RESUMEN

Background and Purpose: Growth hormone-releasing hormone (GHRH) agonist, a 29-amino acid peptide, shows significant potential in treating myocardial infarction (MI) by aiding the repair of injured heart tissue. The challenge lies in the effective on-site delivery of GHRH agonist. This study explores the use of a targetable delivery system employing ROS-responsive PEG-PPS-PEG polymers to encapsulate and deliver GHRH agonist MR409 for enhanced therapeutic efficacy. Methods: We synthesized a self-assembling poly (ethylene glycol)-poly (propylene sulfide)-poly (ethylene glycol) polymer (PEG-PPS-PEG) amphiphilic polymer responsive to reactive oxygen species (ROS). The hydrophilic peptide GHRH agonist MR409 was encapsulated within these polymers to form nano PEG-PPS-PEG@MR409 vesicles (NPs). Cardiomyocyte apoptosis was induced under hypoxia and serum-free culture condition for 24 hours, and their production of ROS was detected by fluorescence dye staining. The cellular uptake of PEG-PPS-PEG@MR409 NPs was observed using fluorescence-labeled MR409. Targeting ability and therapeutic efficacy were evaluated using a mouse MI model. Results: PEG-PPS-PEG@MR409 NPs were efficiently internalized by cardiomyocytes, reducing ROS levels and apoptosis. These NPs exhibited superior targeting to the infarcted heart compared to naked MR409 peptide. With a reduced injection frequency (once every three days), PEG-PPS-PEG@MR409 NPs significantly promoted cardiac function recovery post-MI, matching the efficacy of daily MR409 injections. Conclusion: ROS-responsive PEG-PPS-PEG polymers provide a novel and effective platform for the targeted delivery of GHRH agonist peptides, improving cardiac function and offering a new approach for peptide therapy in MI treatment.

Asunto(s)

Infarto del Miocardio , Miocitos Cardíacos , Polietilenglicoles , Especies Reactivas de Oxígeno , Animales , Polietilenglicoles/química , Polietilenglicoles/farmacocinética , Polietilenglicoles/farmacología , Especies Reactivas de Oxígeno/metabolismo , Ratones , Infarto del Miocardio/tratamiento farmacológico , Miocitos Cardíacos/efectos de los fármacos , Modelos Animales de Enfermedad , Hormona Liberadora de Hormona del Crecimiento/agonistas , Hormona Liberadora de Hormona del Crecimiento/farmacocinética , Hormona Liberadora de Hormona del Crecimiento/administración & dosificación , Apoptosis/efectos de los fármacos , Sulfuros/química , Sulfuros/farmacocinética , Sulfuros/farmacología , Sulfuros/administración & dosificación , Péptidos/química , Péptidos/farmacología , Péptidos/farmacocinética , Péptidos/administración & dosificación , Masculino , Ratones Endogámicos C57BL

RESUMEN

Subcutaneous (SC) injection is a common route of administration for drug compounds with poor oral bioavailability. However, bioavailability is often variable and incomplete, and there is as yet no standard accepted medium for simulation of the human SC environment. In this work we evaluate a FRAP based method for quantitative determination of local self-diffusion coefficients within extracellular matrix (ECM) mimetic hydrogels, potentially useful as in vitro models for drug transport in the ECM after SC injection. Gels were made consisting of either agarose, cross-linked collagen (COL) and hyaluronic acid (HA) or cross-linked HA. The diffusivities of uncharged FITC-dextran (FD4), the highly charged poly-lysine (PLK20) and poly-glutamic acid (PLE20) as well as the GLP-1 analogue exenatide were determined within the gels using FRAP. The diffusion coefficients in uncharged agarose gels were in the range of free diffusion in PBS. The diffusivity of cationic PLK20 in gels containing anionic HA was substantially decreased due to strong electrostatic interactions. Peptide aggregation could be observed as immobile fractions in experiments with exenatide. We conclude that the FRAP method provides useful information of peptides' interactions and transport properties in hydrogel networks, giving insight into the mechanisms affecting absorption of drug compounds after subcutaneous injection.

Asunto(s)

Dextranos , Exenatida , Matriz Extracelular , Ácido Hialurónico , Hidrogeles , Péptidos , Hidrogeles/química , Difusión , Matriz Extracelular/metabolismo , Inyecciones Subcutáneas , Exenatida/farmacocinética , Exenatida/química , Exenatida/administración & dosificación , Ácido Hialurónico/química , Dextranos/química , Dextranos/farmacocinética , Péptidos/química , Péptidos/farmacocinética , Péptidos/administración & dosificación , Ácido Poliglutámico/química , Ácido Poliglutámico/análogos & derivados , Polilisina/química , Colágeno/química , Sefarosa/química , Fluoresceína-5-Isotiocianato/análogos & derivados , Fluoresceína-5-Isotiocianato/química , Fluoresceína-5-Isotiocianato/administración & dosificación , Fluoresceína-5-Isotiocianato/farmacocinética , Humanos

RESUMEN

Monoclonal antibodies (mAbs) have high binding specificity and affinity, making them attractive for treating brain diseases. However, their effectiveness is limited by poor blood-brain barrier (BBB) penetration and rapid central nervous system (CNS) clearance. Our group identified blood-brain barrier modulator (BBBM) peptides that improved mAb penetration across the BBB into the brain. In this study, we investigated the pharmacokinetics of a mAb delivered to the brain using BBBMs after intravenous (IV) administration and explored the impact of antibody format (size, neonatal Fc receptor (FcRn) binding, hyaluronic acid binding) on brain clearance following direct injection into the central nervous system (CNS) via intracerebroventricular (ICV) injection. IRDye800CW-labeled antibodies were administered into C57BL/6 mice via ICV or IV injection, and organ concentrations were measured after various time points. When a mAb was coadministered with a BBBM peptide, the permeation of mAb across the BBB was increased compared to mAb alone at early time points; however, the mAb was cleared within 2 h from the brain. ICV experiments revealed that an antibody Fab fragment had a higher brain exposure than a mAb, and that a Fab fused to a hyaluronic acid binding domain (Fab-VG1) showed remarkable improvement in brain exposure. These findings suggest that BBBMs and antibody format optimization may be promising strategies for enhancing brain retention of therapeutic antibodies.

