bims-cepepe 2024-04-21 papers

bims-cepepe

Biomed News

on Cell-penetrating peptides

Issue of 2024–04–21
twelve papers selected by
Henry Lamb, Queensland University of Technology

Structure and Activity of Reconstructed Pseudo-Ancestral Cyclotides.
Incorporation of Three Extracyclic Arginine Residues into a Melanocortin Macrocyclic Agonist (c[Pro-His-DPhe-Arg-Trp-Dap-Lys(Arg-Arg-Arg-Ac)-DPro]) Decreases Food Intake When Administered Intrathecally or Subcutaneously Compared to a Macrocyclic Ligand Lacking Extracyclic Arginine Residues (c[Pro-His-DPhe-Arg-Trp-Dap-Ala-DPro)].
The use of a selective, nontoxic dual-acting peptide for breast cancer patients with brain metastasis.
Genetically encoded libraries and spider venoms as emerging sources for crop protective peptides.
Cyclic Peptides from Graspetide Biosynthesis and Native Chemical Ligation.
Application of parallel artificial membrane permeability assay technique and chemometric modeling for blood-brain barrier permeability prediction of protein kinase inhibitors.
Prediction of blood-brain barrier penetrating peptides based on data augmentation with Augur.
Illuminating Neuropeptide Y Y4 Receptor Binding: Fluorescent Cyclic Peptides with Subnanomolar Binding Affinity as Novel Molecular Tools.
Surface Modification of PEEKs with Cyclic Peptides to Support Endothelialization and Antithrombogenicity.
Combined, yet separate: cocktails of carriers (not drugs) for actinium-225 α-particle therapy of solid tumors expressing moderate-to-low levels of targetable markers.
De novo purine metabolism is a metabolic vulnerability of cancers with low p16 expression.
Hi1a Improves Sensorimotor Deficit following Endothelin-1-Induced Stroke in Rats but Does Not Improve Functional Outcomes following Filament-Induced Stroke in Mice.

ChemMedChem. 2024 Apr 17. e202400124

Structure and Activity of Reconstructed Pseudo-Ancestral Cyclotides.

Zhihao Jiang, Yen-Hua Huang, Quentin Kaas, David J Craik, Conan K Wang.

  Cyclotides are cyclic peptides that are promising scaffolds for the design of drug candidates and chemical tools. However, despite there being hundreds of reported cyclotides, drug design studies have commonly focussed on a select few prototypic examples. Here, we explored whether ancestral sequence reconstruction could be used to generate new cyclotides for further optimization. We show that the reconstructed 'pseudo-ancestral' sequences, named Ancy-m (for the ancestral cyclotide of the Möbius sub-family) and Ancy-b (for the bracelet sub-family), have well-defined structures like their extant members, comprising the core structural feature of a cyclic cystine knot. This motif underpins efforts to re-engineer cyclotides for agrochemical and therapeutic applications. We further show that the reconstructed sequences are resistant to temperatures approaching boiling, bind to phosphatidyl-ethanolamine lipid bilayers at micromolar affinity, and inhibit the growth of insect cells at inhibitory concentrations in the micromolar range. Interestingly, the Ancy-b cyclotide had a higher oxidative folding yield than its comparator cyclotide cyO2, which belongs to the bracelet cyclotide subfamily known to be notoriously difficult to fold. Overall, this study provides new cyclotide sequences not yet found naturally that could be valuable starting points for the understanding of cyclotide evolution and for further optimization as drug leads.

Keywords:  ancestral sequence reconstruction; cyclic peptide; disulfide-rich peptide

DOI:  https://doi.org/10.1002/cmdc.202400124
ACS Pharmacol Transl Sci. 2024 Apr 12. 7(4): 1114-1125

Incorporation of Three Extracyclic Arginine Residues into a Melanocortin Macrocyclic Agonist (c[Pro-His-DPhe-Arg-Trp-Dap-Lys(Arg-Arg-Arg-Ac)-DPro]) Decreases Food Intake When Administered Intrathecally or Subcutaneously Compared to a Macrocyclic Ligand Lacking Extracyclic Arginine Residues (c[Pro-His-DPhe-Arg-Trp-Dap-Ala-DPro)].

