Development of an Improved Fabrication Method for an Ultrasound-Sensitive Platform for Drug Delivery to Hypoxic Tumor Sites
Authors:   , Vaidya Purva , Wheatley Margaret A. 
Published: July 2021

The efficacy of cancer treatments such as chemotherapy and radiation therapy can be limited by the presence of hypoxic regions within the tumor, leading to poor clinical outcomes. Our group has developed SE61_[O2], a microbubble comprised of a Span 60 (sorbitan monostearate) and water-soluble vitamin E (TPGS) shell with an oxygen gas core, for delivery of oxygen to hypoxic tumor sites. In vivo studies have shown that mice receiving SE61_[O2] with ultrasound followed by radiation have decreased tumor growth and increased survival, suggesting the potential of SE61_[O2] to sensitize tumors to radiation therapy. However, the duration of oxygenation after administration of SE61_[O2] does not last long enough for it to be clinically viable. To increase the duration of oxygenation, our group is investigating lonidamine-loaded SE61_[O2] (SE61_[O2]-LND), to capitalize on the fact that lonidamine targets tumor metabolism and has been shown to increase sensitivity to radiation therapy. The current fabrication method for SE61_[O2]-LND required the use of methanol and resulted in low drug loading that had high inter-batch variability, leading to a need for an improved methodology. The purpose of this study was to develop an improved fabrication method of SE61_[O2] MBs for drug loading applications. The requirements for drug loaded microbubbles fabricated with the improved methodology are that they needed to meet the minimum acoustic requirements (enhancement [greater than or equal to] 15 dB and half-life [greater than or equal to] 1.5 min), have a diameter < 6 [mu]m, be able to support an oxygen gas core, and meet the minimum drug loading requirement of 2 [mu]g/mL. Since TPGS, one of the shell-forming molecules, forms micelles at a low critical micelle concentration, it was hypothesized that we could use these micelles to solubilize lonidamine, as it is a hydrophobic drug, and capitalize on the fact that the concentration of TPGS used to fabricate SE61 is greater than the critical micelle concentration of TPGS. The design resulted in MBs that had acoustic and size properties that met the requirements and that were not significantly different from those made with the standard method. Furthermore, it was found that the TPGS micelle method greatly improved the yield and produced twice as many microbubbles. Initially, it was believed that this may be due to the different fabrication methods resulting in microbubbles with different shell compositions; however, NMR studies showed that the molar ratios of the two surfactants were similar in microbubbles made by either method. Next, Nile red was used as a model hydrophobic drug to assess the loading capabilities of both methods. Drug loading did not affect the acoustic or size properties of the microbubbles for either method. Both methods exceeded the minimum drug loading requirement, but microbubbles made with the TPGS micelle method had loaded twice as much Nile red per batch as the standard method. This could be attributed to the increased microbubble yield attained with the TPGS micelle method. The improved methodology was then applied to lonidamine loading. While the resulting microbubbles met all the necessary requirements, lonidamine loading was lower than expected for the TPGS micelle method based on the Nile red results. This suggests that lonidamine may not intercalate into the TPGS micelles or SE61 microbubble shell the same way Nile red does. Further studies must be completed to characterize lonidamine loading and explore alternative methods for formation of drug loaded micelles.