(I-323) Investigating the Penetration of Collagenase-Attached ICG Liposomes Through Gelatin Hydrogels Using Photoacoustic Imaging

Introduction:: In the US, pancreatic cancer accounts for approximately 3% of all cancer cases 1. Common forms of treatment include chemotherapeutics and radiotherapy, however, many aggressive tumors have the potential to be resistant to these treatments 2. Although chemotherapy gradually reduces the progression of tumors, common side effects include damage of heart, follicle, digestive, and other types of growing cells. Fortunately, there has been a rapid increase in cancer therapeutics in the bioengineering field. Liposomes have proven to be efficient drug delivery agents due to their encapsulation, controlled release, and biocompatibility. In order to optimize the delivery of drugs, collagenase would be a viable solution to deteriorate the collagen fibers in the dense extracellular matrices and provide greater access to the necrotic core.  The objective of this study is to determine whether the Indocyanine Green (ICG) loaded liposomes, with collagenase attached to the shell, could penetrate through a gelatin (denatured collagen) hydrogel. Using photoacoustic imaging (PA), the movement of the solution can be monitored through the encapsulated ICG.

Materials and Methods::

Liposomes: Two experimental liposomes groups are created through thin film hydration. The first group contains DPPC, cholesterol, and DSPE-PEG (2000) with an inside composition of 90:45:15, respectively. The second group consists of DPPC, cholesterol, and DSPE-PEG2000 Carboxy NHS at the same ratio. The films are rehydrated with an ICG solution at 60C, exceeding the transition temperature of DPPC (40°C). The liposomes are extruded through a 100 nm filter, and are dialyzed for 24-48 hours. 

Hydrogels: Fabrication of the hydrogel required gelatin mixed with deionized water at 40 °C, and solution is placed and poured into a petri dish to create 0.9 % w/v hydrogel. It is stored in the fridge overnight for gelation. 

Injection & Imaging: The liposomes are injected into the hydrogel using a 24 gauge needle. PA imaging is used from 680 nm to 970 nm to monitor the movement. Images are taken at 10 minute intervals until 60 minutes.

Results, Conclusions, and Discussions::

The preliminary experiments depict the proficiency of PA imaging when analyzing solutions containing ICG. Specifically, images are taken at 710 nm and 780 nm because of the high absorbance of the monomeric ICG at these wavelengths, making it easier to track the high intensity signal at different points in time. At this time, the two control groups have been tested: free ICG and free ICG and collagenase. The free ICG, as shown as Figure 1 seems to be in place for the entire hour. However, Figure 2, represents the free ICG and collagenase. It can be seen that there is movement of the ICG signal from start to finish. The percent change of the signal per area for both solutions is represented by Figure 3. Within the 60 min interval, there was a greater percentage change of signal per area for the solution with collagenase than the solution without, for both 710 and 780nm. The collagenase is shown to actively penetrate the gelatin hydrogel, resulting in greater flow through the region of interest. As these are preliminary results, further testing is required to ensure the validity of the data.

Figure 1: This image shows the movement of only ICG throughout the hydrogel for a time interval of 60 min at 710 nm and 780 nm.

Figure 2: This image shows the movement of ICG and collagenase throughout the hydrogel for a time interval of 60 min at 710 nm and 780 nm.

Figure 3: This graph shows the percent change in the movement of the ICG throughout the hydrogel. 

Conclusion. 

The preliminary results are indicative of greater flow within the collagenase solution than without, leading to the conclusion that when embedded on the surface of the liposome, collagenase would act in a similar manner. Furthermore, utilizing photoacoustic imaging to record flow parameters has been shown to be effective. For the future and by the time of the conference, our team aims to perform similar testing with non-collagenase embedded liposomes and collagenase-embedded liposomes to determine the potential benefits and limitations of the hypothesized theory, along with conducting more trials.

Acknowledgements (Optional): : University of Texas at Austin Biomedical Engineering Department

References (Optional): :

Key statistics for pancreatic cancer. American Cancer Society. (n.d.). https://www.cancer.org/cancer/types/pancreatic-cancer/about/key-statistics.html#:~:text=of%20pancreatic%20cancer-,How%20common%20is%20pancreatic%20cancer%3F,will%20die%20of%20pancreatic%20cancer. 

Winstead, E. (2023, March 17). Preventing resistance to cancer targeted therapies. National Cancer Institute. https://www.cancer.gov/news-events/cancer-currents-blog/2023/preventing-resistance-cancer-targeted-therapies