Mark D Ericson, Katie T Freeman, Courtney M Larson, Jacob L Bouchard, Kristen John, Mary M Lunzer, Zoe M Koerperich, Carrie Haskell-Luevano.

Of the three Food and Drug Administration-approved melanocortin peptide drugs, two possess a cyclic scaffold, demonstrating that cyclized melanocortin peptides have therapeutic relevance. An extracyclic Arg residue, critical for pharmacological activity in the approved melanocortin cyclic drug setmelanotide, has also been demonstrated to increase the signal when fluorescently labeled cell-penetrating cyclic peptides are incubated with HeLa cells, with the maximal signal observed with three extracyclic Arg amino acids. Herein, a branching Lys residue was substituted into two macrocyclic melanocortin peptide agonists to incorporate 0-3 extracyclic Arg amino acids. Incorporation of the Arg residues resulted in equipotent or increased agonist potency at the mouse melanocortin receptors in vitro, suggesting that these substitutions were tolerated in the macrocyclic scaffolds. Further in vivo evaluation of one parent ligand (c[Pro-His-DPhe-Arg-Trp-Dap-Ala-Pro]) and the three Arg derivative (c[Pro-His-DPhe-Arg-Trp-Dap-Lys(Ac-Arg-Arg-Arg)-Pro)] demonstrated that the three Arg derivative further decreased food intake compared to the parent macrocycle when the compounds were administered either via intrathecal injection or subcutaneous dosing. This suggests that three extracyclic Arg amino acids may be beneficial in the design of cyclic melanocortin ligands and that in vitro pharmacological profiling may not predict the in vivo efficacy of melanocortin ligands.

DOI: https://doi.org/10.1021/acsptsci.4c00011
Biomed Pharmacother. 2024 Apr 12. pii: S0753-3322(24)00457-8. [Epub ahead of print]174 116573

The use of a selective, nontoxic dual-acting peptide for breast cancer patients with brain metastasis.

Marco Cavaco, Clara Pérez-Peinado, Javier Valle, Ruben D M Silva, Lurdes Gano, João D G Correia, David Andreu, Miguel A R B Castanho, Vera Neves.

  Triple-negative breast cancer (TNBC) is an aggressive subtype characterized by the absence of commonly targeted receptors. Unspecific chemotherapy is currently the main therapeutic option, with poor results. Another major challenge is the frequent appearance of brain metastasis (BM) associated with a significant decrease in patient overall survival. The treatment of BM is even more challenging due to the presence of the blood-brain barrier (BBB). Here, we present a dual-acting peptide (PepH3-vCPP2319) designed to tackle TNBC/BM, in which a TNBC-specific anticancer peptide (ACP) motif (vCPP2319) is joined to a BBB peptide shuttle (BBBpS) motif (PepH3). PepH3-vCPP2319 demonstrated selectivity and efficiency in eliminating TNBC both in monolayers (IC50≈5.0 µM) and in spheroids (IC50≈25.0 µM), with no stringent toxicity toward noncancerous cell lines and red blood cells (RBCs). PepH3-vCPP2319 was also able to cross the BBB in vitro and penetrate the brain in vivo, and was stable in serum with a half-life above 120 min. Tumor cell-peptide interaction is fast, with quick peptide internalization via clathrin-mediated endocytosis without membrane disruption. Upon internalization, the peptide is detected in the nucleus and the cytoplasm, indicating a multi-targeted mechanism of action that ultimately induces irreversible cell damage and apoptosis. In conclusion, we have designed a dual-acting peptide capable of brain penetration and TNBC cell elimination, thus expanding the drug arsenal to fight this BC subtype and its BM.

Keywords:  Anticancer peptides; BBB peptide shuttle; Blood-brain barrier; Brain metastasis; Cell-penetrating peptides; PepH3; Triple-negative breast cancer; vCPP2319

DOI:  https://doi.org/10.1016/j.biopha.2024.116573
J Pept Sci. 2024 Apr 16. e3600

Genetically encoded libraries and spider venoms as emerging sources for crop protective peptides.

