Search Results (27)

Peptide–drug conjugates (PDCs) have emerged as a next-generation therapeutic platform, combining the target specificity of peptides with the pharmacological potency of small-molecule drugs. As an evolution beyond antibody–drug conjugates (ADCs), PDCs offer distinct advantages, including enhanced cellular permeability, improved drug selectivity, and versatile design flexibility. This review provides a comprehensive analysis of the fundamental components of PDCs, including homing peptide selection, linker engineering, and payload optimization, alongside strategies to address their inherent challenges, such as stability, bioactivity, and clinical translation barriers. Therapeutic applications of PDCs span oncology, infectious diseases, metabolic disorders, and emerging areas like COVID-19, with several conjugates advancing in clinical trials and achieving regulatory milestones. Innovations, including bicyclic peptides, supramolecular architectures, and novel linker technologies, are explored as promising avenues to enhance PDC design. Additionally, this review examines the clinical trajectory of PDCs, emphasizing their therapeutic potential and highlighting ongoing trials that exemplify their efficacy. By addressing limitations and leveraging emerging advancements, PDCs hold immense promise as targeted therapeutics capable of addressing complex disease states and driving progress in precision medicine. Full article

Background/Objectives: Integrin α_Vβ₃ plays a crucial role in tumor angiogenesis and cancer progression, making it a key target for radiolabeled probes used in imaging and therapy. A previously developed probe, [¹²⁵I]bcRGD, exhibited high selectivity for α_Vβ₃ but limited tumor accumulation due to rapid blood clearance. This study aimed to address this issue through two strategies: (1) conjugating albumin-binding molecules to enhance systemic circulation and (2) dimerizing RGD peptides to improve binding affinity via multivalency effects. Methods: Three [¹²⁵I]bcRGD derivatives were synthesized: [¹²⁵I]bcRGD_pal (with palmitic acid), [¹²⁵I]bcRGD_iba (with 4-(p-iodophenyl)butyric acid), and [¹²⁵I]bcRGD_dimer (a dimeric bicyclic RGD peptide). Their physicochemical properties, α_Vβ₃-selectivity, albumin-binding capacity, and biodistribution were assessed in vitro and in vivo using tumor-bearing mice. Tumor models included α_Vβ₃-high U-87 MG and α_Vβ₃-low A549 xenografts. Results: [¹²⁵I]bcRGD_pal and [¹²⁵I]bcRGD_iba exhibited prolonged blood retention (30-fold and 55-fold vs. [¹²⁵I]bcRGD, respectively) and increased tumor accumulation (3.9% ID/g and 3.6% ID/g at 2 h, respectively). Despite improved systemic circulation, tumor-to-blood ratios remained low (<1), indicating limited tumor retention. [¹²⁵I]bcRGD_dimer achieved significantly greater tumor accumulation (4.2% ID/g at 2 h) and favorable tumor-to-blood (22) and tumor-to-muscle (14) ratios, with a 5.4-fold higher uptake in U-87 MG tumors compared to A549 tumors. Conclusions: Dimerization was more effective than albumin binding in enhancing bcRGD’s tumor-targeting potential. The dimeric probe demonstrated improved tumor accumulation, favorable pharmacokinetics, and preserved integrin selectivity. These findings provide a foundation for further structural optimization of bicyclic RGD peptides for integrin α_Vβ₃-targeted imaging and therapy applications. Full article

This work explores the self-assembly and optical properties of a novel chiral, aromatic-rich Boc-Phe-Phe dipeptide derivative functionalized with a benzothiazole bicyclic ring that forms supramolecular structures. Leveraging the well-known self-assembling capabilities of diphenylalanine dipeptides, this modified derivative introduces a heterocyclic benzothiazole unit that significantly enhances the fluorescence of the resulting nanostructures. The derivative’s rich aromatic character drives the formation of supramolecular structures through self-organization mechanisms influenced by quantum confinement. By adjusting the solvent system, the nanostructures exhibit tunable morphologies, ranging from nanospheres to nanobelts. The nonlinear optical properties of these self-assembled structures were studied and an estimated $d_{eff}$ of ~0.9 pm/V was obtained, which is comparable to that reported for the highly aromatic triphenylalanine peptide. Full article