Asunto(s)

Anticuerpos Monoclonales , Barrera Hematoencefálica , Encéfalo , Ratones Endogámicos C57BL , Receptores Fc , Animales , Barrera Hematoencefálica/metabolismo , Ratones , Encéfalo/metabolismo , Anticuerpos Monoclonales/farmacocinética , Anticuerpos Monoclonales/administración & dosificación , Receptores Fc/metabolismo , Antígenos de Histocompatibilidad Clase I/metabolismo , Antígenos de Histocompatibilidad Clase I/inmunología , Ácido Hialurónico/química , Masculino , Fragmentos Fab de Inmunoglobulinas , Péptidos/química , Péptidos/farmacocinética , Distribución Tisular

RESUMEN

The purpose of this study was to examine how the introduction of ibuprofen (IBU) affected tumor-targeting and biodistribution properties of 177Lu-labeled IBU-conjugated alpha-melanocyte-stimulating hormone peptides. The IBU was used as an albumin binder and conjugated to the DOTA-Lys moiety without or with a linker to yield DOTA-Lys(IBU)-GG-Nle-CycMSHhex {1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid-Lys(IBU)-Gly-Gly-Nle-c[Asp-His-DPhe-Arg-Trp-Lys]-CONH2}, DOTA-Lys(Asp-IBU)-GGNle-CycMSHhex, DOTA-Lys(Asn-IBU)-GGNle-CycMSHhex, and DOTA-Lys(Dab-IBU)-GGNle-CycMSHhex peptides. Their melanocortin-receptor 1 (MC1R) binding affinities were determined on B16/F10 melanoma cells first. Then the biodistribution of 177Lu-labeled peptides was determined on B16/F10 melanoma-bearing C57 mice at 2 h postinjection to choose the lead peptide for further examination. The full biodistribution and melanoma imaging properties of 177Lu-DOTA-Lys(Asp-IBU)-GGNle-CycMSHhex were further evaluated using B16/F10 melanoma-bearing C57 mice. DOTA-Lys(IBU)-GG-Nle-CycMSHhex, DOTA-Lys(Asp-IBU)-GGNle-CycMSHhex, DOTA-Lys(Asn-IBU)-GGNle-CycMSHhex, and DOTA-Lys(Dab-IBU)-GGNle-CycMSHhex displayed the IC50 values of 1.41 ± 0.37, 1.52 ± 0.08, 0.03 ± 0.01, and 0.58 ± 0.06 nM on B16/F10 melanoma cells, respectively. 177Lu-DOTA-Lys(Asp-IBU)-GGNle-CycMSHhex exhibited the lowest liver and kidney uptake among all four designed 177Lu peptides. Therefore, 177Lu-DOTA-Lys(Asp-IBU)-GGNle-CycMSHhex was further evaluated for its full biodistribution and melanoma imaging properties. The B16/F10 melanoma uptake of 177Lu-DOTA-Lys(Asp-IBU)-GGNle-CycMSHhex was 19.5 ± 3.12, 24.12 ± 3.35, 23.85 ± 2.08, and 10.80 ± 2.89% ID/g at 0.5, 2, 4, and 24 h postinjection, respectively. Moreover, 177Lu-DOTA-Lys(Asp-IBU)-GGNle-CycMSHhex could clearly visualize the B16/F10 melanoma lesions at 2 h postinjection. The conjugation of IBU with or without a linker to GGNle-CycMSHhex affected the MC1R binding affinities of the designed peptides. The charge of the linker played a key role in the liver and kidney uptake of 177Lu-Asp-IBU, 177Lu-Asn-IBU, and 177Lu-Dab-IBU. 177Lu-Asp-IBU exhibited higher tumor/liver and tumor/kidney uptake ratios than those of 177Lu-Asn-IBU and 177Lu-Dab-IBU, underscoring its potential evaluation for melanoma therapy in the future.

Asunto(s)

Ibuprofeno , Lutecio , alfa-MSH , Animales , Ratones , alfa-MSH/química , alfa-MSH/farmacocinética , Lutecio/química , Distribución Tisular , Ibuprofeno/química , Ibuprofeno/farmacocinética , Ibuprofeno/farmacología , Línea Celular Tumoral , Melanoma Experimental/tratamiento farmacológico , Melanoma Experimental/metabolismo , Ratones Endogámicos C57BL , Radioisótopos/química , Melanoma/metabolismo , Melanoma/tratamiento farmacológico , Albúminas/química , Radiofármacos/farmacocinética , Radiofármacos/química , Radiofármacos/farmacología , Péptidos/química , Péptidos/farmacocinética , Péptidos/farmacología , Femenino

RESUMEN

Brain-targeted drug delivery poses a great challenge due to the blood-brain barrier (BBB). In a previous study, we have developed a peptide-modified stealth liposome (SP-sLip) to enhance BBB penetration via the adsorption of apolipoproteins in plasma. SP is an 11-amino acid peptide derived from 25 to 35 of the Amyloid ß peptide (Aß1-42), which is a nature ligand of apolipoproteins. Although freshly prepared SP-sLip exhibited efficient brain targeting performance, it occured self-aggregation and instability in storage. In this study, we developed a D-peptide ligand according to the reverse sequence of SP with D-amino acids, known as DSP, to improve the stability in storage. Notably, DSP exhibited a reduced tendency for self-aggregation and improved stability in comparison to the SP peptide. Furthermore, compared to SP-sLip, DSP-modified sLip (DSP-sLip) demonstrated enhanced stability (>2 weeks), prolonged blood circulation (AUC increased 44.4%), reduced liver and spleen accumulation (reduced by 2.23 times and 1.86 times) with comparable brain-targeting efficiency. Similar to SP-sLip, DSP-sLip selectively adsorbed apolipoprotein A1, E, and J in the blood to form functionalized protein corona, thus crossing BBB via apolipoprotein receptor-mediated transcytosis. These findings underscored the importance of ligand stability in the in vitro and in vivo performance of brain-targeted liposomes, therefore paving the way for the design and optimization of efficient and stable nanocarriers.

