Cell-penetrating compounds are substances that enhance the cellular uptake of various molecular cargoes that do not easily cross the cellular membrane. The majority of cell-penetrating compounds described in the literature are cell-penetrating peptides (CPPs). This review summarizes the various structural types of cell-penetrating compounds, with the main focus on CPPs. The authors present a brief overview of the history of CPPs, discuss the various types of conjugation of CPPs to biologically active cargoes intended for cell internalization, examine the cell-entry mechanisms of CPPs, and report on the applications of CPPs in research and in preclinical and clinical studies.

This report describes the design, synthesis and evaluation of tumor-targeted polymer probes to visualize epidermal growth factor receptor (EGFR)-positive malignant tumors for successful resection via fluorescence guided endoscopic surgery. Fluorescent polymer probes of various molecular weights enabling passive accumulation in tumors via enhanced permeability and retention were prepared and evaluated, showing an optimal molecular weight of 200,000 g/mol for passive tumor targeting. Moreover, poly(N-(2-hydroxypropyl)methacrylamide)-based copolymers labeled with fluorescent dyes were targeted with the EGFR-binding oligopeptide GE-11 (YHWYGYTPQNVI), human EGF or anti-EGFR monoclonal antibody cetuximab were all able to actively target the surface of EGFR-positive tumor cells. Nanoprobes targeted with GE-11 and cetuximab showed the best targeting profile but differed in their tumor accumulation kinetics. Cetuximab increased tumor accumulation after 15 min, whereas GE 11 needed at least 4 h. Interestingly, after 4 h, there were no significant differences in tumor targeting, indicating the potential of oligopeptide targeting for fluorescence-navigated surgery. In conclusion, fluorescent polymer probes targeted by oligopeptide GE-11 or whole antibody are excellent tools for surgical navigation during oncological surgery of head and neck squamous cell carcinoma, due to their relatively simple design, synthesis and cost, as well as optimal pharmacokinetics and accumulation in tumors.

Cell-penetrating peptides (CPPs) are commonly used substances enhancing the cellular uptake of various cargoes that do not easily cross the cellular membrane. CPPs can be either covalently bound directly to the cargo or they can be attached to a transporting system such as a polymer carrier together with the cargo. In this work, several CPP–polymer conjugates based on copolymers of N-(2-hydroxypropyl)methacrylamide (pHPMA) with HIV-1 Tat peptide (TAT), a minimal sequence of penetratin (PEN), IRS-tag (RYIRS), and PTD4 peptide, and the two short hydrophobic peptides VPMLK and PFVYLI were prepared and characterized. Moreover, the biological efficacy of fluorescently labeled polymer carriers decorated with various CPPs was compared. The experiments revealed that the TAT–polymer conjugate and the PEN–polymer conjugate were internalized about 40 times and 15 times more efficiently than the control polymer, respectively. Incorporation of dodeca(ethylene glycol) spacer improved the cell penetration of both studied polymer–peptide conjugates compared to the corresponding spacer-free polymer conjugates, while the shorter tetra(ethylene glycol) spacer improved only the penetration of the TAT conjugate but it did not improve the penetration of the PEN conjugate. Finally, a significantly improved cytotoxic effect of the polymer conjugate containing anticancer drug pirarubicin and TAT attached via a dodeca(ethylene glycol) was observed when compared with the analogous polymer–pirarubicin conjugate without TAT.

Microbial resistance is one of the main problems of modern medicine. Recently, antimicrobial peptides have been recognized as a novel approach to overcome the microbial resistance issue, nevertheless, their low stability, toxicity, and potential immunogenic response in biological systems have limited their clinical application. Herein, we present the design, synthesis, and preliminary biological evaluation of polymer-antibacterial peptide constructs. The antimicrobial GKWMKLLKKILK-NH₂ oligopeptide (PEP) derived from halictine, honey bee venom, was bound to a polymer carrier via various biodegradable spacers employing the pH-sensitive or enzymatically-driven release and reactivation of the PEP’s antimicrobial activity. The antibacterial properties of the polymer-PEP constructs were assessed by a determination of the minimum inhibitory concentrations, followed by fluorescence and transmission electron microscopy. The PEP exerted antibacterial activity against both, gram-positive and negative bacteria, via disruption of the bacterial cell wall mechanism. Importantly, PEP partly retained its antibacterial efficacy against Staphylococcus epidermidis, Escherichia coli, and Acinetobacter baumanii even though it was bound to the polymer carrier. Indeed, to observe antibacterial activity similar to the free PEP, the peptide has to be released from the polymer carrier in response to a pH decrease. Enzymatically-driven release and reactivation of the PEP antimicrobial activity were recognized as less effective when compared to the pH-sensitive release of PEP.

Nano‐sized polymer systems are often used as carriers for drugs and contrast agents to increase circulation time and solubility and to reduce possible side effects. These nanomedicines usually accumulate in tumor tissue due to the enhanced permeability and retention (EPR) effect. However, a targeting group may be attached to the polymer carrier in addition to the active substance to further increase tumor accumulation and specificity. In this study, the oligopeptide sequence RGD was chosen to target α_vβ₃ integrins overexpressed in the tumor vasculature and on some tumor cells. A set of polymer conjugates bearing a fluorescent dye and RGD peptide of different structures (linear, cyclic, branched) was prepared for use in tumor diagnosis, with a potential future application in navigated surgery. The accumulation of the most promising candidate, a targeted fluorescent nanoprobe, increased by 35% in glioblastoma tumors compared to the non‐targeted control, which accumulated only due to the EPR effect. However, the administration of a polymer‐bound modified cilengitide as an antiangiogenic treatment did not show a beneficial effect in the suppression of angiogenesis.

Nano‐sized carriers are widely studied as suitable candidates for the advanced delivery of various bioactive molecules such as drugs and diagnostics. Herein, the development of long‐circulating stimuli‐responsive polymer nanoprobes tailored for the fluorescently‐guided surgery of solid tumors is reported. Nanoprobes are designed as long‐circulating nanosystems preferably accumulated in solid tumors due to the Enhanced permeability and retention effect, so they act as a tumor microenvironment‐sensitive activatable diagnostic. This study designs polymer probes differing in the structure of the spacer between the polymer carrier and Cy7 by employing pH‐sensitive spacers, oligopeptide spacers susceptible to cathepsin B‐catalyzed enzymatic hydrolysis, and non‐degradable control spacer. Increased accumulation of the nanoprobes in the tumor tissue coupled with stimuli‐sensitive release behavior and subsequent activation of the fluorescent signal upon dye release facilitated favorable tumor‐to‐background ratio, a key feature for fluorescence‐guided surgery. The probes show excellent diagnostic potential for the surgical removal of intraperitoneal metastasis and orthotopic head and neck tumors with very high efficacy and accuracy. In addition, the combination of macroscopic resection followed by fluorescence‐guided surgery using developed probes enable the identification and resection of most of the CAL33 intraperitoneal metastases with total tumor burden reduced to 97.2%.