Background: Ribosomally synthesized and post-translationally modified peptides (RiPPs) are a growing class of natural products biosynthesized from a genetically encoded precursor peptide. RiPPs have attracted attention for the ability to generate and screen libraries of these compounds for useful biological activities. To facilitate this screening, it is useful to be able to do so with the leader peptide still present. We assessed the suitability of the microviridin family for these screening experiments by determining their activity with the leader peptide still present. Methods: Modified precursor peptides with the leader present were heterologously expressed in Escherichia coli. Their ability to inhibit elastase was tested with a fluorogenic substrate. HPLC was used to monitor degradation of the modified precursor peptides by elastase. SDS-PAGE was used to determine the ability of immobilized modified precursor peptide to pull down elastase. Results: We found that the fully modified precursor peptide of microviridin B can inhibit the serine protease elastase with a low nanomolar IC₅₀, and that the fully modified precursor with an N-terminal His-tag can mediate interactions between elastase and Ni-NTA resin, all indicating leader peptide removal is not necessary for microviridins to bind their target proteases. Additionally, we found that a bicyclic variant was able to inhibit elastase with the leader peptide still present, although with a roughly 100-fold higher IC₅₀ and being subject to hydrolysis by elastase. Conclusions: These results open a pathway to screening libraries of microviridin variants for improved protease inhibition or other characteristics that can serve as, or as inspirations for, new pharmaceuticals. Full article

Kappa opioid receptor (KOR) antagonists have potential therapeutic applications in the treatment of stress-induced relapse to substance abuse and mood disorders. The dynorphin A analog arodyn (Ac[Phe^1,2,3,Arg⁴,D-Ala⁸]dynorphin A-(1–11)-NH₂) exhibits potent and selective kappa opioid receptor antagonism. Multiple cyclizations in longer peptides, such as dynorphin and its analogs, can extend the conformational constraint to additional regions of the peptide beyond what is typically constrained by a single cyclization. Here, we report the design, synthesis, and pharmacological evaluation of a bicyclic arodyn analog with two constraints in the opioid peptide sequence. The peptide, designed based on structure–activity relationships of monocyclic arodyn analogs, was synthesized by solid-phase peptide synthesis and cyclized by sequential ring-closing metathesis (RCM) in the C- and N-terminal sequences. Molecular modeling studies suggest similar interactions of key aromatic and basic residues in the bicyclic peptide with KOR as found in the cryoEM structure of KOR-bound dynorphin, despite substantial differences in the backbone conformations of the two peptides. The bicyclic peptide’s affinities at KOR and mu opioid receptors (MOR) were determined in radioligand binding assays, and its KOR antagonism was determined in the [³⁵S]GTPγS assay in KOR-expressing cells. The bicyclic analog retains KOR affinity and selectivity (K_i = 26 nM, 97-fold selectivity over MOR) similar to arodyn and exhibits potent KOR antagonism in the dynorphin-stimulated [³⁵S]GTPγS assay. This bicyclic peptide represents a promising advance in preparing cyclic opioid peptide ligands and opens avenues for the rational design of additional bicyclic opioid peptide analogs. Full article

Bicyclic peptides have attracted the interest of pharmaceutical companies because of their remarkable properties, putting them on a new path in medicine. Their conformational rigidity improves proteolytic stability and leads to rapid penetration into tissues via any possible route of administration. Moreover, elimination of renal metabolism is of great importance, for example, for people with a history of liver diseases. In addition, each ring can function independently, making bicyclic peptides extremely versatile molecules for further optimization. In this paper, we compared the potentiometric and spectroscopic properties studied by UV–vis, MCD, and EPR of four synthetic analogues of the bi-cyclic peptide c(PKKHP-c(CFWKTC)-PKKH) (BCL). In particular, we correlated the structural and spectral properties of complexes with coordinating abilities toward Cu(II) ions of MCL1 (Ac-PKKHPc(CFWKTC)PKKH-NH₂) that contains the unbinding cycle and N- and C-terminal linear parts with two histidine residues, one per part; two monocyclic ligands containing one histidine residue, both in the N-terminal position, i.e., MCL2 (Ac-PKKHPc(CFWKTC)PKKS-NH₂) and in the C-terminal position, i.e., MCL3 (Ac-PKKSPc(CFWKTC)PKKH-NH₂), respectively; and the linear structure LNL (Ac-PKKHPSFWKTSPKKH-NH₂). Potentiometric results have shown that the bicyclic structure promotes the involvement of the side chain imidazole donors in Cu(II) binding. On the other hand, the results obtained for the mono-cyclic analogues lead to the conclusion that the coordination of the histidine moiety as an anchoring group is promoted by its location in the peptide sequence further from the nonbinding cycle, strongly influencing the involvement of the amide donors in Cu(II) coordination. Full article