Asunto(s)

Péptidos beta-Amiloides , Barrera Hematoencefálica , Encéfalo , Sistemas de Liberación de Medicamentos , Liposomas , Animales , Encéfalo/metabolismo , Ligandos , Péptidos beta-Amiloides/metabolismo , Péptidos beta-Amiloides/administración & dosificación , Barrera Hematoencefálica/metabolismo , Masculino , Péptidos/administración & dosificación , Péptidos/química , Péptidos/farmacocinética , Fragmentos de Péptidos/administración & dosificación , Fragmentos de Péptidos/farmacocinética , Fragmentos de Péptidos/química , Distribución Tisular , Apolipoproteínas/administración & dosificación , Apolipoproteínas/química , Ratones , Estabilidad de Medicamentos

RESUMEN

Nectin cell adhesion molecule 4 (Nectin-4) is overexpressed in various malignant tumors and has emerged as a promising target for tumor imaging. Bicyclic peptides, known for their conformational rigidity, metabolic stability, and membrane permeability, are ideal tracers for positron emission tomography (PET) imaging. In this study, we evaluated the feasibility of visualizing Nectin-4-positive tumors using radiolabeled bicyclic peptide derivatives and optimized the pharmacokinetics of radiotracers by introducing PEG chains of different lengths. Five PEGylated radiotracers radiolabeled with 68Ga3+ exhibited high radiochemical purity and stability. As the chain length increased, the Log D values decreased from -2.32 ± 0.13 to -2.50 ± 0.16, indicating a gradual increase in the hydrophilicity of the radiotracers. In vitro cell-binding assay results showed that the PEGylated bicyclic peptide exhibits nanomolar affinity, and blocking experiments confirmed the specific binding of the tracers to the Nectin-4 receptor. In vivo PET imaging and biodistribution studies in SW780 and 5637 xenograft mice showed that [68Ga]Ga-NOTA-PEG12-BP demonstrated optimal pharmacokinetics, characterized by rapid and good tumor uptake, faster background clearance, and improved tumor-to-tissue contrast. Finally, compared with 18F-FDG, PET imaging, in vivo blocking assays of [68Ga]Ga-NOTA-PEG12-BP and histological staining confirmed that specific tumor uptake was mediated by Nectin-4 receptors. The results indicated that [68Ga]Ga-NOTA-PEG12-BP was a promising PET radiotracer for Nectin-4 targeting, with applications for clinical translation.

Asunto(s)

Radioisótopos de Galio , Polietilenglicoles , Tomografía de Emisión de Positrones , Animales , Radioisótopos de Galio/farmacocinética , Radioisótopos de Galio/química , Ratones , Humanos , Tomografía de Emisión de Positrones/métodos , Polietilenglicoles/química , Distribución Tisular , Línea Celular Tumoral , Moléculas de Adhesión Celular/metabolismo , Ratones Desnudos , Radiofármacos/farmacocinética , Radiofármacos/química , Femenino , Péptidos/química , Péptidos/farmacocinética , Ratones Endogámicos BALB C , Nectinas

RESUMEN

Macrocyclic drugs can address an increasing range of molecular targets but enabling central nervous system (CNS) access to these drugs has been viewed as an intractable problem. We designed and synthesized a series of quinolinium-modified cyclosporine derivatives targeted to the mitochondrial cyclophilin D protein. Modification of the cation to enable greater delocalization was confirmed by x-ray crystallography of the cations. Critically, greater delocalization improved brain concentrations. Assessment of the compounds in preclinical assays and for pharmacokinetics identified a molecule JP1-138 with at least 20 times the brain levels of a non-delocalized compound or those reported for cyclosporine. Levels were maintained over 24 hours together with low hERG potential. The paradigm outlined here could have widespread utility in the treatment of CNS diseases.

Asunto(s)

Compuestos de Quinolinio , Animales , Humanos , Compuestos de Quinolinio/química , Compuestos de Quinolinio/farmacocinética , Ciclosporina/química , Ciclosporina/farmacocinética , Sistema Nervioso Central/metabolismo , Sistema Nervioso Central/efectos de los fármacos , Cristalografía por Rayos X , Péptidos/química , Péptidos/farmacocinética , Encéfalo/metabolismo , Encéfalo/efectos de los fármacos , Ratones

RESUMEN

Introduction: Glioblastoma multiforme (GBM), a highly invasive and prognostically challenging brain cancer, poses a significant hurdle for current treatments due to the existence of the blood-brain barrier (BBB) and the difficulty to maintain an effective drug accumulation in deep GBM lesions. Methods: We present a biomimetic nanoplatform with angiopep-2-modified macrophage membrane, loaded with indocyanine green (ICG) templated self-assembly of SN38 (AM-NP), facilitating active tumor targeting and effective blood-brain barrier penetration through specific ligand-receptor interaction. Results: Upon accumulation at tumor sites, these nanoparticles achieved high drug concentrations. Subsequent combination of laser irradiation and release of chemotherapy agent SN38 induced a synergistic chemo-photothermal therapy. Compared to bare nanoparticles (NPs) lacking cell membrane encapsulation, AM-NPs significantly suppressed tumor growth, markedly enhanced survival rates, and exhibited excellent biocompatibility with minimal side effects. Conclusion: This NIR-activatable biomimetic camouflaging macrophage membrane-based nanoparticles enhanced drug delivery targeting ability through modifications of macrophage membranes and specific ligands. It simultaneously achieved synergistic chemo-photothermal therapy, enhancing treatment effectiveness. Compared to traditional treatment modalities, it provided a precise, efficient, and synergistic method that might have contributed to advancements in glioblastoma therapy.

