bims-cepepe Biomed News
on Cell-penetrating peptides
Issue of 2025–03–09
ten papers selected by
Henry Lamb, Queensland University of Technology
  1. Bulk measurement of membrane permeability for random cyclic peptides in living cells to guide drug development.
  2. Comprehensive invitro evaluation of the inhibitory effects of relatively high molecular weight peptides on drug-drug interaction-associated four liver transporters and its association with physicochemical properties.
  3. Reniochalistatin E: A potentiating agent and a potential novel drug delivery platform.
  4. Enzymatic crosslinking of histidine side chains in peptide natural products.
  5. Application of phage surface display for the identification of Eu3+-binding peptides.
  6. Design, synthesis, and biological evaluation of novel and highly potent peptides targeting syntenin.
  7. Structure-based discovery of hydrocarbon-stapled paxillin peptides that block FAK scaffolding in cancer.
  8. Membrane selectivity and pore formation of SprA1 and SprA2 hemolytic peptides from Staphylococcus aureus type I toxin-antitoxin systems.
  9. Rationally designed peptides inhibit the formation of α-synuclein fibrils and oligomers.
  10. Cell-penetrating peptide-grafted AAV2 capsids for improved retinal delivery via intravitreal injection.
  1. Angew Chem Int Ed Engl. 2025 Mar 07. e202500493
    Bulk measurement of membrane permeability for random cyclic peptides in living cells to guide drug development.
    Alexander L Nielsen, Christian R O Bartling, Anne Zarda, Nathan De Sadeleer, Rebecca M Neeser, Phillippe Schwaller, Kristian Strømgaard, Christian Heinis.
      Cyclic peptides are attractive for drug discovery due to their excellent binding properties and the potential to cross cell membranes. However, by far not all cyclic peptides are cell permeable, and measuring or predicting their membrane permeability is not trivial. In this work, we assessed the membrane permeability of thioether-cyclized peptides, a widely used format in drug discovery. We developed a strategy for synthesizing hundreds of cyclic peptides carrying a short chloroalkane tag for the bulk quantification of membrane permeability in live cells using the chloroalkane penetration assay. Permeability data for random cyclic peptides established design rules, namely that the probability of peptides entering cells is strongly increasing if the molecular weight is below 800 Da, the polar surface is smaller than 250 Å2, or if there are less than six H-bond donors. From this, machine learning could predict the membrane permeability of random peptides with good confidence, facilitating the future development of membrane permeable cyclic peptide drugs.
    Keywords:  CAPA; chloroalkane tag; cyclic peptide; macrocycle; membrane permeability
    DOI:  https://doi.org/10.1002/anie.202500493
  2. Drug Metab Pharmacokinet. 2025 Feb 03. pii: S1347-4367(25)00005-9. [Epub ahead of print]61 101055
    Comprehensive invitro evaluation of the inhibitory effects of relatively high molecular weight peptides on drug-drug interaction-associated four liver transporters and its association with physicochemical properties.
    Rika Ishikawa, Takashi Misawa, Yosuke Demizu, Yoshiro Saito, Ruri Kikura-Hanajiri, Kosuke Saito.
      In recent years, advances in peptide synthesis have enabled the construction of relatively high molecular weight (Mw; >1 kDa) peptides using various types of amino acids (AAs), including proteinogenic/natural and nonnatural AAs. This advancement helps in obtaining peptides with improved stability, cell membrane permeability, and/or target-binding affinity. However, drug-drug interaction (DDI) information for these peptides remains scarce. Therefore, we focused on relatively high Mw peptides to examine their potential in inhibiting liver transporters, organic anion transporting polypeptide (OATP) 1B1, OATP1B3, P-glycoprotein, and breast cancer resistant protein (BCRP) in vitro. We addressed the inhibitory effects of various types of cyclic peptides containing non-natural AAs and cell-penetrating peptides composed of proteinogenic/natural AAs. Our results demonstrated that several peptides inhibited transport activities, indicating that they can potentially cause DDI. We further evaluated the relationship between their inhibition potency and physicochemical properties (Mw and hydrophobicity or charge of the constituting AA) to characterize the specific physicochemical properties contributing to their inhibition potency. The hydrophobic AA contents of the peptides correlated with the inhibition potencies for all four transporters. Our findings demonstrate the transporter-mediated DDI potential of peptides and the necessity of their evaluation for drug development.
    Keywords:  Cell penetrating peptide; Cyclic peptide; DDI-Associated transporters; Drug–drug interaction; Physicochemical property
    DOI:  https://doi.org/10.1016/j.dmpk.2025.101055
  3. Bioorg Chem. 2025 Mar 01. pii: S0045-2068(25)00211-1. [Epub ahead of print]157 108331
    Reniochalistatin E: A potentiating agent and a potential novel drug delivery platform.