Recombinant peptide synthesis allows for large-scale production of peptides with therapeutic potential. However, access to dicarba peptidomimetics via sidechain-deprotected sequences becomes challenging with exposed Lewis basicity presented by amine and sulfur-containing residues. Presented here is a combination of strategies which can be used to deactivate coordinative residues and achieve high-yielding Ru-catalyzed ring-closing metathesis. The chemistry is exemplified using α-conotoxin EpI, a native bicyclic disulfide-containing sequence isolated from the marine conesnail Conus episcopatus. Replacement of the loop I disulfide with E/Z–dicarba bridges was achieved with high conversion via solution-phase ring-closing metathesis of the unprotected linear peptide after simple chemoselective oxidation and ion-exchange masking of problematic functionality. Metathesis was also attempted in green solvent choices to further improve the sustainability of dicarba peptide synthesis. Full article

Novel monoterpene-based hydroxamic acids of two structural types were synthesized for the first time. The first type consisted of compounds with a hydroxamate group directly bound to acyclic, monocyclic and bicyclic monoterpene scaffolds. The second type included hydroxamic acids connected with the monoterpene moiety through aliphatic (hexa/heptamethylene) or aromatic linkers. An in vitro analysis of biological activity demonstrated that some of these molecules had powerful HDAC6 inhibitory activity, with the presence of a linker area in the structure of compounds playing a key role. In particular, it was found that hydroxamic acids containing a hexa- and heptamethylene linker and (-)-perill fragment in the Cap group exhibit excellent inhibitory activity against HDAC6 with IC₅₀ in the submicromolar range from 0.56 ± 0.01 µM to 0.74 ± 0.02 µM. The results of the study of antiradical activity demonstrated the presence of moderate ability for some hydroxamic acids to scavenge 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2ROO^• radicals. The correlation coefficient between the DPPH radical scavenging activity and oxygen radical absorbance capacity (ORAC) value was R² = 0.8400. In addition, compounds with an aromatic linker based on para-substituted cinnamic acids, having a monocyclic para-menthene skeleton as a Cap group, 35a, 38a, 35b and 38b, demonstrated a significant ability to suppress the aggregation of the pathological β-amyloid peptide 1-42. The 35a lead compound with a promising profile of biological activity, discovered in the in vitro experiments, demonstrated neuroprotective effects on in vivo models of Alzheimer’s disease using 5xFAD transgenic mice. Together, the results obtained demonstrate a potential strategy for the use of monoterpene-derived hydroxamic acids for treatment of various aspects of Alzheimer’s disease. Full article

CD59 is an abundant immuno-regulatory human protein that protects cells from damage by inhibiting the complement system. CD59 inhibits the assembly of the Membrane Attack Complex (MAC), the bactericidal pore-forming toxin of the innate immune system. In addition, several pathogenic viruses, including HIV-1, escape complement-mediated virolysis by incorporating this complement inhibitor in their own viral envelope. This makes human pathogenic viruses, such as HIV-1, not neutralised by the complement in human fluids. CD59 is also overexpressed in several cancer cells to resist the complement attack. Consistent with its importance as a therapeutical target, CD59-targeting antibodies have been proven to be successful in hindering HIV-1 growth and counteracting the effect of complement inhibition by specific cancer cells. In this work, we make use of bioinformatics and computational tools to identify CD59 interactions with blocking antibodies and to describe molecular details of the paratope–epitope interface. Based on this information, we design and produce paratope-mimicking bicyclic peptides able to target CD59. Our results set the basis for the development of antibody-mimicking small molecules targeting CD59 with potential therapeutic interest as complement activators. Full article

