Anionic linear-globular dendrimer-cis-platinum (II) conjugates promote cytotoxicity in vitro against different cancer cell lines

Abstract

Due to their unique properties, Anticancer dendrimer-based drugs have been displaying promising results in both in vitro and in vivo in the treatment of cancerous cells, as compared to the traditional polymers. In this report, two conjugates (G1+Pt and G2+Pt) of cisplatin [cis-diaminedichloroplatinum; (CDDP)] with two generations (G1, G2) of a biocompatible anionic dendrimer were prepared in an aqueous media. Their potential cytotoxic effects, in two sensitive cancer cell lines HT1080 and CT26 together with one resistant cancer cell line SKOV3, using MTT (methyl thiazolyl tetrazolium) assay were examined. Hemolytic impacts and cell death mechanisms of the conjugates on human blood and HT1080 cell line were also investigated. The conjugate G2+Pt showed greater toxicity up to 9x and 2x in the sensitive and resistant cell lines (IC(50) comparison, inhibitory concentration) respectively when compared to the parent drug. The G1+Pt conjugate showed greater toxicity only in the sensitive HT1080 (2x) and CT26 (3.7x) cell lines. Moreover, the G1+Pt conjugate was less toxic approximately one third of the cisplatin in SKOV3 after 48 hrs of incubation. In summary, the G2+Pt conjugate had greater toxicity than the G1+Pt conjugate and cisplatin, based on the in vitro results. Approximately the same hemolysis behavior was observed for both conjugates and cisplatin. Both apoptosis and necrosis mechanisms (about 2x more than cisplatin) were attributed to conjugates and cisplatin in a direct correlation between the concentration and the degree of cell death. In conclusion, these conjugates with such high potency and minimum hemolysis would be suitable candidates for use against these cancerous cell lines as efficient and novel antitumor agents.

Keywords: apoptosis-necrosis; cis-platinum (II); dendrimer; hemolysis; in vitro cytotoxicity.

Figure 1

Schematic diagrams of cisplatin (CDDP), the dendrimers G1, G2 and their possible reactions (generation of different conjugates) in aqueous media.

Figure 2

The comparative release of chloride ions from cisplatin (CDDP) alone and in reaction with the dendrimers G1 and G2.

Figure 3

In vitro release of platinum from the G1+Pt and G2+Pt conjugates at two different pHs. Abbreviation: h, hours.

Figure 4

FT-IR spectra of the dendrimers before and after the reaction with cisplatin (CDDP). IR: (A) G1, (B) G1+Pt, (C) G2, (D) G2+Pt. Abbreviations: FTIR, Fourier transform infrared; IR, infra-red.

Figure 5

Viability of the different cancer cell lines according to the mitochondrial activity of the cells incubating with different kinds of Pt derivatives and free cisplatin (CDDP) after 24 and 48 hrs. (A) HT1080 cell line, (B) CT26 cell line, (c) SKOV3.

Figure 5

Viability of the different cancer cell lines according to the mitochondrial activity of the cells incubating with different kinds of Pt derivatives and free cisplatin (CDDP) after 24 and 48 hrs. (A) HT1080 cell line, (B) CT26 cell line, (c) SKOV3.

Figure 6

The IC₅₀ of some of the conjugates and free cisplatin (CDDP) in different cancer cell lines at different times. Abbreviation: h, hours.

Figure 7

In vitro release of hemoglobin from human erythrocytes in contact with various concentrations of free cisplatin (CDDP) and Pt derivatives after 2 hrs of incubation.

Figure 8

Dot plot diagrams related to the amount of apoptosis-necrosis assays. Impact of the conjugates and free cisplatin (CDDP) at two concentrations on the percentage of apoptosis and necrosis. The data were analyzed with a version 2.4 of Flomax software. (A) negative control, (B) G1(0.75 μg/ml), (C) G1(1.5 μg/ml), (D) G2 (0.75 μg/ml), (E) G2 (1.5 μg/ml). (F) Platinum (0.75 μg/ml), (G) Platinum (1.5 μg/ml). Lower right: apoptotic cells, Upper right: necrotic and secondary apoptotic cells, Lower left: viable cells, Upper left: damaged cells.