The emergence of liposomes has provided a breakthrough for the clinical use of highly toxic chemotherapeutic drugs . However, the clinical effects of conventional drug-loaded liposomes are unsatisfactory due to the lack of specific targeting of cancer cells and the inability to achieve endosome escape and controllable release in vivo. In this paper, a cRGD-PEOz-Hz-DOB (cPzD) molecule with specific targeting, endosomal escape, and endogenous acid-sensitive controlled release ability was creatively designed and synthesized to prepare drug-loaded liposomes cPzDGX encapsulating the anticancer drug Doxorubicin (DOX) in order to enhance the anticancer performance of DOX. The study found that the hydrazone bond in cPzD reached a breakage rate of 42% within 48 h at pH 5.0, so the cumulative release rate of cPzDGX at pH 5.0 for 48 h was as high as 82.9%, which was 22.5% higher than that at pH 7.4. Moreover, the uptake of cPzDGX in A549/ADR cells was increased by 62% at 2 h, and the expression of the green fluorescent protein gene EGFP was also increased by 36% relative to the targeting control group, indicating that cRGD modification could effectively improve the targeting ability of liposomes to cancer cells. In addition, it was demonstrated that PEOz modification could help drug carriers achieve endosome escape because the green fluorescent protein expression in the PEOz-modified liposome group was 60.5% higher than that in the PEG-modified group. Finally, the cell viability of the cPzDGX group decreased by 17.2% relative to the free DOX group when the DOX concentration was 1 μg/mL in the cytotoxicity assay, and the diameter reduction of 3D tumor MCs increased by 34.4%. And cPzDGX could effectively enhance the permeability of free DOX to solid tumors. The above results indicate that the DOX-loaded liposomes cPzDGX we designed have great prospects for the accurate and efficient treatment of cancer.
The resistance of cancer cells to anticancer drugs has been recognized as one of the main reasons for chemotherapy failure. Multidrug combination therapy is one of the most effective ways to solve this problem. Therefore, in this article, we designed and synthesized a pH/GSH dual-responsive camptothecin/doxorubicin (CPT/DOX) dual pro-drug synergistic treatment system with the aim of overcoming the resistance of non-small cell lung cancer A549/ADR cells to DOX. The pro-drug cRGD-PEOz-S-S-CPT (cPzT) was obtained by linking CPT to poly(2-ethyl-2-oxazoline) (PEOz) with endosomal escape properties through a GSH-responsive disulfide bond and modifying it with the targeted peptide cRGD. The pro-drug mPEG–NH–N=C-DOX (mPX) was synthesized by attaching DOX to polyethylene glycol (PEG) through acid-sensitive hydrazone bonds. The dual pro-drug micelles cPzT/mPX configured according to the CPT/DOX mass ratio of 3:1 showed a strong synergistic therapeutic effect at IC50 with a combined therapy index CI = 0.49, far less than 1. Moreover, with the further improvement of the inhibition rate, the 3:1 ratio showed a stronger synergistic therapeutic effect than other ratios. The cPzT/mPX micelles not only had better targeted uptake ability but also showed a better therapeutic effect in both 2D and 3D tumor suppression assays relative to free CPT/DOX and significantly enhanced the penetration ability into solid tumors. In addition, the results of confocal laser scanning microscopy (CLSM) showed that cPzT/mPX could effectively overcome the resistance of A549/ADR cells to DOX by delivering DOX into the nucleus to exert its effect. Thus, this dual pro-drug synergistic therapy system combining targeting and endosomal escape ability provides a possible strategy to overcome tumor drug resistance.
Polyethylene glycol (PEG), which can impart long cycling performance to drug carriers, has been commonly used for drug carrier modification. However, PEG modification impedes cellular uptake of drug carriers and intracellular release of drugs, a problem known as the “PEG dilemma” that limits its application. In this study, we designed and synthesized mPEG-NCH-DOB molecules with acid-sensitive hydrazone bonding for the responsive removal of PEG. Drug-loaded lipid PHDGX for doxorubicin (DOX) delivery was obtained by compounding mPEG-NCH-DOB with lipid DOPG, which possess the capability for long cycling, and acid-sensitive PEG-removal to accelerate drug release and enhance cellular uptake. mPEG-NCH-DOB could reach 50% fracture rate after 24 h at pH 5.0. The cytotoxicity assay and flow cytometry (FCM) results showed that PHDGX could enter A549/ADR cells more easily than free DOX at low DOX concentrations and has stronger anti-cancer ability. Through a confocal laser-scanning microscopy (CLSM) study, PHDGX was found to enter the nucleus of A549/ADR cells more easily than free DOX at the same time point, which indicated that PHDGX could overcome the DOX resistance of A549/ADR at low DOX concentrations. The 3D tumor microsphere (MC) growth inhibition assay showed that PHDGX not only has the best MC inhibition ability but also has better MC penetration ability than free DOX. Therefore, this drug-loaded liposome has good stability under conventional conditions and responsive PEG removal in the tumor acid environment, offering a potential strategy for utilizing PEG in drug carrier modification and overcoming tumor resistance to free drugs.
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