Design, Synthesis, and Characterization of Novel Pyrazole Cross-Linked Chitosan Derivatives Modified with Zinc Oxide Nanoparticles for Boosting Their Anticancer Activity

Abstract

A new series of chitosan-based pyrazole derivatives was successfully prepared via crosslinking chitosan using either malonopyrazole (MPy-Cs) or thiopyrazole (TPy-Cs). Three derivatives of TPy-Cs were produced based on their content of TPy, namely TPy-Cs1, TPy-Cs2, and TPy-Cs3 of crosslinking degrees of 71, 48, and 29%, respectively. Further, various weight ratios of ZnO nanoparticles were loaded into some of these derivatives to obtain the corresponding ZnONP bio-composites. FTIR, XRD, SEM, and TEM techniques were employed to emphasize the chemical, internal, and morphological structure of these derivatives. Although MPy-Cs derivatives did not show any activity against all the examined cancer cell lines, TPy-Cs derivatives exhibited an appreciable anticancer activity which greatly improved with increasing their TPy content, i.e., from TPy-Cs3 to TPy-Cs1. The TPy-Cs1 displayed IC₅₀ (14.4 μg/mL) against the HN9 cell line that was comparable to the Doxorubicin (DOX) standard drug (12.6 μg/mL). Among all the prepared composites, TPy-Cs3/ZnONPs-5% was the most potent anticancer candidate against all the tested cancer cell lines, although it does not exceed the anticancer activity of DOX. Tpy-Cs2 and its ZnONP composites were safe on normal human skin fibroblast (HSF) cell lines. Thus, the inclusion of both TPy and ZnONPs into the chitosan matrix fostered its anticancer efficiency.

Keywords: ZnO nanoparticles; anticancer activity; chitosan; cytotoxicity; pyrazole; synthesis.

Figure 1

FTIR spectrum of MPy.

Figure 2

¹H-NMR spectrum of MPy.

Figure 3

¹³C-NMR spectrum of MPy.

Scheme 1

Synthesis of novel MPy-Cs derivative: (A) synthesis of MPy; (B) synthesis of MPy-Cs.

Scheme 2

Synthesis of novel TPy-Cs derivatives.

Scheme 3

Presumptive presentation of the interaction between TPy-Cs and ZnONPs in their composites.

Figure 4

FTIR spectrum of MPy-Cs derivative.

Figure 5

FTIR spectrum of TPy-Cs1 derivative.

Figure 6

FTIR spectra of TPy-Cs2/ZnONP composites.

Figure 7

XRD patterns of MPy-Cs and TPy-Cs derivatives.

Figure 8

XRD patterns of MPy-Cs and TPy-Cs3-based ZnONP composites.

Figure 9

SEM images of MPy-Cs, and TPy-Cs derivatives.

Figure 10

TEM images of TPy-Cs2/ZnONPs-5% composite at different magnifications.

Figure 11

IC₅₀ for the investigated MPy-Cs, TPy-Cs derivatives, and their ZnONP composites, beside DOX (as reference drug) against HCT₁₁₆ cancer cell line at 48 h incubation. The results are expressed as the mean ± SD of three separate experiments performed in three replicates.

Figure 12

IC₅₀ for the investigated MPy-Cs, TPy-Cs derivatives, and their ZnONP composites, beside DOX (as reference drug) against A375 cancer cell line at 48 h incubation. The results are expressed as the mean ± SD of three separate experiments performed in three replicates.

Figure 13

IC₅₀ for the investigated MPy-Cs, TPy-Cs derivatives, and their ZnONP composites, beside DOX (as reference drug) against HN9 cancer cell line at 48 h incubation. The results are expressed as the mean ± SD of three separate experiments performed in three replicates.

Figure 14

IC₅₀ for TPy-Cs3/ZnONPs-5% against the three cancer cell lines at 48 h incubation. The results are expressed as the mean ± SD of three separate experiments performed in three replicates.

Figure 15

Cytotoxicity of TPy-Cs2 derivative and its ZnONP composites on the normal human skin fibroblast (HSF) cell line. The results are expressed as the mean ± SD of three separate experiments performed in three replicates.

Figure 16

Structure–activity relationship for the TPy-Cs derivatives.