A two-pronged approach against glioblastoma: Drug repurposing and nanoformulation design for in situ‑controlled release

Glioblastoma (GB) is one of the most lethal types of neoplasms. Its biologically aggressive nature and the presence of the blood-brain barrier (BBB) limit the efficacy of standard therapies. Several strategies are currently being developed to both overcome the BBB and deliver drugs site-specifically to tumor cells. This work hypothesizes a two‑pronged approach to tackle glioblastoma: drug repurposing with celecoxib and a nanoformulation using ultra-small nanostructured lipid carriers (usNLCs).

Celecoxib antitumor druggable activity was inspected bioinformatically and screened in four glioma cell lines aiming at the comparison with temozolomide, as standard of care. Delving into formulation design, it was tailored aiming at (i) improving the drug solubility/loading properties, (ii) assigning a thermal-triggerable drug release based on a lipid matrix with a low melting point, and (iii) enhancing the cytotoxic effect by selecting a template targetable to tumor cells. For this purpose, an integrated analysis of the critical material attributes (CMAs), critical process parameters (CPPs) and critical quality attributes (CQAs) was conducted under the umbrella of a Quality by Design approach.

CMAs that demonstrate a high-risk level for the final quality and performance of the usNLCs include the drug solubility in lipids (solid and liquid), the lipid composition (envisioning a thermoresponsive approach), the ratio between lipids (solid vs. liquid), and the surfactant type and concentration. Particle size was shown to be governed by the interaction lipid-surfactant followed by surfactant type. The drug encapsulation did not influence colloidal characteristics, making it a promising carrier for lipophilic drugs. In general,usNLCs exhibited a controlled drug release during the 72 hours at 37 ºC with a final release of ca. 25 %, while at 45 ºC this was doubled.

The in vitro cellular performance depended on the surfactant type and lipid composition, with the formulations containing a sole solid lipid (Suppocire^® NB) and Kolliphor^® RH40 as surfactant being the most cytotoxic. usNLCs with an average diameter of ca. 70 nm and a narrow size distribution (PdI lower than 0.2) were yielded, exhibiting, high stability, drug protection, sustained and thermo‑sensitive release properties, and high cytotoxicity to glioma cells, meeting the suitable CQAs for parenteral administration. This formulation may pave the way to a multi-addressable purpose to improve address GB treatment.

2. Materials

Polysorbate 80 (Tween® 80), Kolliphor® RH40, IR780, and octadecylamine were provided by Sigma-Aldrich (Missouri, USA). Oleic acid was acquired from Fluka (Buchs, Switzerland). Labrasol® (Caprylocaproyl Polyoxyl-8 glycerides), Compritol® 888 ATO (glyceryl dibehenate), Suppocire®NB (mono-, di- and triglyceride esters of C10 to C18 fatty acids, the triester fraction being predominant), Capryol® 90 (propylene glycol monocaprylate), CapryolTM PGMC (propylene glycol monocaprylate-type I), Capmul MCM C8, Labrafil, and Lauroglycol 90 were kindly offered by Gattefossé (Gennevilliers, France). Lipoid S 75® (soy phospholipid) was provided by Lipoid GmbH (Ludwigshafen, Germany).

Dulbecco’s Modified Eagle’s Medium (DMEM), fetal bovine serum (FBS), 0.4% trypan blue solution, trypsin-EDTA solution, sodium bicarbonate, phosphate-buffered saline (PBS), penicillin, and streptomycin were obtained from Sigma-Aldrich (Missouri, USA). Five glioma cell lines were acquired from the American Type Culture Collection (ATCC, Manassas, VA, USA). A172 (ATCC CRL-1620), H4 (ATCC HTB-148) were used as non-tumorigenic cells, and U118 (ATCC HTB-15) and U87 (ATCC HTB-14) were used as tumorigenic cells. Ultrapure water (HPLC grade, >18.2 MΏ was prepared using a Milli-Q water apparatus (Millipore®, USA) and filtered through a 0.22μm nylon filter before use. All other reagents and solvents were of analytical or high-performance liquid chromatography (HPLC)
grade.

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