2. We introduce a framework for analyzing de novo peptides-evaluation along a replication/restitution axis-and apply it to the DexDesign-generated D-peptides. Notably, the peptides we generated are predicted to bind their targets tighter than their targets' endogenous ligands, validating the peptides' potential as lead inhibitors. We also provide an implementation of DexDesign in the free and open source computational protein design software OSPREY.
    Keywords:  D-peptide; DexDesign; OSPREY; algorithms; peptide design
    DOI:  https://doi.org/10.1093/protein/gzae007
  2. Eur J Med Chem. 2024 May 14. pii: S0223-5234(24)00381-7. [Epub ahead of print]272 116501
    Macrocyclization strategy for improving candidate profiles in medicinal chemistry.
    Om Darlami, Rabin Pun, Sung-Hoon Ahn, Seok-Ho Kim, Dongyun Shin.
      Macrocycles are defined as cyclic compounds with 12 or more members. In medicinal chemistry, they are categorized based on their core chemistry into cyclic peptides and macrocycles. Macrocycles are advantageous because of their structural diversity and ability to achieve high affinity and selectivity towards challenging targets that are often not addressable by conventional small molecules. The potential of macrocyclization to optimize drug-like properties while maintaining adequate bioavailability and permeability has been emphasized as a key innovation in medicinal chemistry. This review provides a detailed case study of the application of macrocyclization over the past 5 years, starting from the initial analysis of acyclic active compounds to optimization of the resulting macrocycles for improved efficacy and drug-like properties. Additionally, it illustrates the strategic value of macrocyclization in contemporary drug discovery efforts.
    Keywords:  Macrocycle; Macrocyclization; Pharmacokinetics; Potency; Selectivity; hERG
    DOI:  https://doi.org/10.1016/j.ejmech.2024.116501
  3. J Am Chem Soc. 2024 May 16.
    Synthesis of Membrane-Permeable Macrocyclic Peptides via Imidazopyridinium Grafting.
    Bo Li, Joshua Parker, Joel Tong, Thomas Kodadek.
      Macrocyclic peptides (MPs) are a class of compounds that have been shown to be particularly well suited for engaging difficult protein targets. However, their utility is limited by their generally poor cell permeability and bioavailability. Here, we report an efficient solid-phase synthesis of novel MPs by trapping a reversible intramolecular imine linkage with a 2-formyl- or 2-keto-pyridine to create an imidazopyridinium (IP+)-linked ring. This chemistry is useful for the creation of macrocycles of different sizes and geometries, including head-to-side and side-to-side chain configurations. Many of the IP+-linked MPs exhibit far better passive membrane permeability than expected for "beyond Rule of 5" molecules, in some cases exceeding that of much lower molecular weight, traditional drug molecules. We demonstrate that this chemistry is suitable for the creation of libraries of IP+-linked MPs and show that these libraries can be mined for protein ligands.
    DOI:  https://doi.org/10.1021/jacs.4c01920
  4. J Nat Prod. 2024 May 15.
    Isolation and Characterization of Insecticidal Cyclotides from Viola communis.
    Negin Khatibi, Yen-Hua Huang, Conan K Wang, Thomas Durek, Edward K Gilding, David J Craik.
      Cyclotides are cysteine-rich plant-derived peptides composed of 28-37 amino acids with a head-to-tail cyclic backbone and a knotted arrangement of three conserved disulfide bonds. Their beneficial biophysical properties make them promising molecules for pharmaceutical and agricultural applications. The Violaceae plant family is the major cyclotide-producing family, and to date, every examined plant from this family has been found to contain cyclotides. The presence of cyclotides in Viola communis was inferred by mass spectroscopy previously, but their sequences and properties had yet to be explored. In this study, the occurrence of cyclotides in this plant was investigated using proteomics and transcriptomics. Twenty cyclotides were identified at the peptide level, including two new members from the bracelet (Vcom1) and Möbius (Vcom2) subfamilies. Structural analysis of these newly identified peptides demonstrated a similar fold compared with cyclotides from the same respective subfamilies. Biological assays of Vcom1 and Vcom2 revealed them to be cytotoxic to Sf9 insect cell lines, with Vcom1 demonstrating higher potency than Vcom2. The results suggest that they could be further explored as insecticidal agents and confirm earlier general findings that bracelet cyclotides have more potent insecticidal activity than their Möbius relatives. Seven new cyclotide-like sequences were observed in the transcriptome of V. communis, highlighting the Violaceae as a rich source for new cyclotides with potential insecticidal activity. An analysis of sequences flanking the cyclotide domain in the various precursors from V. communis and other Violaceae plants revealed new insights into cyclotide processing and suggested the possibility of two alternative classes of N-terminal processing enzymes for cyclotide biosynthesis.
