Design of a novel theranostic nanomedicine: synthesis and physicochemical properties of a biocompatible polyphosphazene–platinum(II) conjugate

Abstract

To develop a theranostic nanomedicine involving the antitumor-active moiety (dach)Pt(II) (dach: trans-(±)-1,2-diaminocyclohexane) of oxaliplatin (OX), a new biocompatible polyphosphazene carrier polymer was designed by grafting with a methoxy poly(ethylene glycol) (MPEG) to increase duration of circulation in the blood and with aminoethanol (AE) as a spacer group. The antitumor (dach)Pt moiety was conjugated to the carrier polymer using cis-aconitic acid (AA) as a linker, resulting in a polymer conjugate formulated as [NP(MPEG550)(AE-AA)Pt(dach)]_n, named “Polyplatin” (PP). PP was found to self-assemble into very stable polymeric nanoparticles with a mean diameter of 55.1 nm and a critical aggregation concentration of 18.5 mg/L in saline. PP could easily be labeled with a fluorescence dye such as Cy5.5 for imaging studies. The time-dependent ex vivo image studies on organ distributions and clearance of Cy-labeled PP have shown that PP accumulated in the tumor with high selectivity by the enhanced permeability and retention effect but was cleared out from all the major organs including the liver in about 4 weeks postinjection. Another time-dependent bioimaging study on distribution and clearance of PP in mouse kidney using laser ablation inductively coupled plasma mass spectroscopy has shown that PP accumulates much less in kidney and is more rapidly excreted than monomeric OX, which is in accord with the very low acute toxicity of PP as shown by its high LD₅₀ value of more than 2000 mg/kg. The pharmacokinetic study of PP has shown that it has a much longer half-life (t_1/2β) of 13.3 hours compared with the 5.21 hours of OX and about a 20 times higher area under the curve value of 42,850.8 ng h/mL compared with the 2,320.4 ng h/mL of OX. The nude mouse xenograft trials of PP against the gastric MKN-28 tumor cell line exhibited remarkably better tumor efficacy compared with OX at the higher tolerated dose, with lower systemic toxicity.

Keywords:

• polyphosphazene
• anticancer agent
• theranostics
• nanomedicine
• oxaliplatin

Supplementary materials

Figure S1 ¹H NMR spectra of the polyphosphazene carrier polymer[NP(PEG550)(AE)(AA)]_n (A) and its (dach)Pt(II) conjugate [NP(MPEG550)(AE)(AA)Pt(dach)]_n (B).

Abbreviations:¹H NMR, proton nuclear magnetic resonance; Pt, platinum; AA, cis-aconitic acid; PEG, poly(ethylene glycol); AE, 2-aminoethanol.

Figure S2 ³¹P NMR spectra of the half PEGylated polymer [NP(MPEG550)Cl]_n (A), carrier polymer [NP(MPEG550)(AE)(AA)]_n (B), and its Pt conjugate [NP(MPEG550)(AE) (AA)Pt(dach)]_n (C).

Notes: The mole ratio of MPEG550 and AA as side groups in polymer was estimated by the integration ratio of the methoxy protons of MPEG550 appearing at 3.27 ppm and methylene protons of AA as well as aminoethanol moiety appearing at 3.41 ppm and 3.44–3.53 ppm. The ³¹P NMR spectrum of the half PEGylated polyphosphazene precursor exhibits one single sharp peak at −10.9 ppm due to the O–P–Cl fragments, but the carrier polymer and its (dach)Pt conjugate show very broad single peaks at −3.95 and −5.19 ppm, respectively, due to the phosphorus resonance of the –O–P–O–fragments.

Abbreviations: ³¹P NMR, phosphorus-31 nuclear magnetic resonance; Pt, platinum; PEG, polyethylene glycol; AA, cis-aconitic acid; AE, sodium salt of 2-aminoethanol.

Figure S3 IR spectra of [(dach)Pt(OOCCH₃)₂] (A), carrier polymer [NP(MPEG550)(AE)(AA)]_n (B), and its Pt conjugate Polyplatin [NP(MPEG550)(AE)(AA)Pt(dach)]_n (C).

Abbreviations: IR, infrared; Pt, platinum; AA, cis-aconitic acid; PEG, poly(ethylene glycol); AE, 2-aminoethanol.

Figure S4 Clearance rates of Polyplatin: time-dependent fluorescence intensities of each major organ.

Abbreviation: h, hour.

Figure S5 Time-dependent plasma concentrations of platinum after single intravenous injection of Polyplatin and oxaliplatin at the dose of 1 mg (dach)Pt/kg to male rats (plasma was pooled from three animals per treatment group): Polyplatin (■); oxaliplatin(●).

Abbreviations: h, hour; Pt, platinum.

Figure S6 Changes in body weight of CRC mice treated with Polyplatin at predetermined doses according to the OECD guideline.

Abbreviations: CRC, colorectal cancer; OECD, Organisation for Economic Co-operation and Development.

Figure S7 The body weight changes of the mice treated with Polyplatin and oxaliplatin as reference.

Acknowledgments

This study was supported by a grant of the Korean Health Technology R&D Project, Ministry of Health and Welfare, Republic of Korea (HI11C0532), by a National Research Foundation of Korea (NRF) Grant funded by the Korean Government (MOE) (2014R1A1A2055876), and by C & Pharm.

Disclosure

The authors report no conflicts of interests in this work.