Asunto(s)

Barrera Hematoencefálica , Neoplasias Encefálicas , Liberación de Fármacos , Glioblastoma , Verde de Indocianina , Nanopartículas , Terapia Fototérmica , Glioblastoma/terapia , Glioblastoma/tratamiento farmacológico , Glioblastoma/metabolismo , Animales , Verde de Indocianina/química , Verde de Indocianina/farmacocinética , Verde de Indocianina/farmacología , Neoplasias Encefálicas/terapia , Neoplasias Encefálicas/tratamiento farmacológico , Neoplasias Encefálicas/metabolismo , Humanos , Línea Celular Tumoral , Ratones , Nanopartículas/química , Terapia Fototérmica/métodos , Barrera Hematoencefálica/efectos de los fármacos , Barrera Hematoencefálica/metabolismo , Irinotecán/farmacocinética , Irinotecán/química , Irinotecán/farmacología , Péptidos/química , Péptidos/farmacología , Péptidos/farmacocinética , Rayos Infrarrojos , Materiales Biomiméticos/química , Materiales Biomiméticos/farmacocinética , Materiales Biomiméticos/farmacología , Sistemas de Liberación de Medicamentos/métodos , Macrófagos/efectos de los fármacos , Macrófagos/metabolismo , Ratones Desnudos , Terapia Combinada/métodos

RESUMEN

Achieving therapeutic efficacy in protein replacement therapies requires sustaining pharmacokinetic (PK) profiles, while maintaining the bioactivity of circulating proteins. This is often achieved via PEGylation in protein-based therapies, but it remains challenging for proteins produced in vivo in mRNA-based therapies due to the lack of a suitable post-translational modification method. To address this issue, we integrated a genetically encoded zwitterionic polypeptide, EKP, into mRNA constructs to enhance the PK properties of product proteins. Composed of alternating glutamic acid (E), lysine (K), and proline (P), EKP exhibits unique superhydrophilic properties and low immunogenicity. Our results demonstrate that EKP fusion significantly extends the circulation half-life of proteins expressed from mRNA while preserving their bioactivity using human interferon alpha and Neoleukin-2/15 as examples. This EKP fusion technology offers a new approach to overcoming the current limitations in mRNA therapeutics and has the potential to significantly advance the development of mRNA-based protein replacement therapy.

Asunto(s)

Péptidos , ARN Mensajero , Humanos , ARN Mensajero/genética , ARN Mensajero/química , Péptidos/química , Péptidos/farmacocinética , Animales , Interferón-alfa/farmacocinética , Interferón-alfa/química , Interferón-alfa/genética , Ratones

RESUMEN

It is well known that the oral bioavailability of hydrophilic and macromolecular drugs is generally very poor due to their poor membrane permeability characteristics. Among these poorly absorbed drugs, peptide and protein drugs are typical poorly absorbed drugs which have low stability and poor permeability in the gastrointestinal tract. Consequently, the clinical administration of peptide and protein drugs is presently limited to administration by injection. However, such frequent administration subjects the patients to considerable pain, and there is also the possibility of the manifestation of serious side effects. Therefore, various approaches have been examined to overcome the poor absorption characteristics of these drugs. These approaches include (1) to use additives including absorption enhancers and protease inhibitors, (2) to modify the chemical structure of peptide and protein drugs, and (3) to apply dosage forms to these drugs, (4) to develop a novel administration method for these drugs that can serve as an alternative to oral and injection administration. We demonstrated that intestinal and transmucosal absorption of peptide and protein drugs could be improved by using these approaches. These approaches may give us useful basic information to improve the intestinal and transmucosal absorption of peptide and protein drugs.

Asunto(s)

Disponibilidad Biológica , Absorción Intestinal , Péptidos , Proteínas , Humanos , Péptidos/farmacocinética , Péptidos/administración & dosificación , Proteínas/administración & dosificación , Proteínas/farmacocinética , Inhibidores de Proteasas/administración & dosificación , Inhibidores de Proteasas/farmacocinética , Permeabilidad , Administración Oral , Mucosa Intestinal/metabolismo , Formas de Dosificación

RESUMEN

Different from most antiretroviral drugs that act as passive defenders to inhibit HIV-1 replication inside the host cell, virus inactivators can attack and inactivate HIV-1 virions without relying on their replication cycle. Herein, we describe the discovery of a hydrocarbon double-stapled helix peptide, termed D26. D26 is based on the HIV-1 gp41 protein lentiviral lytic peptide-3 motif (LLP3) sequence, which can efficiently inhibit HIV-1 infection and inactivate cell-free HIV-1 virions. It was noted that D26 was highly resistant to proteolytic degradation and exhibited a remarkably extended in vivo elimination half-life. Additionally, relative to its linear, nonstapled version, D26 exhibited much higher exposure in sanctuary sites for HIV-1. Amazingly, this lead compound also demonstrated detectable oral absorption. Thus, it can be concluded that D26 is a promising candidate for further development as a long-acting, orally applicable HIV-1 inactivator for the treatment of HIV-1 infection.