    DOI:  https://doi.org/10.1021/acs.jnatprod.4c00168
  5. Angew Chem Int Ed Engl. 2024 May 16. e202403647
    An Amphiphilic Cell-Penetrating Macrocycle for Efficient Cytosolic Delivery of Proteins, DNA and CRISPR Cas9.
    Somayah Qutub, Imtiyaz Bhat, Batoul Maatouk, Basem Moosa, Kashif Nawaz, Escarlet Diaz Galicia, Aliyah Fakim, Weibin Lin, Raik Grünberg, Stefan Arold, Niveen M Khashab.
      The discovery of safe platforms that can circumvent the endocytic pathway is of great significance for biological therapeutics that are usually degraded during endocytosis. Here we show that self-assembled and dynamic macrocycles can passively diffuse through the cell membrane and deliver a broad range of biologics including proteins, CRISPR Cas9 and ssDNA directly to the cytosol while retaining their bioactivity. Cell penetrating macrocycles (CPMs) can be easily prepared from the room temperature condensation of diketopyrrolopyrrole lactams with diamines. We attribute the high cellular permeability of CPMs to their amphiphilic nature and chameleonic properties. They adopt conformations that partially bury polar groups and expose hydrophobic side chains thus self-assembling into micellar-like structures. Their superior fluorescence renders CPMs trackable inside cells where they follow the endomembrane system. CPMs outperformed commercial reagents for biologics delivery and showed high RNA knockdown efficiency of CRISPR Cas9. We envisage that this class of macrocycles will be an ideal starting point to design and synthesize biomimetic macrocyclic tags that can readily facilitate the interaction and uptake of biomolecules and overcome endosomal digestion.
    Keywords:  Fluorescence; Passive delivery; Self-Assembly; biomimetic; proteins
    DOI:  https://doi.org/10.1002/anie.202403647
  6. Nat Commun. 2024 May 17. 15(1): 4217
    Bioinformatics leading to conveniently accessible, helix enforcing, bicyclic ASX motif mimics (BAMMs).
    Tianxiong Mi, Duyen Nguyen, Zhe Gao, Kevin Burgess.
      Helix mimicry provides probes to perturb protein-protein interactions (PPIs). Helical conformations can be stabilized by joining side chains of non-terminal residues (stapling) or via capping fragments. Nature exclusively uses capping, but synthetic helical mimics are heavily biased towards stapling. This study comprises: (i) creation of a searchable database of unique helical N-caps (ASX motifs, a protein structural motif with two intramolecular hydrogen-bonds between aspartic acid/asparagine and following residues); (ii) testing trends observed in this database using linear peptides comprising only canonical L-amino acids; and, (iii) novel synthetic N-caps for helical interface mimicry. Here we show many natural ASX motifs comprise hydrophobic triangles, validate their effect in linear peptides, and further develop a biomimetic of them, Bicyclic ASX Motif Mimics (BAMMs). BAMMs are powerful helix inducing motifs. They are synthetically accessible, and potentially useful to a broad section of the community studying disruption of PPIs using secondary structure mimics.
    DOI:  https://doi.org/10.1038/s41467-024-48323-z
  7. Methods Mol Biol. 2024 ;2807 271-283
    An In Vitro Model of Blood-Brain Barrier for Studies on HIV Neuroinflammation and CNS Antibody Penetration.