Elena Marone Fassolo, Shaodong Guo, Yachen Wang, Stefano Rosa, Volker Herzig.

  Agricultural crops are targeted by various pathogens (fungi, bacteria, and viruses) and pests (herbivorous arthropods). Antimicrobial and insecticidal peptides are increasingly recognized as eco-friendly tools for crop protection due to their low propensity for resistance development and the fact that they are fully biodegradable. However, historical challenges have hindered their development, including poor stability, limited availability, reproducibility issues, high production costs, and unwanted toxicity. Toxicity is a primary concern because crop-protective peptides interact with various organisms of environmental and economic significance. This review focuses on the potential of genetically encoded peptide libraries like the use of two-hybrid-based methods for antimicrobial peptides identification and insecticidal spider venom peptides as two main approaches for targeting plant pathogens and pests. We discuss some key findings and challenges regarding the practical application of each strategy. We conclude that genetically encoded peptide library- and spider venom-derived crop protective peptides offer a sustainable and environmentally responsible approach for addressing modern crop protection needs in the agricultural sector.

Keywords:  antimicrobial peptides; bioinsecticides; crop protection; genetically encoded peptide libraries; pesticides; spider venom peptides; two‐hybrid assays

DOI:  https://doi.org/10.1002/psc.3600
J Am Chem Soc. 2024 Apr 18.

Cyclic Peptides from Graspetide Biosynthesis and Native Chemical Ligation.

Brian Choi, Arthur Acuña, A James Link.

The ribosomally synthesized and post-translationally modified peptide (RiPP) superfamily of natural products includes many examples of cyclic peptides with diverse macrocyclization chemistries. The graspetides, one family of macrocyclized RiPPs, harbor side chain-side chain ester or amide linkages. We recently reported the structure and biosynthesis of the graspetide pre-fuscimiditide, a 22-amino-acid (aa) peptide with two ester cross-links forming a stem-loop structure. These cross-links are introduced by a single graspetide synthetase, the ATP-grasp enzyme ThfB. Here we show that ThfB can also catalyze the formation of amide or thioester cross-links in prefuscimiditide, with thioester formation being especially efficient. We further show that upon proteolysis to reveal an N-terminal cysteine residue, the thioester-linked peptide rapidly and quantitatively rearranges via native chemical ligation into an isopeptide-bonded head-to-tail cyclic peptide. The solution structure of this rearranged peptide was determined by using 2D NMR spectroscopy experiments. Our methodology offers a straightforward recombinant route to head-to-tail cyclic peptides.

DOI: https://doi.org/10.1021/jacs.4c02745
Future Med Chem. 2024 Apr 19.

Application of parallel artificial membrane permeability assay technique and chemometric modeling for blood-brain barrier permeability prediction of protein kinase inhibitors.

Milan Jovanović, Milica Radan, Marija Čarapić, Nenad Filipović, Katarina Nikolic, Milkica Crevar.

  Aim: This study aims to investigate the passive diffusion of protein kinase inhibitors through the blood-brain barrier (BBB) and to develop a model for their permeability prediction. Materials & methods: We used the parallel artificial membrane permeability assay to obtain logPe values of each of 34 compounds and calculated descriptors for these structures to perform quantitative structure-property relationship modeling, creating different regression models. Results: The logPe values have been calculated for all 34 compounds. Support vector machine regression was considered the most reliable, and CATS2D_09_DA, CATS2D_04_AA, B04[N-S] and F07[C-N] descriptors were identified as the most influential to passive BBB permeability. Conclusion: The quantitative structure-property relationship-support vector machine regression model that has been generated can serve as an efficient method for preliminary screening of BBB permeability of new analogs.

Keywords:  ANN; MLR; PAMPA-BBB; QSPR; SVM; protein kinase inhibitors

DOI:  https://doi.org/10.4155/fmc-2023-0390
BMC Biol. 2024 Apr 19. 22(1): 86

Prediction of blood-brain barrier penetrating peptides based on data augmentation with Augur.

Zhi-Feng Gu, Yu-Duo Hao, Tian-Yu Wang, Pei-Ling Cai, Yang Zhang, Ke-Jun Deng, Hao Lin, Hao Lv.

