Design of a Novel Theranostic Nanomedicine (III): Synthesis and Physicochemical Properties of Tumor-Targeting Cisplatin Conjugated to a Hydrophilic Polyphosphazene.

PURPOSE: A new theranostic nanomedicine involving anticancer-active cisplatin moiety was designed to study its tumor-targeting properties as well as its drug efficacy and toxicity. METHODS: A cisplatin carrier polymer was prepared by grafting equimolar polyethylene glycol of a molecular weight of 550 (PEG550) and aminoethanol to the poly(dichlorophosphazene) backbone. Cisplatin was conjugated to the carrier polymer using cis-aconitic acid as a linker. RESULTS: The cisplatin-loaded polyphosphazene, named "Polycisplatin" was found to be amphiphilic in aqueous solution and self-assembled into nanoparticles with an average particle size of 18.6 nm in diameter. The time-dependent organ distribution study of Cy5.5-labeled Polycisplatin in the A549-tumor-bearing mice exhibited a high tumor selectivity of Polycisplatin by EPR effect despite the relatively small particle size. In order to compare the in vivo efficacy of Polycisplatin and cisplatin, their xenograft trials were performed using nude mice against the human gastric cell line MKN-28. Polycisplatin exhibited slightly less tumor suppression effect compared with cisplatin at the same dose of 1.95 mg Pt/kg, which is the maximum tolerate dose of cisplatin, but at the higher double dose of 3.9 mg Pt/kg, Polycisplatin exhibited a little better efficacy than cisplatin. Furthermore, mice treated with cisplatin at the dose of 1.95 mg Pt/kg exhibited severe body weight decrease by about 25%, while mice treated with Polycisplatin did not show serious body weight decrease even at its double dose of 3.9 mg Pt/kg. Furthermore, kidney indicators including kidney index, BUN, and creatinine values measured displayed that Polycisplatin is much less nephrotoxic than cisplatin. CONCLUSION: Nanoparticular Polycisplatin was successfully prepared by conjugating cisplatin to a hydrophilic polyphosphazene carrier polymer using the acid-cleavable cis-aconitic acid. Polycisplatin nanoparticles exhibit excellent tumor-targeting properties by EPR effect. The xenograft trials exhibited excellent antitumor efficacy and reduced systemic toxicity of Polycisplatin.

Design of theranostic nanomedicine (II): synthesis and physicochemical properties of a biocompatible polyphosphazene-docetaxel conjugate.

To prepare an efficient theranostic polyphosphazene-docetaxel (DTX) conjugate, a new drug delivery system was designed by grafting a multifunctional lysine ethylester (LysOEt) as a spacer group along with methoxy poly(ethylene glycol) (MPEG) to the polyphosphazene backbone ([NP]n), and then DTX was conjugated to the carrier polymer using acid-cleavable cis-aconitic acid (AA) as a linker. The resultant polyphosphazene-DTX conjugate, formulated as [NP(MPEG550)3(Lys-OEt)(AA)(DTX)]n and named "Polytaxel", exhibited high water solubility and stability by forming stable polymeric micelles as shown in its transmission electron microscopy image and dynamic light scattering measurements. Another important aspect of Polytaxel is that it can easily be labeled with various imaging agents using the lysine amino group, enabling studies on various aspects, such as its organ distribution, tumor-targeting properties, pharmacokinetics, toxicity, and excretion. The pharmacokinetics of Polytaxel was remarkably improved, with prolonged elimination half-life and enhanced area under the curve. Ex vivo imaging study of cyanine dye-labeled Polytaxel showed that intravenously injected Polytaxel is long circulating in the blood stream and selectively accumulates in tumor tissues. Polytaxel distributed in other organs was cleared from all major organs at ~6 weeks after injection. The in vitro study of DTX release from the carrier polymer showed that >95% of conjugated DTX was released at pH 5.4 over a period of 7 days. Xenograft trials of Polytaxel using nude mice against the human gastric tumor cell line MKN-28 showed complete tumor regression, with low systemic toxicity. Polytaxel is currently in preclinical study.

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

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 1/2ß) 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.

Synthesis and properties of a new micellar polyphosphazene-platinum(II) conjugate drug.

Aiming at tumor targeting delivery of oxaliplatin using polymer therapy, a new monomeric platinum(II) complex (dach)Pt[HEDM] (dach: trans-(±)-1,2-diaminocyclohexane; HEDM: 2-hydroxyethoxydiethylmalate) was designed to include the antitumor moiety (dach)Pt and HEDM as a linker to the polyphosphazene backbone. This monomeric Pt-complex could easily be grafted to the PEGylated polyphosphazene backbone to prepare a novel polyphosphazene-Pt conjugate, [NP(MPEG550)(dach)Pt(EM)]n [MPEG550: methoxy poly(ethylene glycol) with an average molecular weight of 550; EM: ethoxymalate]. This amphiphilic polyphosphazene-Pt conjugate was found to self-assemble into stable polymeric micelles of a mean diameter of 130nm, which is suitable for passive tumor targeting by enhanced permeability and retention (EPR) effect. Pharmacokinetic study of this polymer conjugate exhibited long blood circulation as expected and longer half-life (t1/2ß=9.52h) compared with oxaliplatin (3.47h), and much larger AUC (area under the curve) value (25,831ng·h/mL) compared with oxaliplatin (1194ng·h/mL). Biodistribution study of the polymer conjugate has shown excellent tumor selectivity with the tumor to tissue ratio of 3.84 at 2h post injection and 11.7 at 24h post injection probably due to the EPR effect of the polymer conjugate while no tumor selectivity was observed for monomeric oxaliplatin. Furthermore, accumulation of this polymer conjugate in kidney was much lower compared with oxaliplatin. Also the nude mouse xenograft trial of the polymer conjugate has shown higher antitumor efficacy compared with oxaliplatin.

Synthesis and physicochemical properties of new tripodal amphiphiles bearing fatty acids as a hydrophobic group.

Saturated fatty acids (FA) were grafted using tyrosine as a spacer group to the cyclotriphosphazene ring along with equimolar hydrophilic methoxy poly(ethylene glycol) (MPEG) in cis-nongeminal way. Seven new cyclotriphosphazene amphiphiles were prepared from combinations of hydrophilic MPEGs with different molecular weights of 350, 550, 750 and 1000 and four different fatty acids of different hydrophobicity including lauric, myristic, palmitic and stearic acids. These steric amphiphiles bearing fatty acids as a hydrophobic group were found to form more stable micelles with very low critical micelle concentrations (CMC) (2.95-7.80mg/L) compared with oligopeptide analogues, and their highly hydrophobic core environment is unique and potentially useful for various biomedical applications.