Applications and Challenges for Use of Cell-Penetrating Peptides as Delivery Vectors for Peptide and Protein Cargos.

The hydrophilic nature of peptides and proteins renders them impermeable to cell membranes. Thus, in order to successfully deliver peptide and protein-based therapeutics across the plasma membrane or epithelial and endothelial barriers, a permeation enhancing strategy must be employed. Cell-penetrating peptides (CPPs) constitute a promising tool and have shown applications for peptide and protein delivery into cells as well as across various epithelia and the blood-brain barrier (BBB). CPP-mediated delivery of peptides and proteins may be pursued via covalent conjugation of the CPP to the cargo peptide or protein or via physical complexation obtained by simple bulk-mixing of the CPP with its cargo. Both approaches have their pros and cons, and which is the better choice likely relates to the physicochemical properties of the CPP and its cargo as well as the route of administration, the specific barrier and the target cell. Besides the physical barrier, a metabolic barrier must be taken into consideration when applying peptide-based delivery vectors, such as the CPPs, and stability-enhancing strategies are commonly employed to prolong the CPP half-life. The mechanisms by which CPPs translocate cell membranes are believed to involve both endocytosis and direct translocation, but are still widely investigated and discussed. The fact that multiple factors influence the mechanisms responsible for cellular CPP internalization and the lack of sensitive methods for detection of the CPP, and in some cases the cargo, further complicates the design and conduction of conclusive mechanistic studies.

Quality by design thinking in the development of long-acting injectable PLGA/PLA-based microspheres for peptide and protein drug delivery.

Adopting the Quality by Design (QbD) approach in the drug development process has transformed from "nice-to-do" into a crucial and required part of the development, ensuring the quality of pharmaceutical products throughout their whole life cycles. This review is discussing the implementation of the QbD thinking into the production of long-acting injectable (LAI) PLGA/PLA-based microspheres for the therapeutic peptide and protein drug delivery. Various key elements of the QbD approaches are initially elaborated using Bydureon®, a commercial product of LAI PLGA/PLA-based microspheres, as a classical example. Subsequently, the factors influencing the release patterns and the stability of the peptide and protein drugs are discussed. This is followed by a summary of the state-of-the-art of manufacturing LAI PLGA/PLA-based microspheres and the related critical process parameters (CPPs). Finally, a landscape of generic product development of LAI PLGA/PLA-based microspheres is reviewed including some major challenges in the field.

Cell-penetrating peptides as tools to enhance non-injectable delivery of biopharmaceuticals.

Non-injectable delivery of peptide and protein drugs is hampered by their labile nature, hydrophilicity, and large molecular size; thus limiting their permeation across mucosae, which represent major biochemical and physical barriers to drugs administered via e.g. the oral, nasal, and pulmonary routes. However, in recent years cell-penetrating peptides (CPP) have emerged as promising tools to enhance mucosal delivery of co-administered or conjugated peptide and protein cargo and more advanced CPP-cargo formulations are emerging. CPPs act as transepithelial delivery vectors, but the mechanism(s) by which CPPs mediate cargo translocation across an epithelium is so far poorly understood; both due to the fact that multiple factors influence the resulting uptake and trafficking mechanisms as well as to the complicated nature of sensitive studies of this. In addition to a proper mechanistic understanding, documentation of CPP-mediated delivery in higher animal species than rodent as well as extensive toxicological studies are necessary for CPP-containing non-injectable DDSs to reach the clinic.

Enhanced oral bioavailability of insulin using PLGA nanoparticles co-modified with cell-penetrating peptides and Engrailed secretion peptide (Sec).

Biodegradable polymer nanoparticle drug carriers are an attractive strategy for oral delivery of peptide and protein drugs. However, their ability to cross the intestinal epithelium membrane is largely limited. Therefore, in the present study, cell-penetrating peptides (R8, Tat, penetratin) and a secretion peptide (Sec) with N-terminal stearylation were introduced to modify nanoparticles (NPs) on the surface to improve oral bioavailability of peptide and protein drugs. In vitro studies conducted in Caco-2 cells showed the value of the apparent permeability coefficient (Papp) of the nanoparticles co-modified with Sec and penetratin (Sec-Pen-NPs) was about two-times greater than that of the nanoparticles modified with only penetratin (Pen-NPs), while the increase of transcellular transport of nanoparticles modified together with Sec and R8 (Sec-R8-NPs), or Sec and Tat (Sec-Tat-NPs), was not significant compared with nanoparticles modified with only R8 (R8-NPs) or Tat (Tat-NPs). Using insulin as the model drug, in vivo studies performed on rats indicated that compared to Pen-NPs, the relative bioavailability of insulin for Sec-Pen-NPs was 1.71-times increased after ileal segments administration, and stronger hypoglycemic effects was also observed. Therefore, the nanoparticles co-modified with penetratin and Sec could act as attractive carriers for oral delivery of insulin.

Pharmacokinetic, pharmacodynamic and biodistribution following oral administration of nanocarriers containing peptide and protein drugs.

The influence of nanoparticle (NP) formulations on the pharmacokinetic, pharmacodynamic and biodistribution profiles of peptide- and protein-like drugs following oral administration is critically reviewed. The possible mechanisms of absorption enhancement and the effects of the physicochemical properties of the NP are examined. The potential advantages and challenges of physiologically-based pharmacokinetic (PBPK) modelling to help predict efficacy in man are discussed. The importance of developing and expanding the regulatory framework to help translate the technology into the clinic and accelerate the availability of oral nanoparticulate formulations is emphasized. In conclusion, opportunities for future work to improve the state of the art of oral nanomedicines are identified.