Asunto(s)

Fármacos Anti-VIH , Disponibilidad Biológica , Proteína gp41 de Envoltorio del VIH , VIH-1 , Péptidos , VIH-1/efectos de los fármacos , Fármacos Anti-VIH/farmacología , Fármacos Anti-VIH/química , Fármacos Anti-VIH/farmacocinética , Humanos , Animales , Administración Oral , Proteína gp41 de Envoltorio del VIH/metabolismo , Proteína gp41 de Envoltorio del VIH/química , Péptidos/química , Péptidos/farmacología , Péptidos/farmacocinética , Descubrimiento de Drogas , Infecciones por VIH/tratamiento farmacológico , Infecciones por VIH/virología , Semivida

RESUMEN

Bioactive peptides derived from food are promising health-promoting ingredients that can be used in functional foods and nutraceutical formulations. In addition to the potency towards the selected therapeutic target, the bioavailability of bioactive peptides is a major factor regarding clinical efficacy. We have previously shown that a low molecular weight peptide fraction (LMWPF) from poultry by-product hydrolysates possesses angiotensin-1-converting enzyme (ACE-1) and dipeptidyl-peptidase 4 (DPP4) inhibitory activities. The present study aimed to investigate the bioavailability of the bioactive peptides in the LMWPF. Prior to the investigation of bioavailability, a dipeptide YA was identified from this fraction as a dual inhibitor of ACE-1 and DPP4. Gastrointestinal (GI) stability and intestinal absorption of the bioactive peptides (i.e., YA as well as two previously reported bioactive dipeptides (VL and IY)) in the LMWPF were evaluated using the INFOGEST static in vitro digestion model and intestinal Caco-2 cell monolayer, respectively. Analysis of peptides after in vitro digestion confirmed that the dipeptides were resistant to the simulated GI conditions. After 4 hours of incubation, the concentration of the peptide from the apical side of the Caco-2 cell monolayer showed a significant decrease. However, the corresponding absorbed peptides were not detected on the basolateral side, suggesting that the peptides were not transported across the intestinal monolayer but rather taken up or metabolized by the Caco2 cells. Furthermore, when analyzing the gene expression of the Caco-2 cells upon peptide stimulation, a down-regulation of peptide transporters, the transcription factor CDX2, and the tight junction protein-1 (TJP1) was observed, suggesting the specific effects of the peptides on the Caco-2 cells. The study demonstrated that bioactive dipeptides found in the LMWPF were stable through in vitro GI digestion; however, the overall bioavailability may be hindered by inadequate uptake across the intestinal barrier.

Asunto(s)

Inhibidores de la Enzima Convertidora de Angiotensina , Dipeptidil Peptidasa 4 , Inhibidores de la Dipeptidil-Peptidasa IV , Absorción Intestinal , Hidrolisados de Proteína , Animales , Humanos , Inhibidores de la Enzima Convertidora de Angiotensina/química , Inhibidores de la Enzima Convertidora de Angiotensina/metabolismo , Inhibidores de la Enzima Convertidora de Angiotensina/farmacocinética , Inhibidores de la Enzima Convertidora de Angiotensina/farmacología , Disponibilidad Biológica , Células CACO-2 , Digestión , Dipéptidos/química , Dipéptidos/metabolismo , Dipéptidos/farmacocinética , Dipéptidos/farmacología , Dipeptidil Peptidasa 4/metabolismo , Inhibidores de la Dipeptidil-Peptidasa IV/química , Inhibidores de la Dipeptidil-Peptidasa IV/metabolismo , Inhibidores de la Dipeptidil-Peptidasa IV/farmacocinética , Inhibidores de la Dipeptidil-Peptidasa IV/farmacología , Tracto Gastrointestinal/metabolismo , Absorción Intestinal/efectos de los fármacos , Péptidos/química , Péptidos/metabolismo , Péptidos/farmacocinética , Péptidos/farmacología , Peptidil-Dipeptidasa A/metabolismo , Aves de Corral , Hidrolisados de Proteína/química , Hidrolisados de Proteína/farmacología

RESUMEN

Nanoparticles, in particular PEGylated, show great potential for in vivo brain targeted drug delivery. Nevertheless, how polyethylene glycol (PEG) length of nanoparticles affects their blood brain barrier (BBB) penetration or brain targeting is still unclear. In this study, we investigated the power of PEG chain-lengths (2, 3.4, 5, 10 kDa) in BBB penetration and brain targeting using Angiopep-2 peptide decorated liposomes. We found that PEG chain-length is critical, where the shorter PEG enabled the Angiopep-2 decorated liposomes to display more potent in vitro cell uptake via endocytosis. In contrast, their in vitro BBB penetration via transcytosis was much weaker relative to the liposomes with longer PEG chains, which result from their ineffective BBB exocytosis. Interestingly, the in vivo brain targeting aligns with the in vitro BBB penetration, as the long chain PEG-modified liposomes exerted superior brain accumulation both in normal or orthotropic glioblastoma (GBM) bearing mice, which could be ascribed to the combinational effect of prolonged circulation and enhanced BBB penetration of long chain PEG attached liposomes. These results demonstrate the crucial role of PEG length of nanoparticles for BBB penetration and brain targeting, providing guidance for PEG length selection in the design of nanocarrier for brain diseases treatment.

Asunto(s)

Barrera Hematoencefálica , Encéfalo , Liposomas , Péptidos , Polietilenglicoles , Animales , Polietilenglicoles/química , Barrera Hematoencefálica/metabolismo , Encéfalo/metabolismo , Péptidos/química , Péptidos/administración & dosificación , Péptidos/farmacocinética , Humanos , Línea Celular Tumoral , Neoplasias Encefálicas/tratamiento farmacológico , Neoplasias Encefálicas/metabolismo , Sistemas de Liberación de Medicamentos , Glioblastoma/tratamiento farmacológico , Glioblastoma/metabolismo , Glioblastoma/patología , Masculino , Ratones , Ratones Desnudos , Nanopartículas/química , Nanopartículas/administración & dosificación , Ratones Endogámicos BALB C

