(T0930-11-73) Implantable Albumin Gels for Biomimetic Delivery of Anticancer Drug for Breast Therapy

Purpose: lbumin is a natural drug carrier in the human body and occupies nearly 60% of the plasma proteome. Our previous studies investigated various dosage forms (nanoparticles, microbubbles, hydrogels, and others) for cancer therapies (1, 2). In the present work, we have studied the impact of mono and divalent salts and their concentration on gelling behavior of native and doxorubicin (DOX) loaded albumin gel. Detailed rheological and physicochemical characterization of native and DOX-loaded albumin was performed.
Methods: 1. Synthesis of Albumin and Plasma Hydrogel; We have investigated the effect of various concentrations of mono and divalent ions on the gelling nature of albumin when exposed to 85°C at different time points in a water bath. Specific quantities of DOX loaded 6% w/v of albumin (pH 7.8 to 8) and the gelling nature was also studied in various salt concentrations. 2. Self-Assembly and Rheological Evaluations; The elastic and viscoelastic nature of albumin (native and drug-loaded) and plasma gels were analyzed by frequency sweep, thixotropy, strain sweep, and shear stress versus rate measurements. The rheology of native and DOX-loaded gel (prepared using KCl, MgCl₂, and CaCl₂) was investigated. The data was compared to the rheological nature of native and DOX-loaded albumin gel prepared in 25 mM NaCl as standard. Further, native albumins gel formation through protein aggregation mediated self-assembly mechanism was also mapped using a particle size analyzer. 3. Stability studies and Drug release kinetics; Frequency sweep (storage and loss modulus) and viscosity measurement were used to study the viscoelastic nature of native and DOX-loaded gel (made in 25 mM NaCl solution) at ambient and at 37°C temperature over 31 days. This was also done for DOX-albumin gels made in other salt solutions. DOX interaction with albumin in various salt solutions while forming gel was evaluated using FTIR. Further, HPLC based assay was developed for the quantitation of DOX. Drug loading and release kinetics was performed in phosphate buffer saline (PBS) at pH 7.4.

Results: Our shows the formation of gel using 6% w/v albumin solution (in 25 mM NaCl), pH 7.8, and exposure at 85℃, for 20min. It was observed that the concentration of albumin in salt solution, and time of exposure to heat, changes the viscosity that makes the resultant gels either elastic or viscoelastic (1). For instance, increased heat exposure (to 30 minutes) causes a structured gel system with more elastic behavior (Figure 1). The use of 25 mM KCl produces a similar gelling effect as NaCl but it makes gel in 15 minutes. Whereas, the use of divalent salts like MgCl₂ and CaCl₂ makes translucent gel within 1 minute of exposure to 85℃ (Figure 2). This data suggests quicker gelling nature of albumin in the presence of divalent salts and the role of salts as ionic crosslinkers. Rheology data (frequency sweep, shear stress vs. rate, and thixotropy) show that divalent salts make relatively harder gel within a minute of exposure to 85℃ compared to monovalent salts. The DOX-loaded albumin solutions in mono-divalent salts were also investigated using rheology, FTIR, and drug release kinetics. It was observed that DOX interacts through non-covalent interaction while the formation of gels.

Conclusion: Albumin gel can be used for the preparation of both injectable and elastic hydrogels by changing salt and time of exposure to heat. This study showed that native albumin gel 6% w/v albumin gel showed viscoelastic behavior, which was comparable to the rheology of human breast tissue. Divalent salt solution of albumin 6%w/v produce gels quicker and at less time of exposure compared to monovalent salts gels. However, mono valent salts produce clear transparent gel compared to divalent salts. DOX-loaded gels were synthesized and rheology data suggests that it can serve as the localized tissue-specific delivery system for anticancer therapy.

References: 1) Ankit Rochani, Aditya Bajaj, Miriam Hernandez Meadows, Gagan Kaushal. Doxorubicin loaded Albumin and Human Plasma Hydrogels as Implant for Breast Cancer Therapy. Controlled Release Society; 2022; Montreal, Canada.
2) Ankit Rochani, Sivakumar Balasubramanian, Aswathy Ravindran Girija, Toru Maekawa, Gagan Kaushal, and D. Sakthi Kumar. Heat Shock Protein 90 (Hsp90)-Inhibitor-Luminespib-Loaded-Protein-Based Nanoformulation for Cancer Therapy. Polymers (Basel). 2020 Aug; 12(8): 1798.
3) Murata M, Tani F, Higasa T, Kitabatake N, Doi E. Heat-induced Transparent Gel Formation of Bovine Serum Albumin. Biosci Biotechnol Biochem. 1993 Jan;57 (1):43-6.

Acknowledgements:Authors of this work pose no conflict of interest. Funding for this work provided by Scholarly Innovation Grant (SIG) of Wegmans School of Pharmacy. The work as also supported by Thomas Jefferson University.

Figure 1: Injectable albumin gel was synthesized by exposing 6%w/v albumin solution (in 25mM NaCl) at 85℃ for 20min. When same solution was exposed at 85℃ for 30min (in water bath), it produces moldable spelling “HAPPY” 6% w/v albumin gel. The gel was molded using silicone molds.

Figure 2 (a): Physical morphology of albumin gel in mono and divalent ions, (b and c) Frequency sweep suggests gels from mono valent salts are less viscoelastic compared to divalent salts. G’= Storage Modulus (Pa) and G’’= Loss Modulus (Pa)