Telodendrimer nanocarrier for co-delivery of paclitaxel and cisplatin: A synergistic combination nanotherapy for ovarian cancer treatment

Abstract

Cisplatin (CDDP) and paclitaxel (PTX) are two established chemotherapeutic drugs used in combination for the treatment of many cancers, including ovarian cancer. We have recently developed a three-layered linear-dendritic telodendrimer micelles (TM) by introducing carboxylic acid groups in the adjacent layer via "thio-ene" click chemistry for CDDP complexation and conjugating cholic acids via peptide chemistry in the interior layer of telodendrimer for PTX encapsulation. We hypothesize that the co-delivery of low dosage PTX with CDDP could act synergistically to increase the treatment efficacy and reduce their toxic side effects. This design allowed us to co-deliver PTX and CDDP at various drug ratios to ovarian cancer cells. The in vitro cellular assays revealed strongest synergism in anti-tumor effects when delivered at a 1:2 PTX/CDDP loading ratio. Using the SKOV-3 ovarian cancer xenograft mouse model, we demonstrate that our co-encapsulation approach resulted in an efficient tumor-targeted drug delivery, decreased cytotoxic effects and stronger anti-tumor effect, when compared with free drug combination or the single loading TM formulations.

Keywords: Cisplatin; Combination chemotherapy; Drug delivery; Ovarian cancer; Paclitaxel; Synergism.

Fig. 1

¹H NMR spectra of telodendrimer I, II and III in DMSO-d6 at a concentration of 5 mg/mL, detected by 600M Bruker NMR. The protons on Fmoc were marked in telodendrimer I; OH and OCH of CA and vinyl protons appeared in telodendrimer II; the Me of CA and emerging COOH and disappearing vinyl groups were shown in telodendrimer III.

Fig. 2

(A) MALDI-TOF MS spectrum of telodendrimer III (in black) with the baseline corrected (in red). (B) Zeta potential of telodendrimer III in red solid line and CDDP-loaded telodendrimer III in green solid line. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

Fig. 3

DLS particle sizes and TEM images of the empty TM (A, D) and single CDDP loaded TM (B, E) and PTX-CDDP co-loaded TMs (C, F). Scale bar: 100 nm.

Fig. 4

(A) DLS monitoring of particle size of TM_{(CDDP/PTX 2:1)} micelle solution upon storage at 4 °C; The insert background is cryo-TEM image of the stored sample; (B) Drug release profiles of PTX and CDDP from the co-loaded nanoformulation in comparison with free CDDP.

Fig. 5

(A) Confocal fluorescence microscopy images of SKOV-3 cells incubated with free DiI and DiI-loaded micelles at 37 and 4 °C for 30 min and 2 h; (B) The overlay pictures of the cells incubated at 37 °C for 2 h cell nuclear and lysosome were stained with DAPI (Blue) and Lysotracker (green) respectively. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

Fig. 6

Cell viability of SKOV-3 ovarian cancer cells after incubated for 72 h with free CDDP, free PTX, single loading of TM_(PTX) and TM_(CDDP) and the coloading TM (CDDP:PTX0 at different ratios. The cell viabilities were displayed against PTX concentration (A) and CDDP concentration (B), respectively. The combination index of the co-loading TM formulations with different ratio of CDDP/PTX in SKOV-3(C), ES-2 cells (D) and in Hela cells (E).

Fig. 7

In vivo (A) and ex vivo (C) NIRF optical images of Raji lymphoma bearing mice injected intravenously with free DiD and DiD-PTX-CDDP coloaded TM formulations, respectively. The in vivo tumor targeting (B) and ex vivo tumor and organ uptake (D) were quantitatively analyzed.

Fig. 8

(A) In vivo pharmacokinetics profiles of platinum concentration in the plasma after i.v. administration of free CDDP and TM _(CDDP/PTX). (B) Tissue distribution of platinum concentration in the plasma on day 2 after i.v. administration of free CDDP and PB-CDDP-PTX. Each drug was administered to Nude mice bearing human SKOV3 ovarian cancer tumor (female, n = 3) at a dose of 6 mg/kg on CDDP basis. Data were expressed as mean ± SE (**p < 0.01; ***p < 0.005).

Fig. 9

(A) The body weight changes for animals treated with TM_{(CDDP/PTX = 2:1)} at two dosage levels, e.g. 4 and 6 mg CDDP/kg for three dosage in MTD study; (B) the body weight changes of animals treated with a single dose of free drug mixture of CDDP/PTX 2:1 w/w and TM_{(CDDP/PTX = 2:1)} at 10 mg CDDP/kg level in comparison with PBS control group; (C) the serum AST and ALT enzyme levels and BUN level of animals in the acute toxicity studies treated with free CDDP/PTX and TM_{(CDDP/PTX = 2:1)}, respectively, at 10 mg CDDP/kg level; The blood cell counting analysis for WBC (D), PLT (E) and RBC (F) for the mice in MTD and acute toxicity studies. (*p < 0.05).

Fig. 10

Histopathological changes in kidney and liver from the acute toxicity studies in BALB/c mice on day 7 after being treated with PBS (control), free CDDP/PTX and TM_{(CDDP/ PTX = 2:1)} at 10 mg CDDP/kg. Tissues were stained with fixed with 4% paraformaldehyde and stained with hematoxylin and eosin (H&E). Significant tubular dilation with flattening of the epithelium cells were observed only in the kidneys in mice treated with free CDDP/PTX mixture. No abnormal structures were observed in livers.

Fig. 11

In vivo tumor growth inhibition (A), body weight changes (B) and KaplaneMeier survival curve (C) of mice beard SKOV-3 ovarian cancer xenografts (n = 5) after intravenous treatment with different CDDP and PTX formulations (on day 0, 4, 8). Figure B and C share the same legends with Figure A. (**p < 0.01; ***p < 0.001).

Scheme 1

The structure of telodendrimer PEG^5K(COOH)₈-L-CA₈.