    Yuekun Lang, Dina Mofed, Vasudev R Rao, Samantha J Welninski, Anjelica F Reyes, Jeffrey R Schneider, Vinayaka R Prasad.
      The blood-brain barrier (BBB) is one of several barriers between the brain and the peripheral blood system to maintain homeostasis. Understanding the interactions between infectious agents such as human immunodeficiency virus type 1 (HIV-1), which are capable of traversing the BBB and causing neuroinflammation requires modeling an authentic BBB in vitro. Such an in vitro BBB model also helps develop means of targeting viruses that reside in the brain via natural immune effectors such as antibodies. The BBB consists of human brain microvascular endothelial cells (HBMECs), astrocytes, and pericytes. Here we report in vitro methods to establish a dual-cell BBB model consisting of primary HBMECs and primary astrocytes to measure the integrity of the BBB and antibody penetration of the BBB, as well as a method to establish a single cell BBB model to study the impact of HIV-1 infected medium on the integrity of such a BBB.
    Keywords:  Astrocytes; Human brain microvascular endothelial cells; IgG CNS penetration; In vitro blood-brain barrier
    DOI:  https://doi.org/10.1007/978-1-0716-3862-0_19
  8. J Am Chem Soc. 2024 May 17.
    Merging Flow Synthesis and Enzymatic Maturation to Expand the Chemical Space of Lasso Peptides.
    Kevin Schiefelbein, Jakob Lang, Matthias Schuster, Claire E Grigglestone, Robin Striga, Laurent Bigler, Meredith C Schuman, Oliver Zerbe, Yanyan Li, Nina Hartrampf.
      Many peptidic natural products, such as lasso peptides, cyclic peptides, and cyclotides, are conformationally constrained and show biological stability, making them attractive scaffolds for drug development. Although many peptides can be synthesized and modified through chemical methods, knot-like lasso peptides such as microcin J25 (MccJ25) and their analogues remain elusive. As the chemical space of MccJ25 analogues accessible through purely biological methods is also limited, we proposed a hybrid approach: flow-based chemical synthesis of non-natural precursor peptides, followed by in vitro transformation with recombinant maturation enzymes, to yield a more diverse array of lasso peptides. Herein, we established the rapid, flow-based synthesis of chemically modified MccJ25 precursor peptides (57 amino acids). Heterologous expression of enzymes McjB and McjC was extensively optimized to improve yields and facilitate the synthesis of multiple analogues of MccJ25, including the incorporation of non-canonical tyrosine and histidine derivatives into the lasso scaffold. Finally, using our chemoenzymatic strategy, we produced a biologically active analogue containing three d-amino acids in the loop region and incorporated backbone N-methylations. Our method provides rapid access to chemically modified lasso peptides that could be used to investigate structure-activity relationships, epitope grafting, and the improvement of therapeutic properties.
    DOI:  https://doi.org/10.1021/jacs.4c03898
  9. STAR Protoc. 2024 May 13. pii: S2666-1667(24)00223-5. [Epub ahead of print]5(2): 103058
    Protocol to generate scaffold-free, multicomponent 3D melanoma spheroid models for preclinical drug testing.
    Cristian Angeli, Joanna Patrycja Wroblewska, Eliane Klein, Christiane Margue, Stephanie Kreis.
      Three-dimensional (3D) models play an increasingly important role in preclinical drug testing as they faithfully mimic interactions between cancer cells and the tumor microenvironment (TME). Here, we present a protocol for generating scaffold-free 3D multicomponent human melanoma spheroids. We describe steps for characterizing models using live-cell imaging and histology, followed by drug testing and assessment of cell death through various techniques such as imaging, luminescence-based assays, and flow cytometry. Finally, we demonstrate the models' adaptability for co-cultures with immune cells.
