Studies On Triptolide Loaded PEG-PLA Polymeric Micelles

Triptolide (TP) shows multiple pharmacological activities, such as anti-inflammatory, immuno-suppressive, anti-tumour and male anti-fertility activities. It has been usually used to treat rheumatoid arthritis (RA), leprosy and nephritis. Recently, there has been growing interest in anti-tumour researches and applications of TP due to its broad anti-tumour spectrum and potent anti-tumour activity. However, the clinical use of TP is restricted due to its poor water solubility and some toxic effects on the gastrointestinal, urogenital, cardiovascular, blood circulatory system, immuno-system, bone marrow and so on. In order to develop an intravenous formulation for clinical use to treat solid tumours, TP-PM (triptolide loaded polymeric micelles) was constructed and the anti-tumour activities of TP-PM, the influence of TP-PM on the immunity of tumour-bearing mice, its toxicities were evaluated in the present paper. Besides, a novel TP loaded transdermal delivery system, named TP loaded hydrogel-thickened microemulsion (TP-MTH) was prepared previously to enhance the efficiency of TP and decrease the toxicity of TP. The anti-inflammatory and analgesic effects of TP-PM were also investigated in this paper. The main contents of this paper are as follows:(1) A diblock copolymer (MePEG-PLA) containing one block of methoxypolyethylene glycol (MePEG) and one block of either poly (D,L-lactide) (PLA) was synthesized by a ring opening bulk polymerization by changing the monomer weight ratios in the presence of stannous octoate. The copolymer was characterized by ¹H-NMR, FT-IR, GPC and pyrene. The spectrum showed that the reaction between lactide and MePEG had occurred. The weight-average molecular weight (M_W) of these copolymers were 2.8×10⁴. The critical micelle concentrations (CMC) of the copolymer was was determined to be 8.9×10^-7 mol/L, which was much lower than that of common low-molecular weight surfactants.(2) A novel polymeric micelle system containing TP (TP-PM) was constructed by the solvent evaporation method using methoxypolyethylene glycol-poly (D,L-lactic acid)-block copolymer as the carrier and characterised using photon correlation spectroscopy, transmission electron microscopy and high-performance liquid chromatography. Results demonstrated that TP-PM had an average diameter of 78.9nm, encapsulation efficiency of 66.7%, core-shell morphology and a long-term stability.(3) The anti-tumour and immuno-modulation effects of TP-PM were evaluated in sarcoma 180-bearing mice and A2780 cells. TP-PM could significantly inhibit tumour growth via intravenous injections at dose levels of 0.0375, 0.075 and 0.15 mg/kg, and their inhibition rates were 42.5%, 46.0% and 49.9%, respectively; they showed similar cytotoxicity against A2780 cells compared to that of TP. Simultaneously, TP-PM had no effect on the thymus index, spleen index, spleen lymphocyte proliferation and the TNF-αand IL-2 levels in serum as compared with TP. Therefore, TP encapsulated in polymeric micelles does not demonstrate immuno-suppressive activity but does not lose its anti-tumour effect. These results show that polymeric micelles are a promising carrier for cancer therapy using TP.(4) The acute toxicity in mice and sub-acute toxicity in rat were studied for TP-PM. Results demonstrated that the LD₅₀ for TP-PM and TP administered intravenously were 1.06 mg/kg and 0.83 mg/kg in mice, respectively. In toxicity study in rat, TP-PM and TP were administered intravenously at the dose levels of 0.1 mg/kg and 0.3 mg/kg for 14 d. Compared to the control, there was a significant difference (P<0.05) about the serum AST, ACP levels of the testis, thymus index and spleen index in rats treated with TP, however, TP-PM had no effect on above parameters. Compared to TP, TP-PM significantly increase the ACP activity of the testis and decrease the MDA level in serum. So, TP-PM Showed lower toxicity than TP.(5) The anti-inflammatory and analgesic effects of TP-MTH were evaluated. TP-MTH (0.003%) was administered on the skin at the 0.02～0.12 mg/kg dose levels in rats and mice. TP-MTH could significantly inhibit the adjuvant-induced rat paw edema, xylene-induced mouse ear edema, carrageenan-induced rat paw edema, cotton pellet implantation in rats, acetic acid-induced writhes in mice and the hot plate-induced pain in mice. Results demonstrated that TP-MTH possessed good anti-inflammatory and analgesic activities.This study demonstrated that TP encapsulated in polymeric micelles does not act as an immuno-suppressant without losing its anti-tumour effect and demonstrated that polymeric micelles are a stable and effective drug delivery carrier of TP for the therapy of solid tumours. Good anti-inflammatory and analgesic activities of TP-MTH showed in this study, in combination with lower toxicity described by Hubei center for safety evaluation of drugs, make it a valuable system for the delivery of TP for treating rheumatoid arthritis.