Innovative Rocuronium Bromide Topical Formulation for Targeted Skin Drug Delivery: Design, Comprehensive Characterization, In Vitro 2D/3D Human Cell Culture and Permeation

Abstract

Cutaneous squamous cell carcinoma (cSCC) is the second-most common type of non-melanoma skin cancer and is linked to long-term exposure to ultraviolet (UV) radiation from the sun. Rocuronium bromide (RocBr) is an FDA-approved drug that targets p53-related protein kinase (PRPK) that inhibits the development of UV-induced cSCC. This study aimed to investigate the physicochemical properties and in vitro behavior of RocBr. Techniques such as thermal analysis, electron microscopy, spectroscopy and in vitro assays were used to characterize RocBr. A topical oil/water emulsion lotion formulation of RocBr was successfully developed and evaluated. The in vitro permeation behavior of RocBr from its lotion formulation was quantified with Strat-M® synthetic biomimetic membrane and EpiDerm™ 3D human skin tissue. Significant membrane retention of RocBr drug was evident and more retention was obtained with the lotion formulation compared with the solution. This is the first systematic and comprehensive study to report these findings.

Comprehensive Physicochemical Characterization, In Vitro Membrane Permeation, and In Vitro Human Skin Cell Culture of a Novel TOPK Inhibitor, HI-TOPK-032

“…The thermal transitions of raw HI-TOPK-032 were determined using a Discovery Differential Scanning Calorimeter 250 with ultra-high purity (UHP) nitrogen gas (Airgas, Air Liquide, Palm Beach, FL, USA) with a flow rate of 50 mL/min and TRIOS v5.6.0.87 software for analysis (DSC250 with T-Zero ® Technology (TA Instruments, New Castle, DE, USA). As described in previously published methods [27,28], briefly, 2-5 mg of the sample was packed into hermetic anodized aluminum T-Zero ® DSC pan, and aluminum lids were hermetically sealed using a T-Zero ® hermetic press (TA Instruments, New Castle, DE, USA). The reference pan was an empty, hermetically sealed T-Zero ® aluminum pan.…”

“…The crystallinity of raw HI-TOPK-032 powder was determined by X-ray powder diffraction (XRPD) analysis using similar conditions to those reported previously [17,26,28]. XRPD patterns of raw HI-TOPK-032 powder were recorded using a PANalytical X'pert diffractometer (PANalytical Inc., Westborough, MA, USA) equipped with a programmable incident beam slit and an X'celerator detector at room temperature.…”

“…Following previously published growth conditions and methods [17,27,28,32,38], Ha-CaT cells (immortalized normal human keratinocyte cell line) were grown in collagen Type I-coated (concentration 5-10 µg/cm 2 in PBS) T-75 flasks using Advanced Dulbecco's Modified Eagle's Medium (ADMEM) 1× supplemented with 10% (v/v) FBS, 0.2% v/v Fungizone (0.5 µg/mL Amphotericin B, 0.41 µg/mL Sodium Deoxycholate), and 1% v/v Pen-Strep (100 Unit/mL Penicillin, 100 µg/mL Streptomycin) in a humidified incubator at 37 • C and a 5% CO 2 atmosphere. At 90% confluence, the HaCaT cells were seeded into a 96-well Black/Optical Bottom Plate at a density of 5000 cells/well in 100 µL of supplemented ADMEM, followed by incubation at 37 • C and 5% CO 2 for 48 h to allow the attachment of cells to the plate surface.…”

“…The in vitro permeation behavior of HI-TOPK-032 from a propylene glycol solution was determined using Franz diffusion cells (PermeGear, Inc., Hellertown, PA, USA), following similar conditions to those reported previously [17,28]. The Strat-M ® membrane (Millipore Sigma, Danvers, MA, USA), a synthetic, non-animal-based transdermal diffusion test model membrane that is predictive of diffusion through human skin, was mounted between two O-rings with an orifice of 0.64 cm 2 and sandwiched between the donor and receptor chambers with a clamp.…”