Dithiolopyrrolone antibiotics are well known for their outstanding biological activities, and their biosynthesis has been studied vigorously. However, the biosynthesis mechanism of the characteristic bicyclic scaffold is still unknown after years of research. To uncover this mechanism, a multi-domain non-ribosomal peptide synthase DtpB from the biosynthetic gene cluster of thiolutin was selected as an object to study. We discovered that its adenylation domain not only recognized and adenylated cysteine, but also played an essential role in the formation of the peptide bond. Notably, an eight-membered ring compound was also discovered as an intermediate during the formation of the bicyclic structure. Based on these findings, we propose a new mechanism for the biosynthesis of the bicyclic scaffold of dithiolopyrrolones, and unveil additional functions of the adenylation domain. Full article

In the era of liquid biopsies, the reliable and cost-effective detection and screening of cancer biomarkers has become of fundamental importance, thus paving the way for the advancement of research in the field of point-of-care testing and the development of new methodologies and technologies. Indeed, the latter ones can help designing advanced diagnostic tools that can offer portability, ease of use with affordable production and operating costs. In this respect, impedance-based biosensing platforms might represent an attractive alternative. In this work, we describe a proof-of-concept study aimed at designing portable impedimetric biosensors for the monitoring of human urokinase-type plasminogen activator (h-uPA) cancer biomarker by employing small synthetic receptors. Aberrant levels of h-uPA were correlated with different types of cancers. Herein, we report the use of two bicyclic peptides (P₂ and P₃) which have been engineered to bind h-uPA with high affinity and exquisite specificity. The synthetic receptors were immobilized via biotin-streptavidin chemistry on the surface of commercial screen-printed electrodes. The impedimetric changes in the electrode/solution interface upon incubation of spiked h-uPA samples in the presence of a redox probe were followed via electrochemical impedance spectroscopy. The P₃-based impedimetric assay showed the best outcomes in terms of dynamic range and linearity (0.01–1 μg mL⁻¹) and sensitivity (LOD = 9 ng mL⁻¹). To fully assess the performances of P₃ over P₂, and to compare the label-free architecture vs. labelled architecture, a voltammetric assay was also developed. Full article

Antimicrobial peptides (AMPs) hold promise as novel therapeutics in the fight against multi-drug-resistant pathogens. Cathelicidin-PY (NH₂-RKCNFLCKLKEKLRTVITSHIDKVLRPQG-COOH) is a 29-residue disulfide-cyclised antimicrobial peptide secreted as an innate host defence mechanism by the frog Paa yunnanensis (PY) and reported to possess broad-spectrum antibacterial and antifungal properties, exhibiting low cytotoxic and low hemolytic activity. Herein, we detail the total synthesis of cathelicidin-PY using an entirely on-resin synthesis, including assembly of the linear sequence by rapid flow Fmoc-SPPS and iodine-mediated disulfide bridge formation. By optimising a synthetic strategy to prepare cathelicidin-PY, this strategy was subsequently adapted to prepare a bicyclic head-to-tail cyclised derivative of cathelicidin-PY. The structure-activity relationship (SAR) of cathelicidin-PY with respect to the N-terminally positioned disulfide was further probed by preparing an alanine-substituted linear analogue and a series of lactam-bridged peptidomimetics implementing side chain to side chain cyclisation. The analogues were investigated for antimicrobial activity, secondary structure by circular dichroism (CD), and stability in human serum. Surprisingly, the disulfide bridge emerged as non-essential to antimicrobial activity and secondary structure but was amenable to synthetic modification. Furthermore, the synthetic AMP and multiple analogues demonstrated selective activity towards Gram-negative pathogen E. coli in physiologically relevant concentrations of divalent cations. Full article