    Kameron R Wildeman, Matthew Barnett, Anthony Fatino, Chamitha Weeramange, Ryan J Rafferty.
      Peptides, both linear and cyclic, have begun to emerge as a viable therapeutic for the treatment of various diseases. As such, synthesizing these linear and macrocyclic peptides as natural product targets or related structural analogs can help us to understand their biological importance. Reniochalistatin E is a proline-rich macrocyclic peptide natural product first synthesized in 2017 by our laboratory, and screening against various cancerous cell lines revealed moderate to low cytotoxicity as a sole agent. We next became interested in studying this compound as a potentiating agent to identify any synergistic effects. Indeed, reniochalistatin E was unveiled to show potential intramolecular pi-stacking that enhanced the cytotoxicity of clinically used anti-cancer agents. To exploit this unique characteristic, attention was directed towards the modification of the tryptophan residue to enhance the synergistic effect with additional cytotoxic agents. To this, a phenylalanine-tryptophan exchange was undertaken to probe this hypothesis and was found to possess no detectable cytotoxicity variation. Together, the data collected supports further exploration with drug conjugate handles en route towards a novel drug delivery platform.
    Keywords:  Anticancer; Medicinal chemistry; Natural product; Reniochalistatin E; Synergy
    DOI:  https://doi.org/10.1016/j.bioorg.2025.108331
  4. Nat Prod Rep. 2025 Mar 03.
    Enzymatic crosslinking of histidine side chains in peptide natural products.
    Ying Shi, Yinzheng Xia, Weijiang Gao, Jingxue Wang, Bing Shi, Huan Wang.
      Covering: 2019 to 2024Peptide macrocyclization stands as the pivotal step in the biosynthesis journey of bioactive cyclic peptide natural products, spanning both ribosomal and non-ribosomal origins. Beyond the enzymatic N- to C-terminus macrocyclization, natural cyclic peptides frequently display side chain-to-side chain crosslinks, which markedly bolster their stability and biological potency. Traditionally, histidine, with its imidazole side chain, has been regarded as chemically unreactive, leading to relatively sparse reports of histidine-containing crosslinks in cyclic peptide natural products. However, recent advancements in research have illuminated a novel perspective on the role of histidine (His) residues in peptide macrocyclization, revealing that His participation in this process is far more ubiquitous than previously envisioned. This highlight underscores the significance of His-containing crosslinks in natural cyclic peptides and delves into the enzymatic mechanisms underlying their formation.
    DOI:  https://doi.org/10.1039/d5np00001g
  5. Front Bioeng Biotechnol. 2025 ;13 1508018
    Application of phage surface display for the identification of Eu3+-binding peptides.
    Gerda Techert, Björn Drobot, Robert Braun, Christoph Bloss, Nora Schönberger, Sabine Matys, Katrin Pollmann, Franziska L Lederer.
      Europium as one of the rare earth elements (REE) has outstanding properties in terms of its application for high-tech and renewable energy products. The high supply risk of REE, coupled with their low recovery rates from secondary sources, necessitates innovative recycling approaches. We introduce a phage display-based peptide biosorbent recycling technology that offers a cost-effective and environmentally friendly solution for recovering metal ions, supporting circular economy goals. In this study, we used phage surface display to screen for peptides with high affinity for europium (III) ions (Eu3+). Performing several independent biopanning experiments with the Ph.D.-12 Phage Display Peptide Library and different elution methods as well as combining them with next-generation sequencing, we identified eight peptides with moderate to good affinities for Eu3+ ions, verified by time-resolved laser fluorescence spectroscopy. The peptides EALTVNIKREME as well as DVHHVDGNDLQPFEGGGS and DSIHSDVTKDGRYPVEGGGS, the latter are variants of enriched dodecamers, proved to be the best candidates for future biosorption and selectivity studies. This study underscores the potential of phage surface display for peptide-based REE recovery, laying the foundation for selective recycling technologies from secondary raw materials.
    Keywords:  REE recovery; europium; next generation sequencing; peptides; phage surface display
    DOI:  https://doi.org/10.3389/fbioe.2025.1508018
  6. Eur J Med Chem. 2025 Feb 25. pii: S0223-5234(25)00211-9. [Epub ahead of print]289 117446
    Design, synthesis, and biological evaluation of novel and highly potent peptides targeting syntenin.
    Yang Zhou, Yuting Wang, Juanjuan Liu, Yu Bai, Jinliang Ma, Miao-Miao Niu, Jindong Li, Haijing Jiang.
