Gold nanoparticles stabilize peptide-drug-conjugates for sustained targeted drug delivery to cancer cells

Abstract Background Peptide-drug-conjugates (PDCs) are being developed as an effective strategy to specifically deliver cytotoxic drugs to cancer cells. However one of the challenges to their successful application is the relatively low stability of peptides in the blood, liver and kidneys. Since AuNPs seem to have a longer plasma half-life than PDCs, one approach to overcoming this problem would be to conjugate the PDCs to gold nanoparticles (AuNPs), as these have demonstrated favorable physico-chemical and safety properties for drug delivery systems. We set out to test whether PEG coated-AuNPs could provide a suitable platform for the non-covalent loading of pre-formed PDCs and whether this modification would affect the bioavailability of the PDCs and their cytotoxicity toward target cancer cells. Methods Peptides specifically internalized by A20 murine lymphoma cells were isolated from a phage library displaying 7mer linear peptides. Peptide specificity was validated by flow cytometry and confocal microscopy. PDCs were synthesized containing a selected peptide (P4) and either chlorambucil (Chlor), melphalan (Melph) or bendamustine (Bend). Gold nanoparticles were sequentially coated with citrate, PEG-6000 and then PDC (PDC-PEG-AuNP). The physico-chemical properties of the coated particles were analyzed by electrophoresis, TEM, UV–VIS and FTIR. Stability of free and PDC-coated AuNP was determined. Results Biopanning of the phage library resulted in discovery of several novel peptides that internalized into A20 cells. One of these (P4) was used to synthesize PDCs containing either Chlor, Melph or Bend. All three PDCs specifically killed A20 target cells, however they had short half-lives ranging from 10.6 to 15.4 min. When coated to PEG-AuNPs, the half-lives were extended to 21.0–22.3 h. The PDC-PEG-AuNPs retained cytotoxicity towards the target cells. Moreover, whereas pre-incubation for 24 h of free PDCs almost completely abolished their cytotoxic activity, the PDC-PEG-AuNPs were still active even after 72 h pre-incubation. Conclusions Peptide-drug-conjugates hold potential for improving the target efficacy of chemotherapeutic drugs, however their short half-lives may limit their application. This hurdle can be overcome by easily conjugating them to gold nanoparticles. This conjugation also opens up the possibility of developing slow release formulations of targeted drug delivery systems containing PDCs

Effect of Solubilizing Group on the Antibacterial Activity of Heptamethine Cyanine Photosensitizers

Antibiotic resistance of pathogenic bacteria dictates the development of novel treatment modalities such as antimicrobial photodynamic therapy (APDT) utilizing organic dyes termed photosensitizers that exhibit a high cytotoxicity upon light irradiation. Most of the clinically approved photosensitizers are porphyrins that are poorly excitable in the therapeutic near-IR spectral range. In contrast, cyanine dyes function well in the near-IR region, but their phototoxicity, in general, is very low. The introduction of iodine atoms in the cyanine molecules was recently demonstrated to greatly increase their phototoxicity. Herein, we synthesized a series of the new iodinated heptamethine cyanine dyes (ICy7) containing various solubilizing moieties, i.e., negatively charged carboxylic (ICy7COOH) and sulfonic (ICy7SO3H) groups, positively charged triphenylphosphonium (ICy7PPh3), triethylammonium (ICy7NEt3) and amino (ICy7NH2) groups, and neutral amide (ICy7CONHPr) group. The effect of these substituents on the photodynamic eradication of Gram-positive (S. aureus) and Gram-negative (E. coli and P. aeruginosa) pathogens was studied. Cyanine dyes containing the amide and triphenylphosphonium groups were found to be the most efficient for eradication of the investigated bacteria. These dyes are effective at low concentrations of 0.05 µM (33 J/cm2) for S. aureus, 50 µM (200 J/cm2) for E. coli, and 5 µM (100 J/cm2) for P. aeruginosa and considered, therefore, promising photosensitizers for APDT applications. The innovation of the new photosensitizers consisted of a combination of the heavy-atom effect that increases singlet oxygen generation with the solubilizing group’s effect improving cell uptake, and with effective near-IR excitation. Such a combination helped to noticeably increase the APDT efficacy and should pave the way for the development of more advanced photosensitizers for clinical use

Novel Cyclic Peptides for Targeting EGFR and EGRvIII Mutation for Drug Delivery

The epidermal growth factor–epidermal growth factor receptor (EGF-EGFR) pathway has become the main focus of selective chemotherapeutic intervention. As a result, two classes of EGFR inhibitors have been clinically approved, namely monoclonal antibodies and small molecule kinase inhibitors. Despite an initial good response rate to these drugs, most patients develop drug resistance. Therefore, new treatment approaches are needed. In this work, we aimed to find a new EGFR-specific, short cyclic peptide, which could be used for targeted drug delivery. Phage display peptide technology and biopanning were applied to three EGFR expressing cells, including cells expressing the EGFRvIII mutation. DNA from the internalized phage was extracted and the peptide inserts were sequenced using next-generation sequencing (NGS). Eleven peptides were selected for further investigation using binding, internalization, and competition assays, and the results were confirmed by confocal microscopy and peptide docking. Among these eleven peptides, seven showed specific and selective binding and internalization into EGFR positive (EGFR+ve) cells, with two of them—P6 and P9—also demonstrating high specificity for non-small cell lung cancer (NSCLC) and glioblastoma cells, respectively. These peptides were chemically conjugated to camptothecin (CPT). The conjugates were more cytotoxic to EGFR+ve cells than free CPT. Our results describe a novel cyclic peptide, which can be used for targeted drug delivery to cells overexpressing the EGFR and EGFRvIII mutation

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Additional file 1. Additional figures S1–S9 and table S1