Channel: Focused Ultrasound Foundation clear
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| 1 | Focused Ultrasound Foundation | Cancer Immunotherapy Workshop 2021 – Focused Ultrasound Mediated Immun... | 248 | 2 | 42.6 | 12:21 | Hello everyone. My name is Pavlo Sanasda Sia Des from the Translational Ferributics Research Group at the University of Maryland School of Medicine. I would like to express my warmest thanks to the focused ultrasound foundation and the cancer research institute for hosting it another focused ultrasound and cancer immunotherapy workshop. I have been tasked to give an overview of the FUS immunomodulation consortium efforts. Within this consortium we explored the option of a synergistic effect between FUS and immune responses in a preclinical glioma model. This is a multi-center effort that started a couple of years ago and included six different sites across the United States and Canada. I will take a minute or two to provide background for some colleagues who may have joined recently or who may not be acquainted with the field of brain tumor biology or immunology. Each site was asked to follow the same experimental protocol in terms of tumor implantation and downstream assays. Each one of the six participating centers applied a different mode of FUS to study immune responses in this glioma-muring model. We're going to see more detail this individual modalities later in the presentation. A few words to the immune environment in the setting of glioblastoma. The glioblastoma has been extensively studied as a paradigm for cancer associated immunosuppression. Systemic immunosuppression in GBM has been demonstrated as evidence by impaired cellular immunity in patients. The glioblastomas have a positive infiltrating T-cells and harbor a relatively low number of somatic mutations compared with other solid tumor types. These tumors profoundly affect the immune system both locally and systemically. In addition to tissue resident myeloid cells or microglam, evidence suggests migrating myeloid cells have an important role in glioblastoma associated immunosuppression. The glioblastoma microenvironment is a highly immunosuppressive environment of tumor and immune cells. The lymphoid compartment also contributes to the immunomodulating environment with regulatory T-cells in particular mediating immunosuppressive effects through upregulation of various soluble factors in immune checkpoint molecules and metabolic pathways. The drug cells in the GBM tumor microenvironment can traffic by the tumor draining lymph nodes of the brain to the deep cervical lymph nodes and can present endogen to promote an adaptive endotumor immune response, although this process might be abrogated in the context of the systemic immunosuppression that is intrinsically associated with glioblastoma and can also be potentially added by the current standard of care treatments for this disease. For example, systemic thymazolamide chemotherapy induces a lymphopenia that is exacerbated by bone marrow sequenstration of T-cells. A few words to the experimental protocol in the timeline. Luciferase transduced GL261 cells were stereotyically implanted in a striatum of the brain of albino female mice six to ten weeks old. After cell implantation, tumor growth was monitored by bioluminescence and MRI imaging. On day 14 post implantation, the animals were prepared for a US treatment with one of the six modalities used in a consortium. Treatments were administered in principle under MR image guidance after core registration. On day 21 post implantation, a US treated end control mice were euthanized and their brains, spleens and cervical lymph nodes harvested for using downstream assets to assess changes in immune cell populations. This assets included flow cytometry, immunohistrochemistry and interferon gamma production. In the next slide, you are going to see the participating centers and which modality each one of these centers applied. Moving to the blood brain barrier opening modality. For a more guided focused ultrasound blood brain barrier opening, mice were catheterized for intravenous injections of MRI contrast agent and microbubbles. Mice were treated with a 1.14 megahertz spherical single element transducer within an Amar compatible FUS system after core registration. MRI contrast agent was administered intravenously to confirm tumor location by contrast enhanced T1 weighted amar imaging. A first spot grid of sonications was overlaid on the Amar visible tumor and sonications were carried out at 0.5% duty cycle for 2 minutes at 0.4 and 0.6 mega Pascal. Contrast enhanced T1 weighted amar imaging was repeated to confirm blood brain barrier opening. The combination of FUS plus microbubbles increased in treated cells maturity in the tumor, tumor draining lymph nodes and meninges. This combination also increased entaging experience PD1 expressing CD8 plus T cells in the tumor microenvironment and the percentage of CD8 plus T cells that were producing granzyme B in the superficial draining lymph nodes. Moving on to the microbascular ablation combined with blood brain barrier opening and anti PD1. Anti PD1 and body was administered in combination with FUS at 255 and 300 kilo Pascal. Although there were no effects observed at the lower FUS level, the combination of a PD1 inhibitor and FUS showed little difference to PD1 inhibitor alone for adaptive immunity. However, the combination increased activated M1 microglia in PDL1 microglia. In the spleen