Evaluation of Anticancer Efficacy of D-α-Tocopheryl Polyethylene-Glycol Succinate and Soluplus® Mixed Micelles Loaded with Olaparib and Rapamycin Against Ovarian Cancer

Abstract

Purpose: Ovarian cancer has the highest mortality rate and lowest survival rate among female reproductive system malignancies. There are treatment options of surgery and chemotherapy, but both are limited. In this study, we developed and evaluated micelles composed of D-α-tocopheryl polyethylene-glycol (PEG) 1000 succinate (TPGS) and Soluplus^® (SOL) loaded with olaparib (OLA), a poly(ADP-ribose)polymerase (PARP) inhibitor, and rapamycin (RAPA), a mammalian target of rapamycin (mTOR) inhibitor in ovarian cancer.

Methods: We prepared micelles containing different molar ratios of OLA and RAPA embedded in different weight ratios of TPGS and SOL (OLA/RAPA-TPGS/SOL) were prepared and physicochemical characterized. Furthermore, we performed in vitro cytotoxicity experiments of OLA, RAPA, and OLA/RAPA-TPGS/SOL. In vivo toxicity and antitumor efficacy assays were also performed to assess the efficacy of the mixed micellar system.

Results: OLA/RAPA-TPGS/SOL containing a 4:1 TPGS:SOL weight ratio and a 2:3 OLA:RAPA molar ratio showed synergistic effects and were optimized. The drug encapsulation efficiency of this formulation was > 65%, and the physicochemical properties were sustained for 180 days. Moreover, the formulation had a high cell uptake rate and significantly inhibited cell migration (**p < 0.01). In the in vivo toxicity test, no toxicity was observed, with the exception of the high dose group. Furthermore, OLA/RAPA-TPGS/SOL markedly inhibited tumor spheroid and tumor growth in vivo.

Conclusion: Compared to the control, OLA/RAPA-TPGS/SOL showed significant tumor inhibition. These findings lay a foundation for the use of TPGS/SOL mixed micelles loaded with OLA and RAPA in the treatment of ovarian cancer.

Introduction

Ovarian cancer, which ranks first in mortality among malignant tumors of the female reproductive system, has the lowest survival rate of all gynecologic malignancies despite advances in diagnosis and treatment.1,2 Specifically, epithelial ovarian cancer represents about 90% of all malignant ovarian tumors.3 The prevailing treatment approach for this type of cancer typically involves cytoreductive surgery coupled with chemotherapy based on platinum compounds.4,5 Although these treatments have good initial responses rates, the recurrence rate is as high as 70%,6 and chemotherapy is limited by drug resistance7 and side effects.8

Poly(ADP-ribose) polymerase (PARP) inhibitors, which target the PARP protein,9,10 selectively target tumor cells that are unable to repair DNA double-strand breaks,11,12 and can enhance neoantigen expression to generate an antitumor immune response.13,14 Currently, inhibitors targeting PARP have gained approval for use in treating various cancers, including ovarian,15 breast,16 and pancreatic,17 with several clinical trials enrolled. Olaparib (OLA) is a prime example of a PARP inhibitor and was the first monotherapy approved by the FDA for the treatment of BRCA-mutated ovarian cancer.18 Cells with mutated BRCA function have a homologous recombination (HR) deficiency, which is reported to be present in a significant proportion of non-BRCA-mutated ovarian cancers.19,20 Despite the high therapeutic efficacy of OLA, its oral bioavailability is low due to its low solubility and permeability.21,22

The mammalian target of rapamycin (mTOR), a serine/threonine kinase, orchestrates cellular growth, proliferation, and viability.23,24 Notably, the mTOR signaling pathway may become excessively active in and lead to tumor development, including ovarian cancer.25,26 Because rapamycin (RAPA) regulates the translation of mRNA, it delays cell cycle progression and thus inhibits cell proliferation.27 Therefore, RAPA is considered a potential therapeutic agent to inhibit tumor growth.28,29 However, like OLA, RAPA, also demonstrates the disadvantages of low solubility and bioavailability.30