New strategies facilitate the design of cyclic peptides which can penetrate the brain. We have designed a bicyclic peptide, OL-CTOP, composed of the sequences of a selective μ-opioid receptor antagonist, CTOP (f-cyclo(CYwOTX)T) (X = penicillamine, Pen; O = ornithine) and odorranalectin, OL (YASPK-cyclo(CFRYPNGVLAC)T), optimized its solid-phase synthesis and demonstrated its ability for nose-to-brain delivery and in vivo activity. The differences in reactivity of Cys and Pen thiol groups protected with trityl and/or acetamidomethyl protecting groups toward I₂ in different solvents were exploited for selective disulfide bond formation on the solid phase. Both the single step and the sequential strategy applied to macrocyclization reactions generated the desired OL-CTOP, with the sequential strategy yielding a large quantity and better purity of crude OL-CTOP. Importantly, intranasally (i.n.s.) administered OL-CTOP dose-dependently antagonized the analgesic effect of morphine administered to mice through the intracerebroventricular route and prevented morphine-induced respiratory depression. In summary, the results demonstrate the feasibility of our solid-phase synthetic strategy for the preparation of the OL-CTOP bicyclic peptide containing two disulfide bonds and reveal the potential of odorranalectin for further modifications and the targeted delivery to the brain. Full article

Myometrium plays critical roles in multiple processes such as embryo spacing through peristalsis during mouse implantation, indicating vital roles of smooth muscle in the successful establishment and quality of implantation. Actin, a key element of cytoskeleton structure, plays an important role in the movement and contraction of smooth muscle cells (SMCs). However, the function of peri-implantation uterine smooth muscle and the regulation mechanism of muscle tension are still unclear. This study focused on the molecular mechanism of actin assembly regulation on implantation in smooth muscle. Phalloidin is a highly selective bicyclic peptide used for staining actin filaments (also known as F-actin). Phalloidin staining showed that F-actin gradually weakened in the CD-1 mouse myometrium from day 1 to day 4 of early pregnancy. More than 3 mice were studied for each group. Jasplakinolide (Jasp) used to inhibit F-actin depolymerization promotes F-actin polymerization in SMCs during implantation window and consequently compromises embryo implantation quality. Transcriptome analysis following Jasp treatment in mouse uterine SMCs reveals significant molecular changes associated with actin assembly. Tagln is involved in the regulation of the cell cytoskeleton and promotes the polymerization of G-actin to F-actin. Our results show that Tagln expression is gradually reduced in mouse uterine myometrium from day 1 to 4 of pregnancy. Furthermore, progesterone inhibits the expression of Tagln through the progesterone receptor. Using siRNA to knock down Tagln in day 3 SMCs, we found that phalloidin staining is decreased, which confirms the critical role of Tagln in F-actin polymerization. In conclusion, our data suggested that decreases in actin assembly in uterine smooth muscle during early pregnancy is critical to optimal embryo implantation. Tagln, a key molecule involved in actin assembly, regulates embryo implantation by controlling F-actin aggregation before implantation, suggesting moderate uterine contractility is conducive to embryo implantation. This study provides new insights into how the mouse uterus increases its flexibility to accommodate implanting embryos in the early stage of pregnancy. Full article

The development of peptide inhibitors against intracellular targets depends upon the dual challenge of achieving a high affinity and specificity for the target and maintaining cellular permeability for biological activity. Previous efforts to develop bicyclic peptides targeted to the Grb7 signalling protein implicated in HER2+ve cancer progression have resulted in improved affinity. However, these same peptides demonstrated a lowered activity due to their decreased ability to penetrate cell membranes. Here, we report the testing of a new series of bicyclic G7 peptides designed to possess improved bioactivity. We discovered that the incorporation of two amino acids (Phe-Pro, Phe-Trp or Phe-Arg) within the bicyclic peptide framework maintains an enhanced binding affinity for the Grb7-SH2 domain compared to that of the first-generation monocyclic peptide G7-18NATE. Structure determination using X-ray crystallography revealed that the mode of binding by the expanded bicyclic G7 peptide is analogous to that of G7-18NATE. Interestingly, while the bicyclic peptide containing Phe-Trp did not display the highest affinity for Grb7-SH2 in the series, it was the most potent inhibitor of HER2+ve SKBR3 breast cancer cell migration when coupled to Penetratin. Together, this demonstrates that peptide flexibility as well as the amino acid tryptophan can play important roles in the uptake of peptides into the cell. Full article