   BACKGROUND: The blood-brain barrier serves as a critical interface between the bloodstream and brain tissue, mainly composed of pericytes, neurons, endothelial cells, and tightly connected basal membranes. It plays a pivotal role in safeguarding brain from harmful substances, thus protecting the integrity of the nervous system and preserving overall brain homeostasis. However, this remarkable selective transmission also poses a formidable challenge in the realm of central nervous system diseases treatment, hindering the delivery of large-molecule drugs into the brain. In response to this challenge, many researchers have devoted themselves to developing drug delivery systems capable of breaching the blood-brain barrier. Among these, blood-brain barrier penetrating peptides have emerged as promising candidates. These peptides had the advantages of high biosafety, ease of synthesis, and exceptional penetration efficiency, making them an effective drug delivery solution. While previous studies have developed a few prediction models for blood-brain barrier penetrating peptides, their performance has often been hampered by issue of limited positive data.
RESULTS: In this study, we present Augur, a novel prediction model using borderline-SMOTE-based data augmentation and machine learning. we extract highly interpretable physicochemical properties of blood-brain barrier penetrating peptides while solving the issues of small sample size and imbalance of positive and negative samples. Experimental results demonstrate the superior prediction performance of Augur with an AUC value of 0.932 on the training set and 0.931 on the independent test set.
CONCLUSIONS: This newly developed Augur model demonstrates superior performance in predicting blood-brain barrier penetrating peptides, offering valuable insights for drug development targeting neurological disorders. This breakthrough may enhance the efficiency of peptide-based drug discovery and pave the way for innovative treatment strategies for central nervous system diseases.

Keywords:  Blood–brain barrier; Data augmentation; Feature selection; Information gain; Machine learning; Penetrating peptides

DOI:  https://doi.org/10.1186/s12915-024-01883-4
ACS Pharmacol Transl Sci. 2024 Apr 12. 7(4): 1142-1168

Illuminating Neuropeptide Y Y4 Receptor Binding: Fluorescent Cyclic Peptides with Subnanomolar Binding Affinity as Novel Molecular Tools.

Jakob Gleixner, Sergei Kopanchuk, Lukas Grätz, Maris-Johanna Tahk, Tõnis Laasfeld, Santa Veikšina, Carina Höring, Albert O Gattor, Laura J Humphrys, Christoph Müller, Nataliya Archipowa, Johannes Köckenberger, Markus R Heinrich, Roger Jan Kutta, Ago Rinken, Max Keller.

The neuropeptide Y (NPY) Y4 receptor (Y4R), a member of the family of NPY receptors, is physiologically activated by the linear 36-amino acid peptide pancreatic polypeptide (PP). The Y4R is involved in the regulation of various biological processes, most importantly pancreatic secretion, gastrointestinal motility, and regulation of food intake. So far, Y4R binding affinities have been mostly studied in radiochemical binding assays. Except for a few fluorescently labeled PP derivatives, fluorescence-tagged Y4R ligands with high affinity have not been reported. Here, we introduce differently fluorescence-labeled (Sulfo-Cy5, Cy3B, Py-1, Py-5) Y4R ligands derived from recently reported cyclic hexapeptides showing picomolar Y4R binding affinity. With pKi values of 9.22-9.71 (radioligand competition binding assay), all fluorescent ligands (16-19) showed excellent Y4R affinity. Y4R saturation binding, binding kinetics, and competition binding with reference ligands were studied using different fluorescence-based methods: flow cytometry (Sulfo-Cy5, Cy3B, and Py-1 label), fluorescence anisotropy (Cy3B label), and NanoBRET (Cy3B label) binding assays. These experiments confirmed the high binding affinity to Y4R (equilibrium pKd: 9.02-9.9) and proved the applicability of the probes for fluorescence-based Y4R competition binding studies and imaging techniques such as single-receptor molecule tracking.

DOI: https://doi.org/10.1021/acsptsci.4c00013
Mater Today Commun. 2024 Jun;pii: 108664. [Epub ahead of print]39

Surface Modification of PEEKs with Cyclic Peptides to Support Endothelialization and Antithrombogenicity.