    Keywords:  Cancer; Cell Biology; High Throughput Screening; Organoids
    DOI:  https://doi.org/10.1016/j.xpro.2024.103058
  10. Cancer J. 2024 May-Jun 01;30(3):30(3): 218-223
    Radiopharmaceuticals for Cancer Diagnosis and Therapy: New Targets, New Therapies-Alpha-Emitters, Novel Targets.
    Neil K Taunk, Freddy E Escorcia, Jason S Lewis, Lisa Bodei.
       ABSTRACT: Radiopharmaceutical therapy has emerged as a promising approach for the treatment of various cancers. The exploration of novel targets such as tumor-specific antigens, overexpressed receptors, and intracellular biomolecules using antibodies, peptides, or small molecules has expanded the scope of radiopharmaceutical therapy, enabling precise and effective cancer treatment for an increasing number of tumor types. Alpha emitters, characterized by their high linear energy transfer and short path length, offer unique advantages in targeted therapy due to their potent cytotoxicity against cancer cells while sparing healthy tissues. This article reviews recent advancements in identifying novel targets for radiopharmaceutical therapy and applications in utilizing α-emitters for targeted treatment.
    DOI:  https://doi.org/10.1097/PPO.0000000000000720
  11. ACS Pharmacol Transl Sci. 2024 May 10. 7(5): 1457-1473
    Synthesis and Preclinical Evaluation of Urea-Based Prostate-Specific Membrane Antigen-Targeted Conjugates Labeled with 177Lu.
    Aleksei E Machulkin, Stanislav A Petrov, Vitalina Bodenko, Mariia S Larkina, Evgenii Plotnikov, Feruza Yuldasheva, Maria Tretyakova, Ekaterina Bezverkhniaia, Nikolay Yu Zyk, Elena Stasyuk, Roman Zelchan, Alexander G Majouga, Vladimir Tolmachev, Anna Orlova, Elena K Beloglazkina, Mekhman S Yusubov.
      177Lu-labeled small-molecule prostate-specific membrane antigen (PSMA) targeted tracers are therapeutic agents for metastatic castration-resistant prostate cancer. Optimizing molecular design holds the potential to further enhance the pharmacokinetic properties of PSMA-targeted agents while preserving their potent therapeutic effects. In this study, six novel N-[N-[(S)-1,3-dicarboxypropyl]carbamoyl]-(S)-l-lysine (DCL) urea-based PSMA ligand 2,2',2″,2‴-(1,4,7,10-tetraazacyclododecane-1,4,7,10-tetrayl)tetraacetic acid conjugates were synthesized. These conjugates feature polypeptide linkers containing the Phe-Phe peptide sequence and an aromatic fragment at the ε-NH-Lys group of the DCL fragment. The synthesis yielded products with satisfactory yields ranging from 60% to 72%, paving the way for their preclinical evaluation. The labeling of the new variants of urea-based PSMA inhibitors provided a radiochemical yield of over 95%. The 177Lu-labeled conjugates demonstrated specific and moderate affinity binding to PSMA-expressing human cancer cells PC3-pip in vitro and specific accumulation in PSMA-expressing xenografts in vivo. Based on the results, both the lipophilicity and the type of substituent in the linker significantly influence the binding properties of the PSMA inhibitor and its biodistribution profile. Specifically, the studied variants containing a bromine substituent or a hydroxyl group introduced into the aromatic fragment of the phenylalanyl residue in DCL exhibit higher affinities to PSMA compared to variants with only a chlorine-substituted aromatic fragment or variants without any substituents. The [177Lu]Lu-13C with the bromine substituent was characterized by the highest activity accumulation in blood, salivary glands, muscle, bone, and gastrointestinal tract and had inasmuch as an unfavorable pharmacokinetic profile. The negative charge of the carboxyl group in the phenyl moiety of the [177Lu]Lu-13A variant has demonstrated a positive effect on reducing the retention of activity in the liver and the kidneys (the ratio of tumor to kidneys was 1.3-fold). Low accumulation in normal tissues in vivo indicates that this novel PSMA-targeting inhibitor is a promising radioligand.
    DOI:  https://doi.org/10.1021/acsptsci.4c00070
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