RESUMEN

PURPOSE: 68Ga-Trivehexin is an investigational PET radiopharmaceutical (NCT05799274) targeting αvß6-integrin for PET imaging of carcinomas. 177Lu-D0301 is a structurally related therapeutic peptide tetramer. However, it showed considerable kidney uptake in rodents, impeding clinical applicability. We therefore evaluated the impact of different kidney protection strategies on the biodistribution of both agents in normal and tumor-bearing mice. METHODS: Ex-vivo biodistribution of 68Ga-Trivehexin (90 min p.i.) and 177Lu-D0301 (90 min and 24 h p.i.) was determined in healthy C57BL/6N and H2009 (human lung adenocarcinoma) xenografted CB17-SCID mice without and with co-infusion of 100 µL of solutions containing 2.5% arginine + 2.5% lysine (Arg/Lys), 4% succinylated gelatin (gelofusine, gelo), or combinations thereof. Arg/Lys was injected either i.p. 30 min before and after the radiopharmaceutical, or i.v. 2 min before the radiopharmaceutical. Gelo was administered either i.v. 2 min prior activity, or pre-mixed and injected together with the radiopharmaceutical (n = 5 per group). C57BL/6N mice were furthermore imaged by PET (90 min p.i.) and SPECT (24 h p.i.). RESULTS: Kidney uptake of 68Ga-Trivehexin in C57BL/6N mice was reduced by 15% (Arg/Lys i.p.), 25% (Arg/Lys i.v.), and 70% (gelo i.v.), 90 min p.i., relative to control. 177Lu-D0301 kidney uptake was reduced by 2% (Arg/Lys i.p.), 41% (Arg/Lys i.v.), 61% (gelo i.v.) and 66% (gelo + Arg/Lys i.v.) 24 h p.i., compared to control. Combination of Arg/Lys and gelo provided no substantial benefit. Gelo furthermore reduced kidney uptake of 177Lu-D0301 by 76% (90 min p.i.) and 85% (24 h p.i.) in H2009 bearing SCID mice. Since tumor uptake was not (90 min p.i.) or only slightly reduced (15%, 24 h p.i.), the tumor/kidney ratio was improved by factors of 3.3 (90 min p.i.) and 2.6 (24 h p.i.). Reduction of kidney uptake was demonstrated by SPECT, which also showed that the remaining activity was located in the cortex. CONCLUSIONS: The kidney uptake of both investigated radiopharmaceuticals was more efficiently reduced by gelofusine (61-85%) than Arg/Lys (25-41%). Gelofusine appears particularly suitable for reducing renal uptake of αvß6-integrin targeted 177Lu-labeled peptide multimers because its application led to approximately three times higher tumor-to-kidney ratios. Since the incidence of severe adverse events (anaphylaxis) with succinylated gelatin products (reportedly 0.0062-0.038%) is comparable to that of gadolinium-based MRI or iodinated CT contrast agents (0.008% and 0.04%, respectively), clinical use of gelofusine during radioligand therapy appears feasible if similar risk management strategies as for contrast agents are applied.

Asunto(s)

Gelatina , Integrinas , Riñón , Animales , Ratones , Riñón/metabolismo , Riñón/diagnóstico por imagen , Distribución Tisular , Humanos , Integrinas/metabolismo , Gelatina/química , Radiofármacos/farmacocinética , Marcaje Isotópico , Línea Celular Tumoral , Radioisótopos de Galio , Aminoácidos/química , Aminoácidos/farmacocinética , Transporte Biológico , Péptidos/farmacocinética , Péptidos/química , Seguridad , Ratones Endogámicos C57BL , Lutecio , Tomografía de Emisión de Positrones , Antígenos de Neoplasias , Radioisótopos , Succinatos

RESUMEN

Background: Therapeutic proteins and peptides offer great advantages compared to traditional synthetic molecular drugs. However, stable protein loading and precise control of protein release pose significant challenges due to the extensive range of physicochemical properties inherent to proteins. The development of a comprehensive protein delivery strategy becomes imperative accounting for the diverse nature of therapeutic proteins. Methods: Biodynamers are amphiphilic proteoid dynamic polymers consisting of amino acid derivatives connected through pH-responsive dynamic covalent chemistry. Taking advantage of the amphiphilic nature of the biodynamers, PNCs and DEs were possible to be prepared and investigated to compare the delivery efficiency in drug loading, stability, and cell uptake. Results: As a result, the optimized PNCs showed 3-fold encapsulation (<90%) and 5-fold loading capacity (30%) compared to DE-NPs. PNCs enhanced the delivery efficiency into the cells but aggregated easily on the cell membrane due to the limited stability. Although DE-NPs were limited in loading capacity compared to PNCs, they exhibit superior adaptability in stability and capacity for delivering a wider range of proteins compared to PNCs. Conclusion: Our study highlights the potential of formulating both PNCs and DE-NPs using the same biodynamers, providing a comparative view on protein delivery efficacy using formulation methods.

Asunto(s)

Emulsiones , Péptidos , Péptidos/química , Péptidos/administración & dosificación , Péptidos/farmacocinética , Emulsiones/química , Humanos , Proteínas/química , Proteínas/administración & dosificación , Proteínas/farmacocinética , Sistemas de Liberación de Medicamentos/métodos , Polímeros/química , Nanopartículas/química , Concentración de Iones de Hidrógeno , Aminoácidos/química , Portadores de Fármacos/química , Portadores de Fármacos/farmacocinética , Liberación de Fármacos , Supervivencia Celular/efectos de los fármacos