Emma R Young, Cameron Martin, Joseph Ribaudo, Xiaochao Xia, William R Moritz, Sarah Madira, Mohamed A Zayed, Justin M Sacks, Xiaowei Li.

  Synthetic polymers are often utilized in the creation of vascular devices, and need to possess specific qualities to prevent thrombosis. Traditional strategies for this include surface modification of vascular devices through covalent attachment of substrates such as heparin, antiplatelet agents, thrombolytic agents, or hydrophilic polymers. One promising prosthetic material is polyether ether ketone (PEEK), which is utilized in various FDA-approved medical devices, including vascular and endovascular prostheses. We hypothesized that surface modification of biologically inert PEEK can help improve its endothelial cell affinity and reduce its thrombogenic potential. To evaluate this, we developed an effective surface-modification approach with unique cyclic peptides, such as CCHGGVRLYC and CCREDVC. We treated the PEEK surface with ammonia plasma, which introduced amine groups onto the PEEK surface. Subsequently, we were able to conjugate these peptides to the plasma-modified PEEKs. We observed that cyclic CCHGGVRLYC conjugated on prosthetic PEEK not only supported endothelialization, but minimized platelet adhesion and activation. This technology can be potentially applied for in vivo vascular and endovascular protheses to enhance their utility and patency.

Keywords:  Cyclic Peptides; Endothelium; Polyether Ether Ketone; Surface Modification; Vascular Anastomosis

DOI:  https://doi.org/10.1016/j.mtcomm.2024.108664
Eur J Nucl Med Mol Imaging. 2024 Apr 20.

Combined, yet separate: cocktails of carriers (not drugs) for actinium-225 α-particle therapy of solid tumors expressing moderate-to-low levels of targetable markers.

Rajiv Ranjit Nair, Aprameya Prasad, Omkar Bhatavdekar, Aira Sarkar, Kathleen L Gabrielson, Stavroula Sofou.

  Alpha-particle radionuclide-antibody conjugates are being clinically evaluated against solid tumors even when they moderately express the targeted markers. At this limit of lower tumor-absorbed doses, to maintain efficacy, the few(er) intratumorally delivered alpha-particles need to traverse/hit as many different cancer cells as possible. We complement antibody-radioconjugate therapies with a separate nanocarrier delivering a fraction of the same total injected radioactivity to tumor regions geographically different than those affected by targeting antibodies; these carrier-cocktails collectively distribute the alpha-particle emitters better.
METHODS: The efficacy of actinium-225 delivered by our carrier-cocktails was assessed in vitro and on mice with orthotopic MDA-MB-436 and/or MDA-MB-231 triple-negative breast cancers and/or an ectopic BxPC3 pancreatic cancer. Cells/tumors were chosen to express low-to-moderate levels of HER1, as model antibody-targeted marker.
RESULTS: Independent of cell line, antibody-radioconjugates were most lethal on cell monolayers. On spheroids, with radii greater than alpha-particles' range, carrier-cocktails improved killing efficacy (p < 0.0500). Treatment with carrier-cocktails decreased the MDA-MB-436 and MDA-MB-231 orthotopic tumor volumes by 73.7% and 72.1%, respectively, relative to treatment with antibody-radioconjugates alone, at same total injected radioactivity; these carrier-cocktails completely eliminated formation of spontaneous metastases vs. 50% and 25% elimination in mice treated with antibody-radioconjugates alone. In BxPC3 tumor-bearing mice, carrier-cocktails increased the median survival to 25-26 days (in male-female animals) vs. 20-21 days of mice treated with antibody-radioconjugates alone (vs. 17 days for non-treated animals). Survival with carrier-cocktail radiotherapy was further prolonged by pre-injecting low-dose, standard-of-care, gemcitabine (p = 0.0390).
CONCLUSION: Tumor-agnostic carrier-cocktails significantly enhance the therapeutic efficacy of existing alpha-particle radionuclide-antibody treatments.