Using polymeric micelle formations presents a plausible method for augmenting the solubility and stability of hydrophobic medications31,32 Polymeric micelles exhibit high solubility,33,34 loading capacity,35 blood flow stability,36,37 and therapeutic potential.38 D-α-tocopheryl polyethylene-glycol (PEG) 1000 succinate (TPGS) is a natural water-soluble derivative of vitamin E,39,40 and increases drug solubility and bioavailability.41,42 In addition, it is used as an anticancer agents, as it induces apoptosis and exhibits synergistic effects with other anticancer agent.43 TPGS exhibits anticancer properties akin to those of α-tocopheryl succinate (TOS), with heightened efficiency in triggering apoptosis and producing reactive oxygen species when compared to TOS.44 Furthermore, TPGS suppresses heightened P-glycoprotein (P-gp) expression,45–47 a factor significantly implicated in the emergence of multidrug resistance (MDR) cancerous cells.48,49 While TPGS offers numerous benefits, it possesses a notably high critical micelle concentration (CMC) of 0.02wt%, which may lead to its dissociation in the bloodstream.39 Soluplus® (SOL), a polyvinyl caprolactam–polyvinyl acetate–polyethylene glycol graft copolymer (PCL-PVAc-PEG), is an amphipathic copolymer. As an amphiphilic substance, SOL can improve the bioavailability of poorly soluble drugs and can self-assemble into micelles above the CMC.50 Mixed micelles can decrease CMC values,51 and increase drug activity while decreasing its cytotoxicity.52 Moreover, mixed micelles offer benefits such as improved micelle stability and drug encapsulation efficiency.53,54 Several studies of mixed micellar systems have also achieved clinical-stage research status.54 Studies on various combinations of TPGS and SOL have also been reported.55,56

Figure 1 Schematic diagram of (A) micelle preparation of TPGS, SOL, OLA, and RAPA and (B) representation of the behavior of OLA/RAPA-TPGS/SOL in cancer cells

Tumor spheroids are three-dimensional (3D) structures of cancer cells that closely mimic solid tumors in living organisms, replicating their structural composition and dynamic microenvironment.57 They facilitate cell-matrix interactions, a feature difficult to achieve in traditional two-dimensional (2D) cell cultures.58 Additionally, their use reduces the need for animal experimentation, making them an ethical and efficient platform for drug screening and evaluation.59,60 In this research, we focused on the development and assessment of innovative nanoformulations. These nanoformulations were engineered for enhanced permeability and retention (EPR) by encapsulating a combination of synergistic drugs,61,62 OLA and RAPA, within mixed micelles. The micelles were formulated using two biocompatible copolymers, namely TPGS and SOL. The unique combination of TPGS, SOL, OLA, and RAPA was investigated for the first time in this study, revealing promising potential as a therapeutic approach for the treatment of ovarian cancer (Figure 1).

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Materials and Reagents

OLA and RAPA were procured from LC Laboratories^® (Woburn, MA, USA). TPGS, coumarin 6 (C6), thiazolyl blue tetrazolium bromide (MTT), tert-butanol, triton X-100, sodium hydroxide (NaOH) solution, Cremophor EL^® and dimethyl sulfoxide (DMSO) were procured from Sigma-Aldrich (St. Louis, MO, USA). The SOL was kindly provided by BASF (Ludwigshafen, Rhineland-Palatinate, Germany). Ethanol (EtOH) was procured from Honeywell Burdick & Jackson (Muskegon, MI, USA). Acetonitrile was purchased from Thermo Fisher Scientific (Waltham, MA, USA). Distilled water (DW) of high purity was acquired from Tedia company (Fair-field, OH, USA). All other chemicals and reagents utilized in this study were of analytical reagent grade.

Shin YB, Choi JY, Yoon MS, Yoo MK, Shin DH, Lee JW. Evaluation of Anticancer Efficacy of D-α-Tocopheryl Polyethylene-Glycol Succinate and Soluplus® Mixed Micelles Loaded with Olaparib and Rapamycin Against Ovarian Cancer. Int J Nanomedicine. 2024;19:7871-7893
https://doi.org/10.2147/IJN.S468935