and a cervical lymph nodes, there was an increase in T cell immunoglobulin and I team domain receptor expression on CD3 plus and CD4 plus T cells only with a combination of FUS and checkpoint inhibitor. For thermal ablation, FUS pills were generated using a single element spherically curved air back transducer with a diameter of 25 millimeters and confoical length of 20 millimeters driven in continuous wave. The transducer had a central opening of four millimeters in diameter to facilitate hydrophone insertion. There is an unfrequency of the transducer was 1.06 megahertz but it was driven at the fifth harmonic at 5.51 megahertz to allow FUS mediated heating within the desired depth range. The gliumotumor was identified with gadolinium contrast enhanced T1 weighted in mar imaging to achieve thermal ablation and marguided focused ultrasound sonication were performed at 5.51 megahertz for 15 seconds at an acoustic power level of 2.45 watts. During sonication, a Martha momentary was performed to map the temperature rise in the brain of the focal depth of the transducer. Legion information was confirmed by assuming a conservative thermal dose threshold of 240 cumulative equivalent minutes at 43 degrees Celsius based on a Martha momentary measurements. Thermal ablation led to a slow tumor growth of 21 days post-enotulation. Finally, muonohistochemistry showed no difference in the tumor of CD4 plus and CD48 plus T cells. For pulse focused ultrasound treatments were also carried under the mar image guidance. The sonications were carried out at 1.5 megahertz and 2.3 mega Pascal. 6 to 9 spots were overlaid on the mar visible tumor and each spot was treated for 60 seconds. Regularly T cells were increased in the screen, suggesting redistribution in the cell population. In the draining lymph nodes increased in myeloid derived suppressor cells and a decrease in CD8 plus T cells were observed. In the tumor, there were no observed differences in immune cell population, 7 days post FUS. However, immunohistochemistry of tumor tissues showed increased CD plus 4 and CD8 plus T cells. For hypothermia, the tumors were located using T2 weighted MR imaging. Accurate targeting in vivo was confirmed by sonicating the tumor for 10 seconds and using a mar temperature imaging to localize the focal heating. A binary controller based on a mar temperature imaging allowed to reach the set point temperature of 41.5 Celsius within a few seconds and remained at the set temperature for a duration of 12 minutes in total at which point the treatment was concluded. FUS induced hypothermia caused changes in immune cell trafficking in brain tumors. There were significant increases in effect or CD8 plus T cells and activated natural killer cells. There were no changes observed in immune cell populations outside the brain. For histotrypcy, an 8-element transducer operating at 1 megahertz was used. The transducer had an aperture diameter of 58.6 millimeters and a focal length of 32.5 millimeters. An average of 1.5 cycle pulses were applied with a peak negative pressure of 40 megapascal were for a total of 50 pulses at a single location. Histotrypcy treatments led to a significant reduction in myeloid derived suppressor cells in the tumor and an increase in interferongamam was observed. A few closing remarks. Immunotherapy is clearly nothing short of revolutionary in the treatment of glioblastoma. Multiple findings support the hypothesis that glioblastoma is a called tumor. Glioblastomas are known to have relatively few tumor infiltrating lymphocytes compared with other tumor types suggesting that they are choiescent tumors in terms of immune reactivity. Hence, combination approaches with the aim of making these cold tumors hot and thus augmenting current immunotherapy strategies are desperately needed. The use of FUS as a local therapeutic modality to increase the availability of tumor antigens and immunotherapy to drive an anti-tumor immune response provides the rationale for this combination approach. We observed local and systemic immunologic shifts in glioma-bearing animals including both thermal and mechanical modes. This may indicate that immunotherapy combinations with FUS may be synergistic as FUS may be able to locally activate the tumor immune microenvironment to drive an anti-tumor immune response. With this, I would like to acknowledge the wonderful team that I have been fortunate to work with. In particular, I would like to acknowledge and wholeheartedly thank Kelsey Timbi, Natasha Shebani, Tyler Kerhenson, Tau Soon, Anna Staja Villalopoulou and Marc Santos. I would like to thank my mentor, Dr. Woodworth and all PIs in the consortium for their valuable support. Finally, I would like to thank again the FUS Foundation for moving the field of FUS mediated immunomodulation forward. I thank you all very much for making time to listen to this presentation. Please reach out with questions and please stay tuned as we're reaching the final stages of the manuscript preparation. I look forward to the next and hopefully to many more workshoped in future. Thank you so very much. | ↗ | ||
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| 2 | Focused Ultrasound Foundation | Cancer Immunotherapy Workshop 2021 – Moderated Discussion: Optimizatio... | 52 | 1 | 23.0 | 49:17 | No transcript | ↗ | ||
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| 3 | Focused Ultrasound Foundation | GBM Workshop 2021 – Immunomodulation | 85 | 16.6 | 38:16 | No transcript | ↗ |