      Syntenin, an intracellular scaffold protein, plays a critical role in renal cell carcinoma (RCC) progression, underscoring its potential as a therapeutic target. Herein, we report a novel, highly efficient, and stable peptide inhibitor (PDPP-3) that exhibits excellent inhibitory effects on syntenin. We have constructed a combined virtual screening scheme based on pharmacophore modeling and molecular docking to identify six potential d-amino acid-containing peptide inhibitors targeting syntenin. Among them, PDPP-3 showed the best inhibitory activity against syntenin. Binding affinity experiments and biostability experiments indicated that the interaction between PDPP-3 and syntenin displayed nanomolar-level binding affinity (Kd = 6.15 ± 0.12 nM) and superior biostability in serum. Molecular dynamics simulation results further confirmed that PDPP-3 could stably bind to the active site of syntenin. Additionally, cytotoxicity test results showed that PDPP-3 exhibited potent inhibitory effects on various types of renal cancer cells, with the best inhibitory effect on SK-RC-20 cells. More importantly, PDPP-3 significantly downregulates the expression of matrix metalloenzymes MT1-MMP and MMP2, which are pivotal for tumor invasion, and demonstrates inhibitory effects on tumor growth in SK-RC-20 derived xenografts. These data suggest that PDPP-3 may be a very promising candidate drug for the treatment of RCC.
    Keywords:  Molecular dynamics; Peptide inhibitor; Renal cell carcinoma (RCC); Syntenin; Virtual screening
    DOI:  https://doi.org/10.1016/j.ejmech.2025.117446
  7. Nat Commun. 2025 Feb 28. 16(1): 2060
    Structure-based discovery of hydrocarbon-stapled paxillin peptides that block FAK scaffolding in cancer.
    Lauren Reyes, Lena Naser, Warren S Weiner, Darren Thifault, Erik Stahl, Liam McCreary, Rohini Nott, Colton Quick, Alex Buchberger, Carlos Alvarado, Andrew Rivera, Joseph A Miller, Ruchi Khatiwala, Brian R Cherry, Ronald Nelson, Jose M Martin-Garcia, Nicholas Stephanopoulos, Raimund Fromme, Petra Fromme, William Cance, Timothy Marlowe.
      The focal adhesion kinase (FAK) scaffold provides FAK-targeted cancer therapeutics with greater efficacy and specificity than traditional kinase inhibitors. The FAK scaffold function largely involves the interaction between FAK's focal adhesion targeting (FAT) domain and paxillin, ultimately regulating many hallmarks of cancer. We report the design of paxillin LD-motif mimetics that successfully inhibit the FAT-paxillin interaction. Chemical and biochemical screening identifies stapled peptide 1907, a high affinity binder of the FAT four-helix bundle with ~100-fold greater binding affinity than the native LD2-sequence. The X-ray co-crystal structure of the FAT-1907 complex is solved. Myristoylated 1907-analog, peptide 2012, delocalizes FAK from focal adhesions, induces cancer cell apoptosis, reduces in vitro viability and invasion, and decreases tumor burden in B16F10 melanoma female mice. Enzymatic FAK inhibition produces no comparable effects. Herein, we describe a biologically potent therapeutic strategy to target the FAK-paxillin complex, a previously deemed undruggable protein-protein interaction.
    DOI:  https://doi.org/10.1038/s41467-025-57196-9
  8. FEBS J. 2025 Mar 03.
    Membrane selectivity and pore formation of SprA1 and SprA2 hemolytic peptides from Staphylococcus aureus type I toxin-antitoxin systems.
    Laurence Fermon, Noëlla Germain-Amiot, Charlotte Oriol, Astrid Rouillon, Yoann Augagneur, Stéphane Dréano, Irène Nicolas, Alexandre Chenal, Arnaud Bondon, Soizic Chevance, Marie-Laure Pinel-Marie.