RESUMEN

Peptides, despite their therapeutic potential, face challenges with undesirable pharmacokinetic (PK) properties and biodistribution, including poor oral absorption and cellular uptake, and short plasma elimination half-lives. Lipidation of peptides is a common strategy to improve their physicochemical and PK properties, making them viable drug candidates. For example, the plasma half-life of peptides has been extended via conjugation to lipids that are proposed to promote binding to serum albumin and thus protect against rapid clearance. Recent work has shown that lipid conjugation to oligodeoxynucleotides, polymers and small molecule drugs results in association not only with albumin, but also with lipoproteins, resulting in half-life prolongation and transport from administration sites via the lymphatics. Enhancing delivery into the lymph increases the efficacy of vaccines and therapeutics with lymphatic targets such as immunotherapies. In this study, the plasma PK, lymphatic uptake, and bioavailability of the glucagon-like peptide-1 (GLP-1) receptor agonist peptides, liraglutide (lipidated) and exenatide (non-lipidated), were investigated following subcutaneous (SC) administration to rats. As expected, liraglutide displayed an apparent prolonged plasma half-life (9.1 versus 1 h), delayed peak plasma concentrations and lower bioavailability (∼10 % versus ∼100 %) compared to exenatide after SC administration. The lymphatic uptake of both peptides was relatively low (<0.5 % of the dose) although lymph to plasma concentration ratios were greater than one for several early timepoints suggesting some direct uptake into lymph. The low lymphatic uptake may be due to the nature of the conjugated lipid (a single-chain C16 palmitic acid in liraglutide) but suggests that other peptides with similar lipid conjugations may also have relatively modest lymphatic uptake. If delivery to the lymph is desired, conjugation to more lipophilic moieties with higher albumin and/or lipoprotein binding efficiencies, such as diacylglycerols, may be appropriate.

Asunto(s)

Exenatida , Liraglutida , Péptidos , Ratas Sprague-Dawley , Animales , Exenatida/farmacocinética , Exenatida/administración & dosificación , Exenatida/farmacología , Liraglutida/farmacología , Liraglutida/farmacocinética , Liraglutida/administración & dosificación , Ratas , Masculino , Péptidos/farmacocinética , Péptidos/administración & dosificación , Lípidos/química , Semivida , Ponzoñas/farmacocinética , Ponzoñas/administración & dosificación , Disponibilidad Biológica , Distribución Tisular , Inyecciones Subcutáneas , Linfa/metabolismo , Linfa/efectos de los fármacos , Receptor del Péptido 1 Similar al Glucagón/agonistas , Receptor del Péptido 1 Similar al Glucagón/metabolismo , Péptido 1 Similar al Glucagón/farmacocinética , Péptido 1 Similar al Glucagón/metabolismo , Sistema Linfático/metabolismo , Sistema Linfático/efectos de los fármacos , Hipoglucemiantes/farmacocinética , Hipoglucemiantes/administración & dosificación , Hipoglucemiantes/farmacología

RESUMEN

A critical parameter during the development of protein therapeutics is to endow them with suitable pharmacokinetic and pharmacodynamic properties. Small protein drugs are quickly eliminated by kidney filtration, and in vivo half-life extension is therefore often desired. Here, different half-life extension technologies were studied where PAS polypeptides (PAS300, PAS600), XTEN polypeptides (XTEN288, XTEN576), and an albumin binding domain (ABD) were compared for half-life extension of an anti-human epidermal growth factor receptor 2 (HER2) affibody-drug conjugate. The results showed that extension with the PAS or XTEN polypeptides or the addition of the ABD lowered the affinity for HER2 to some extent but did not negatively affect the cytotoxic potential. The half-lives in mice ranged from 7.3 h for the construct including PAS300 to 11.6 h for the construct including PAS600. The highest absolute tumor uptake was found for the construct including the ABD, which was 60 to 160% higher than the PASylated or XTENylated constructs, even though it did not have the longest half-life (9.0 h). A comparison of the tumor-to-normal-organ ratios showed the best overall performance of the ABD-fused construct. In conclusion, PASylation, XTENylation, and the addition of an ABD are viable strategies for half-life extension of affibody-drug conjugates, with the best performance observed for the construct including the ABD.

Asunto(s)

Péptidos , Receptor ErbB-2 , Animales , Semivida , Receptor ErbB-2/metabolismo , Humanos , Línea Celular Tumoral , Péptidos/química , Péptidos/farmacocinética , Péptidos/administración & dosificación , Femenino , Ratones Desnudos , Albúminas/química , Proteínas Recombinantes de Fusión/farmacocinética , Proteínas Recombinantes de Fusión/química , Proteínas Recombinantes de Fusión/administración & dosificación , Antineoplásicos/administración & dosificación , Antineoplásicos/farmacocinética , Antineoplásicos/química , Ratones , Neoplasias/tratamiento farmacológico , Neoplasias/metabolismo , Inmunoconjugados/farmacocinética , Inmunoconjugados/química , Inmunoconjugados/administración & dosificación , Ratones Endogámicos BALB C , Distribución Tisular

RESUMEN

Antisense oligonucleotide (ASO) has emerged as a promising therapeutic approach for treating central nervous system (CNS) disorders by modulating gene expression with high selectivity and specificity. However, the poor permeability of ASO across the blood-brain barrier (BBB) diminishes its therapeutic success. Here, we designed and synthesized a series of BBB-penetrating peptides (BPP) derived from either the receptor-binding domain of apolipoprotein E (ApoE) or a transferrin receptor-binding peptide (THR). The BPPs were conjugated to phosphorodiamidate morpholino oligomers (PMO) that are chemically analogous to the 2'-O-(2-methoxyethyl) (MOE)-modified ASO approved by the FDA for treating spinal muscular atrophy (SMA). The BPP-PMO conjugates significantly increased the level of full-length SMN2 in the patient-derived SMA fibroblasts in a concentration-dependent manner with minimal to no toxicity. Furthermore, the systemic administration of the most potent BPP-PMO conjugates significantly increased the expression of full-length SMN2 in the brain and spinal cord of SMN2 transgenic adult mice. Notably, BPP8-PMO conjugate showed a 1.25-fold increase in the expression of full-length functional SMN2 in the brain. Fluorescence imaging studies confirmed that 78% of the fluorescently (Cy7)-labelled BPP8-PMO reached brain parenchyma, with 11% uptake in neuronal cells. Additionally, the BPP-PMO conjugates containing retro-inverso (RI) D-BPPs were found to possess extended half-lives compared to their L-counterparts, indicating increased stability against protease degradation while preserving the bioactivity. This delivery platform based on BPP enhances the CNS bioavailability of PMO targeting the SMN2 gene, paving the way for the development of systemically administered neurotherapeutics for CNS disorders.

