(F-202) In vivo mechanochemical dynamic cancer therapy using focused ultrasound-triggered mechanophores

Introduction::

Photodynamic therapy (PDT) is a localized radical therapy that uses light and photosensitizers to generate reactive oxygen species (ROS), proximate to cancer cells. PDT is non-invasive, precisely targeted, and without long-term side effects but is limited by the relatively shallow depth of light penetration in tissues. Sonodynamic therapy (SDT) refers more specifically to sensitizer-dependent sonochemical or sonophotochemical events in an acoustic field that leads to cytotoxicity. It uses an ultrasound-sensitive drug (sensitizers) to treat deep-seated tumors with high-energy ultrasound pulses but suffers from potential tissue thermal damage due to inertial cavitation, low precision, and relatively high energy input.  

Here we develop a new class of nanoparticle sensitizers, termed mechanochemical sensitizers (MeCS), that can be activated and kill tumors with low-energy high intensity focused ultrasound (HIFU) pulses. We report MeCS nanoparticles that are poly(Poly(Ethylene Glycol) Methacrylate) (PPEGMA) grafted silica particles with azo groups incorporated at silica particle-polymer interface. The azo group is a well-studied mechanophore. Upon incorporation in the middle of the polymer chain or at the interface of a polymer and a particle, it can be activated to generate reactive oxygen species (ROS) under force field in the presence of water and oxygen. Utilizing this effect, we find that these azo based MeCS show cytotoxicity under low power HIFU treatment. Finally, the MeCS shows therapeutic effects in an in vivo mouse tumor model using intratumor injection.

Materials and Methods::

The synthesis and characterization of particles involve evaluating their size, morphology, composition, chemical bonds, and ROS production using techniques like TEM, thermal gravimetric analysis, Raman spectroscopy, and a peroxide enzyme ROS assay kit. We utilized a high-intensity focused ultrasound (HIFU) system to activate these particles.

 

We proceeded with in vitro studies using 4T1 cells as a tumor model. Ultrasound was used to activate MeCS particles in cells placed in a 24-well plate. We assessed cell viability through the alamarBlue assay and Calcein AM/PI staining.

 

The research was then extended to in vivo studies, conducted on BALB/c mice with 4T1 tumors. We treated the mice with either MeCS or control particles, followed by irradiation with HIFU. The therapeutic efficacy was evaluated using MRI or BLI imaging, tumor volume and weight measurements, and histopathological analysis. Systemic toxicity and biodistribution were examined via fluorescence imaging or ICP-MS. These findings collectively enhance our comprehension of the potential application of MeCS particles in tumor therapy.

 

Results, Conclusions, and Discussions::

We created MeCS nanoparticles that are poly(Poly(Ethylene Glycol) Methacrylate) (PPEGMA) grafted silica particles with azo groups incorporated at the silica particle-polymer interface. MeCS particles, ~55 nm in diameter.

 

Moreover, MeCS alone shows excellent biocompatibility in the absence of HIFU treatment. We have also proved that the activated MeCS produces sufficient ROS to cause the death of 4T1 cancer cell lines. More importantly, the energy density we have used was more than five times lower than the typical energy density reported in SDT, which further decreases the side effect of traditional HIFU treatment. In the in vitro experiments, the fluorescence outputs from the AlamarBlue assay and Calcein AM/PI showed that the treatment with MeCS + HIFU significantly reduced cell viability, while the control group was hardly affected.

 

Finally, the MeCS shows therapeutic effects in an in vivo mouse tumor model using intratumor injection. Group 4 ( MeCS +HIFU） shows significant tumor growth inhibition during the treatment, while the tumor volumes in other groups increased with similar growth rate. The photograph of harvested tumors after 10 days of the treatment also suggests that MeCS can inhibit the tumor growth under mild HIFU treatment. Importantly, none of the mice showed abnormal body weight changes during the treatment.

 

In summary, we provided a demonstration that MeCS particles with Azo group can release ROS to kill cancer cells under both in vivo and in vitro conditions. This injectable nanoparticle expands the potential of MDT as a non-invasive cancer therapy. In future studies, we consider further development of tumor hosting nanoparticles that can be systematically administered, combined with the high penetration of HIFU, it can precisely treat deep-seated tumors.

 

Acknowledgements (Optional): :

References (Optional): : Kim, G., Wu, Q., Chu, J. L., Smith, E. J., Oelze, M. L., Moore, J. S., & Li, K. C. (2022). Ultrasound controlled mechanophore activation in hydrogels for cancer therapy. Proceedings of the National Academy of Sciences, 119(4), e2109791119.