Keywords:  Actinium-225; Alpha-particle radiotherapy; Antibody-targeted radiotherapy; Intratumor distributions; Low targeted markers; Spheroids

DOI:  https://doi.org/10.1007/s00259-024-06710-0
Cancer Res Commun. 2024 Apr 16.

De novo purine metabolism is a metabolic vulnerability of cancers with low p16 expression.

Naveen Kumar Tangudu, Raquel Buj, Hui Wang, Jiefei Wang, Aidan R Cole, Apoorva Uboveja, Richard Fang, Amandine Amalric, Baixue Yang, Adam Chatoff, Claudia V Crispim, Peter Sajjakulnukit, Maureen A Lyons, Kristine Cooper, Nadine Hempel, Costas A Lyssiotis, Uma R Chandran, Nathaniel W Snyder, Katherine M Aird.

p16 is a tumor suppressor encoded by the CDKN2A gene whose expression is lost in ~50% of all human cancers. In its canonical role, p16 inhibits the G1-S phase cell cycle progression through suppression of cyclin dependent kinases. Interestingly, p16 also has roles in metabolic reprogramming, and we previously published that loss of p16 promotes nucleotide synthesis via the pentose phosphate pathway. However, the broader impact of p16/CDKN2A loss on other nucleotide metabolic pathways and potential therapeutic targets remains unexplored. Using CRISPR KO libraries in isogenic human and mouse melanoma cell lines, we determined several nucleotide metabolism genes essential for the survival of cells with loss of p16/CDKN2A. Consistently, many of these genes are upregulated in melanoma cells with p16 knockdown or endogenously low CDKN2A expression. We determined that cells with low p16/CDKN2A expression are sensitive to multiple inhibitors of de novo purine synthesis, including anti-folates. Finally, tumors with p16 knockdown were more sensitive to the anti-folate methotrexate in vivo than control tumors. Together, our data provide evidence to reevaluate the utility of these drugs in patients with p16/CDKN2Alow tumors as loss of p16/CDKN2A may provide a therapeutic window for these agents.

DOI: https://doi.org/10.1158/2767-9764.CRC-23-0450
ACS Pharmacol Transl Sci. 2024 Apr 12. 7(4): 1043-1054

Hi1a Improves Sensorimotor Deficit following Endothelin-1-Induced Stroke in Rats but Does Not Improve Functional Outcomes following Filament-Induced Stroke in Mice.

Adriana Knezic, Elena Budusan, Natalie J Saez, Brad R S Broughton, Lachlan D Rash, Glenn F King, Robert E Widdop, Claudia A McCarthy.

Activation of acid-sensing ion channel 1a (ASIC1a) plays a major role in mediating acidosis-induced neuronal injury following a stroke. Therefore, the inhibition of ASIC1a is a potential therapeutic avenue for the treatment of stroke. Venom-peptide Hi1a, a selective and highly potent ASIC1a inhibitor, reduces the infarct size and functional deficits when injected into the brain after stroke in rodents. However, its efficacy when administered using a clinically relevant route of administration remains to be established. Therefore, the current investigation aims to examine the efficacy of systemically administered Hi1a, using two different models of stroke in different species. Mice were subjected to the filament model of middle cerebral artery occlusion (MCAO) and treated with Hi1a systemically using either a single- or multiple-dosing regimen. 24 h poststroke, mice underwent functional testing, and the brain infarct size was assessed. Rats were subjected to endothelin-1 (ET-1)-induced MCAO and treated with Hi1a intravenously 2 h poststroke. Rats underwent functional tests prior to and for 3 days poststroke, when infarct volume was assessed. Mice receiving Hi1a did not show any improvements in functional outcomes, despite a trend toward reduced infarct size. This trend for reduced infarct size in mice was consistent regardless of the dosing regimen. There was also a trend toward lower infarct size in rats treated with Hi1a. More specifically, Hi1a reduced the amount of damage occurring within the somatosensory cortex, which was associated with an improved sensorimotor function in Hi1a-treated rats. Thus, this study suggests that Hi1a or more brain-permeable ASIC1a inhibitors are a potential stroke treatment.

DOI: https://doi.org/10.1021/acsptsci.3c00328