Asunto(s)

Apolipoproteínas E , Barrera Hematoencefálica , Ratones Transgénicos , Oligonucleótidos Antisentido , Animales , Barrera Hematoencefálica/metabolismo , Barrera Hematoencefálica/efectos de los fármacos , Oligonucleótidos Antisentido/administración & dosificación , Oligonucleótidos Antisentido/farmacología , Oligonucleótidos Antisentido/farmacocinética , Humanos , Apolipoproteínas E/metabolismo , Ratones , Morfolinos/administración & dosificación , Morfolinos/farmacocinética , Morfolinos/farmacología , Proteína 2 para la Supervivencia de la Neurona Motora/genética , Proteína 2 para la Supervivencia de la Neurona Motora/metabolismo , Atrofia Muscular Espinal/tratamiento farmacológico , Sistemas de Liberación de Medicamentos/métodos , Fibroblastos/metabolismo , Fibroblastos/efectos de los fármacos , Encéfalo/metabolismo , Encéfalo/efectos de los fármacos , Péptidos/administración & dosificación , Péptidos/farmacología , Péptidos/química , Péptidos/farmacocinética , Péptidos de Penetración Celular/química

RESUMEN

Very late antigen-4 (VLA-4) is a transmembrane integrin protein that is highly expressed in aggressive forms of metastatic melanoma. A small-molecule peptidomimetic, LLP2A, was found to have a low pM affinity binding to VLA-4. Because LLP2A itself does not inhibit cancer cell proliferation and survival, it is an ideal candidate for the imaging and delivery of therapeutic payloads. An analog of [177Lu]Lu-labeled-LLP2A was previously investigated as a therapeutic agent in melanoma tumor-bearing mice, resulting in only a modest improvement in tumor growth inhibition, likely due to rapid clearance of the agent from the tumor. To improve the pharmacokinetic profile, DOTAGA-PEG4-LLP2A with a 4-(p-iodophenyl)butyric acid (pIBA) albumin binding moiety was synthesized. We demonstrate the feasibility of this albumin binding strategy by comparing in vitro cell binding assays and in vivo biodistribution performance of [177Lu]Lu-DOTAGA-PEG4-LLP2A ([177Lu]Lu-1) to the albumin binding [177Lu]Lu-DOTAGA-pIBA-PEG4-LLP2A ([177Lu]Lu-2). In vitro cell binding assay results for [177Lu]Lu-1 and [177Lu]Lu-2 showed Kd values of 0.40 ± 0.07 and 1.75 ± 0.40 nM, with similar Bmax values of 200 ± 6 and 315 ± 15 fmol/mg, respectively. In vivo biodistribution data for both tracers exhibited specific uptake in the tumor, spleen, thymus, and bone due to endogenous expression of VLA-4. Compound [177Lu]Lu-2 exhibited a much longer blood circulation time compared to [177Lu]Lu-1. The tumor uptake for [177Lu]Lu-1 was highest at 1 h (∼15%ID/g) and that for [177Lu]Lu-2 was highest at 4 h (∼23%ID/g). Significant clearance of [177Lu]Lu-1 from the tumor occurs at 24 h (<5%ID/g) while[177Lu]Lu-2 is retained for greater than 96 h (∼10%ID/g). An efficacy study showed that melanoma tumor-bearing mice receiving compound [177Lu]Lu-2 given in two fractions (2 × 14.8 MBq, 14 days apart) had a greater median survival time than mice administered a single 29.6 MBq dose of compound [177Lu]Lu-1, while a single 29.6 MBq dose of [177Lu]Lu-2 imparted hematopoietic toxicity. The in vitro and in vivo data show addition of pIBA to [177Lu]Lu-DOTAGA-PEG4-LLP2A slows blood clearance for a higher tumor uptake, and there is potential of [177Lu]Lu-2 as a theranostic in fractionated administered doses.

Asunto(s)

Lutecio , Radioisótopos , Animales , Ratones , Distribución Tisular , Línea Celular Tumoral , Melanoma/tratamiento farmacológico , Melanoma/metabolismo , Humanos , Radiofármacos/farmacocinética , Melanoma Experimental/tratamiento farmacológico , Melanoma Experimental/metabolismo , Femenino , Integrina alfa4beta1/metabolismo , Integrina alfa4beta1/antagonistas & inhibidores , Albúminas , Péptidos/química , Péptidos/farmacocinética , Nanomedicina Teranóstica/métodos , Ratones Endogámicos C57BL , Dipéptidos , Compuestos de Fenilurea

RESUMEN

Phage display is a versatile method often used in the discovery of peptides that targets disease-related biomarkers. A major advantage of this technology is the ease and cost efficiency of affinity selection, also known as biopanning, to identify novel peptides. While it is relatively straightforward to identify peptides with optimal binding affinity, the pharmacokinetics of the selected peptides often prove to be suboptimal. Therefore, careful consideration of the experimental conditions, including the choice of using in vitro, in situ, or in vivo affinity selections, is essential in generating peptides with high affinity and specificity that also demonstrate desirable pharmacokinetics. Specifically, in vivo biopanning, or the combination of in vitro, in situ, and in vivo affinity selections, has been proven to influence the biodistribution and clearance of peptides and peptide-conjugated nanoparticles. Additionally, the marked difference in properties between peptides and nanoparticles must be considered. While peptide biodistribution depends primarily on physiochemical properties and can be modified by amino acid modifications, the size and shape of nanoparticles also affect both absorption and distribution. Thus, optimization of the desired pharmacokinetic properties should be an important consideration in biopanning strategies to enable the selection of peptides and peptide-conjugated nanoparticles that effectively target biomarkers in vivo.

Asunto(s)

Técnicas de Visualización de Superficie Celular , Péptidos , Péptidos/farmacocinética , Péptidos/química , Animales , Técnicas de Visualización de Superficie Celular/métodos , Humanos , Distribución Tisular , Nanopartículas/química , Biblioteca de Péptidos