Synthesis of Polystyrene-Based Cationic Nanomaterials with Pro-Oxidant Cytotoxic Activity on Etoposide-Resistant Neuroblastoma Cells

Abstract

Drug resistance is a multifactorial phenomenon that limits the action of antibiotics and chemotherapeutics. Therefore, it is essential to develop new therapeutic strategies capable of inducing cytotoxic effects circumventing chemoresistance. In this regard, the employment of natural and synthetic cationic peptides and polymers has given satisfactory results both in microbiology, as antibacterial agents, but also in the oncological field, resulting in effective treatment against several tumors, including neuroblastoma (NB). To this end, two polystyrene-based copolymers (P5, P7), containing primary ammonium groups, were herein synthetized and tested on etoposide-sensitive (HTLA-230) and etoposide-resistant (HTLA-ER) NB cells. Both copolymers were water-soluble and showed a positive surface charge due to nitrogen atoms, which resulted in protonation in the whole physiological pH range. Furthermore, P5 and P7 exhibited stability in solution, excellent buffer capacity, and nanosized particles, and they were able to reduce NB cell viability in a concentration-dependent way. Interestingly, a significant increase in reactive oxygen species (ROS) production was observed in both NB cell populations treated with P5 or P7, establishing for both copolymers an unequivocal correlation between cytotoxicity and ROS generation. Therefore, P5 and P7 could be promising template macromolecules for the development of new chemotherapeutic agents able to fight NB chemoresistance.

Broad-Spectrum Bactericidal Activity of a Synthetic Random Copolymer Based on 2-Methoxy-6-(4-Vinylbenzyloxy)-Benzylammonium Hydrochloride

“…Interestingly, the MIC values of P7 against Gram-positive isolates were very similar to those obtained by the recently published P5 copolymer [13]. However, differently from what was observed when testing its anti-tumor activity on NB cells, which was lower than that of P5 [20], when tested on bacteria P7 was found to be more cytotoxic than P5 [13], showing both MIC and MBC values very much lower than those of P5. The lowest MIC values were observed against the genus Enterococcus, including some vancomycinresistant strains (VRE) (MICs = 0.6-1.15 µM on E. faecium and 2.3 µM on E. faecalis) and against the sporogenic Bacillus subtilis (MIC = 1.15 µM), which reported double or overlapping MBC values compared to the MICs.…”

“…The experimental details are also included in Section S1 of the Supplementary Information (SI) (Scheme S1 and Sections S1.1-S1.7) associated with this article. The synthetic intermediates and the final compound M7 were spectrophotometrically characterized by means of FTIR and NMR analyses, as shown in the SI, which confirmed their structures, according to what has been reported in the literature [20]. In addition, other analysis, including HPLC, GC-MS, elemental analysis, as well as melting and boiling point determinations, were carried out when useful or necessary and the results, which are also available in Section S1 (SI), were successfully compared with those that have previously been reported [20].…”

“…However, for the sake of clarity, the main physicochemical properties of P7 have been included in Table 1 below. M7 was prepared by performing the synthetic procedure reported and extensively discussed in a recent study [20]. The experimental details are also included in Section S1 of the Supplementary Information (SI) (Scheme S1 and Sections S1.1-S1.7) associated with this article.…”

“…However, for the sake of clarity, the main physicochemical properties of P7 have been included in Table 1 below. Having shown remarkable cytotoxicity on tumor cells with membranes resembling those of bacteria in terms of charge [20], and differing from those of eukaryotic cells, towards which cationic materials such as P7 have been reported to be non-cytotoxic [21,25], the P7 copolymer appeared to us as an excellent candidate for developing a new antibacterial tool for several reasons. Due to the structure of M7, used as a cationic monomer, P7 possessed primary benzylammonium hydrochloride residues as cationic moieties, which have been reported to confer interesting antimicrobial properties on several homopolymers and copolymers [11,[26][27][28][29].…”

“…Since it is known that the cytoplasmic membrane of cancer cells, including neuroblastoma (NB) cell lines, is more like that of prokaryotes than that of eukaryotes, and cationic material active on bacteria are selectively cytotoxic also on tumor cells, including multidrug-resistant ones [20,21,25], we recently observed the cytotoxic effects of P5 on two cell lines that were differently sensitive to ETO [20]. In that study, we also reported the preparation, physicochemical characterization, and the significant cytotoxic effects of a new watersoluble cationic copolymer (P7) (Figure 2), obtained by polymerizing 2-methoxy-6-(4vinylbenzyloxy)-benzylammonium hydrochloride monomer (M7) with di-methyl acrylamide (DMAA) comonomer [20]. P5 proved to be much more potent than the low-molecular-weight monomer M5 and displayed remarkable antibacterial activity against most of the 61 isolates tested, belonging both to Gram-positive and Gram-negative species [13].…”