      SprA1 and SprA2 are small hydrophobic peptides that belong to the type I toxin-antitoxin systems expressed by Staphylococcus aureus. Both peptides induce S. aureus death when overexpressed. Although they share 71% of amino acids sequence similarity, SprA2 exhibits stronger hemolytic activity than SprA1. In this study, we investigated the mode of action of these toxins on both prokaryotic-like and eukaryotic-like membranes. We first confirmed that SprA2, like SprA1, is an alpha-helical peptide located at the S. aureus membrane. By overexpressing each toxin, we demonstrated that SprA1 forms stable pores in the S. aureus membrane, evidenced by concomitant membrane depolarization, permeabilization and ATP release leading to growth arrest, whereas SprA2 forms transient pores, causing concomitant membrane depolarization, ATP release, and growth arrest. We showed that the unique cysteine residue present in SprA1 and SprA2 is required for toxicity through disulfide bond formation. Next, we found that both synthetic peptides induce slight leakage in anionic DOPC-DOPG lipid vesicles mimicking prokaryotic membranes, concomitant with lipid vesicles aggregation and/or fusion. Moreover, we observed that SprA1 permeabilizes S. aureus protoplasts, via its ability to form stable pores, whereas SprA2 permeabilizes and lyses them. However, no permeabilization of intact bacteria was detected after the addition of SprA1 and SprA2 in the extracellular medium. Finally, we confirmed that SprA2 has strong activity on zwitterionic DOPC lipid vesicles mimicking eukaryotic membranes, without inducing aggregation. This work highlights the strong selectivity of SprA2 for eukaryotic membranes, suggesting that this toxin may play a role in S. aureus virulence.
    Keywords:  Staphylococcus aureus; membrane depolarization; membrane permeabilization; membrane‐active peptide; pore formation; type I toxin–antitoxin system
    DOI:  https://doi.org/10.1111/febs.70001
  9. Eur J Med Chem. 2025 Feb 26. pii: S0223-5234(25)00217-X. [Epub ahead of print]289 117452
    Rationally designed peptides inhibit the formation of α-synuclein fibrils and oligomers.
    Tariq T Ali, Madiha Merghani, Mohammed Al-Azzani, Luisa Maria Gatzemeier, Michael Hoppert, Dora Kaloyanova, Tiago F Outeiro, Piotr Neumann, Blagovesta Popova, Gerhard H Braus.
      Parkinson's Disease (PD) is characterized by the pathological aggregation of α-synuclein (αSyn) into oligomers and amyloid fibrils, making αSyn aggregation a key target for drug development. Peptides have gained recent attention as potential agents to inhibit aggregation. Two previously identified peptide inhibitors, discovered through large-scale yeast screening, were used as templates for in silico mutagenesis aimed at designing novel peptides with improved efficacy in inhibiting αSyn aggregation and cytotoxicity. The newly designed peptides underwent in silico docking analysis, and the most promising candidates were tested in vitro and in cellular models. Peptides T02 and T05 emerged as the most effective inhibitors, with T02 binding αSyn monomers and T05 targeting lower-order oligomers. Both peptides reduce αSyn fibril and oligomer formation in vitro and significantly suppress αSyn aggregation and cytotoxicity in yeast and human H4 cells. These novel peptides represent antagonists of αSyn aggregation with promising potential for therapeutic intervention for PD.
    Keywords:  Alpha-synuclein; Alpha-synuclein oligomerization; Parkinson disease; Peptide drug discovery; Protein aggregation; Rational peptide design
    DOI:  https://doi.org/10.1016/j.ejmech.2025.117452
  10. Mol Ther Methods Clin Dev. 2025 Mar 13. 33(1): 101426
    Cell-penetrating peptide-grafted AAV2 capsids for improved retinal delivery via intravitreal injection.
    Jiang-Hui Wang, Mengtian Cui, Hao Liu, Peiyi Guo, Jackson McGowan, Shun-Yun Cheng, Dominic J Gessler, Jun Xie, Claudio Punzo, Phillip W L Tai, Guangping Gao.
      Recombinant adeno-associated virus (rAAV) is a leading vector for retinal gene therapy due to its favorable safety profile demonstrated by the FDA-approved Luxturna for Leber congenital amaurosis. However, challenges with low transduction efficiency and immunogenicity, coupled with the invasiveness of subretinal injections, have driven efforts to engineer AAV capsids for minimally invasive intravitreal delivery. Intravitreal injections face the barrier of the inner limiting membrane (ILM), particularly with AAV2-based vectors. In this study, we displayed cell-penetrating peptides (CPPs) on AAV2 capsids to enhance retinal cell transduction via intravitreal injection. Through in vivo capsid screening, we identified AAV2.CPP1, which showed significantly improved pan-retinal expression and photoreceptor transduction in mice as well as a reduced immune response compared to the AAV2.7m8 vector. We also revealed that the CPP1 insertion reduced heparan sulfate binding, improving ILM penetration. These findings highlight AAV2.CPP1 as a promising candidate for retinal gene therapy via intravitreal injection, offering enhanced efficiency and a minimized immune response.
    Keywords:  AAV2; capsid screening; cell-penetrating peptide; intravitreal injection; photoreceptor
    DOI:  https://doi.org/10.1016/j.omtm.2025.101426
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