Alginate Sulfate Polycaprolactone Nanoparticles For 5-Fluorouracil Anti-colorectal Cancer Drug Encapsulation

Abstract

Nanoscaled drug delivery systems have great potential in cancer applications, especially for chemotherapy drugs where nanoparticles (NPs) protect the drug from rapid metabolism and increase its lifetime. The anticancer chemotherapy drug 5-fluorouracil (5-FU) has a very short half-life with high toxicity. To overcome this drawback, 5-FU is incorporated into many types of NPs such as lipid, inorganic and polymeric NPs. Alginate sulfate (AlgSulf) is a synthetic mimic of sulfated glycosaminoglycans that can enhance the NP's properties in terms of drug protection and sustained release. Moreover, previous work from our group demonstrated the anticancer properties of AlgSulf on breast cancer and lung adenocarcinoma cells. Therefore AlgSulf/polycaprolactone (PCL) nanoparticles could enhance 5-FU delivery and therapeutic effect by enhanced drug protection and sustained release and could also work in synergy with 5-FU to decrease cancer cell proliferation. In this thesis, we report for the first time the preparation and evaluation of AlgSulf/PCL polymeric NPs loading 5-FU. We prepared double emulsion NPs using high molecular weight PCL containing respectively 0.5%, 1% and 2% concentrations of AlgSulf, 0.5% pure alginate (Alg) and one without AlgSulf nor Alg (PCL NP) and characterized them by particle size, polydispersity index (PDI), zeta potential analysis, drug encapsulation efficiency (%EE), scanning electron microscopy (SEM), attenuated total reflectance (ATR), X-ray powder diffraction (XRD), and release in plates assessment. We also assessed their stability in terms of size and PDI and their effect on HCT116 human colorectal cancer cell by Sulforhodamine B (SRB) and trypan blue assays when loaded and unloaded with 5-FU. AlgSulf was also evaluated in its free form on HCT116 cells using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H- tetrazolium bromide (MTT) assay to assess AlgSulf anticancer effect. The NPs were stable up to three weeks with a size of 200 nm, low PDI, and negative zeta potential which render them suitable for drug delivery. SEM images showed well-defined spherical NPs, and the encapsulation efficiency ranged from 13.82% to 18.28%. All NP’s presented the peaks of 5-FU functional groups when assessing them using ATR, the crystal structure of all NP’s surface did not change by adding 5-FU which indicates that 5-FU is encapsulated in the NPs in an amorphous way. The SRB assay revealed a decrease in HCT116 cell growth whether in non-loaded NPs or loaded with 5-FU for up to 72 hours, with 0.5% AlgSulf loaded with 5-FU NP showing the greatest anticancer effect for a concentration of 25 μM at 72h (P<0.0001). The acquired inhibition in cell proliferation when employing the NPs was less than that of free 5-FU. This may be explained by the results of the release in plate assay which showed a slow release of PCL after 3 days of incubation for the 0.5% AlgSulf NP. The trypan blue exclusion assay confirms that utilizing 0.5% AlgSulf in the NPs core had a more potent anticancer effect compared to higher concentrations of AlgSulf. The current work demonstrates the potential of combining AlgSulf/PCL NPs with chemotherapy drugs like 5-FU for sustained drug release and enhanced cancer therapy. Additional in vitro and in vivo testing are needed to refine the NP formulation and assess its efficacy on healthy and drug-resistant cancer cells.