Enhancing Anti-CTLA-4 Antibody Delivery to the Brain Using Focused Ultrasound and Microbubbles

Maria Afonso Pereira,1 Vanessa Drevenakova,1,2 Qiyixing Ethan Liu,1 Sarina Grewal,1– 3 Sophie V Morse1,2 1Department of Bioengineering, Imperial College London, London, UK; 2UK Dementia Research Institute Centre, Imperial College London, London, UK; 3Department of Brain Sciences, Imperial College London, London, UKCorrespondence: Sophie V Morse, Department of Bioengineering, Imperial College London, London, UK, Email [email protected]: Gliomas, particularly glioblastomas, are characterised by their poor prognosis and low patient survival rate. Cytotoxic T lymphocyte-associated antigen-4 antibodies, a type of immune checkpoint inhibitor, have shown promise as an effective treatment strategy against the most aggressive tumour cells within the microenvironment of gliomas. However, their delivery to the brain is hindered by the blood-brain barrier, which is leaky in a heterogenous way, leading to uneven drug distribution across the tumour. Focused ultrasound, combined with intravenously administered microbubbles, is a technique that can non-invasively, safely, and reversibly increase the permeability of the blood-brain barrier in a targeted area, promoting the delivery of therapeutics, such as immune checkpoint inhibitors, to inaccessible tumour areas and thereby helping to prevent tumour relapse.Methods: We applied two different types of focused ultrasound sequences (long pulses vs rapid short-pulses) with microbubbles to the left hippocampus of wild-type female C57BL/6 mice before administering fluorescently labelled cytotoxic T lymphocyte-associated antigen-4 antibodies.Results: We determined that the targeted brain region had a significant increase in antibody delivery following ultrasound treatment with both pulse sequences. A more uniform delivery was achieved when treating with rapid short-pulse sequences (p-value = 0.0130), where bursts of short 5 μs pulses of focused ultrasound were emitted at a fast repetition frequency (1.25 kHz). We observed a significant increase in anti-tumour immune cells in long-pulse treated brains (p-value = 0.0021 (CD3+ cells) and p-value = 0.0001 (MHC class II+ cells)).Conclusion: These results provide a proof-of-principle for how focused ultrasound with microbubbles can promote homogenous anti-tumour drug delivery and modulate the immune microenvironment. Keywords: focused ultrasound, blood-brain barrier, microbubbles, drug delivery, immunotherapy, immune checkpoint inhibitors