{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,13]],"date-time":"2026-10-13T00:47:27Z","timestamp":1791852447743,"version":"4.3.4"},"reference-count":42,"publisher":"MDPI AG","issue":"23","license":[{"start":{"date-parts":[[2025,11,26]],"date-time":"2025-11-26T00:00:00Z","timestamp":1764115200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"National Institute of Health","award":["R01 CA269394"],"award-info":[{"award-number":["R01 CA269394"]}]},{"name":"National Institute of Health","award":["R01 EB032772"],"award-info":[{"award-number":["R01 EB032772"]}]},{"name":"National Institute of Health","award":["R01-EB034399"],"award-info":[{"award-number":["R01-EB034399"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Cancers"],"abstract":"<jats:p>Background\/Objectives: Preclinical animal studies and clinical case reports have shown evidence of histotripsy being capable of inducing anti-tumor immune responses strong enough to inhibit tumor growth of off-target tumors. Previous studies exploring histotripsy immune stimulation have used a single therapy dose. This study aims to explore how histotripsy dose affects local tumor immune stimulation in a murine melanoma model. Methods: C57BL\/6J mice bearing subcutaneous B16F10 tumors were treated with histotripsy using an ultrasound-guided 8-element 1 MHz transducer operating at a 100 Hz pulse repetition frequency (PRF) and &gt;30 MPa peak-negative pressure. The histotripsy dose was defined by the number of pulses (8, 20, 40, or 100) per focal location (ppl). Tissue damage and residual tissue structure were measured histologically and scored by a trained pathologist. The longitudinal effect of histotripsy dosing was assessed using tumor growth and survival. Acute immune stimulation was measured at days 2 and 7 post-treatment via immunofluorescence staining of the treated tumor. Results: Histotripsy doses at 20, 40, and 100 ppl achieved significant tumor necrosis within the target region (&gt;75%), with residual structure decreasing as the dose increased. Overall, the greatest tumor control was observed in mice that received the 40 ppl dose compared to untreated mice. This correlates with the 40 ppl dose also having the largest increase in CD45+ immune cells and CD8+ T cells 7 days post-treatment compared to untreated mice. Conclusions: The effect of histotripsy dosing on immune infiltration and tumor growth highlights the significant impact of histotripsy dose on clinical effects.<\/jats:p>","DOI":"10.3390\/cancers17233773","type":"journal-article","created":{"date-parts":[[2025,11,26]],"date-time":"2025-11-26T10:04:57Z","timestamp":1764151497000},"page":"3773","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":11,"title":["Histotripsy Dose Impacts Treated Tumor Immune Infiltration and Survival Outcomes in a Murine B16F10 Melanoma Model"],"prefix":"10.3390","volume":"17","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-2508-5050","authenticated-orcid":false,"given":"Reliza","family":"McGinnis","sequence":"first","affiliation":[{"name":"Department of Biomedical Engineering, University of Michigan, Ann Arbor, MI 48109, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Brian","family":"Song","sequence":"additional","affiliation":[{"name":"Department of Surgery, University of Michigan, Ann Arbor, MI 48109, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0654-2949","authenticated-orcid":false,"given":"Hanna","family":"Kim","sequence":"additional","affiliation":[{"name":"Department of Biomedical Engineering, University of Michigan, Ann Arbor, MI 48109, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Anna","family":"Lorenzon","sequence":"additional","affiliation":[{"name":"Department of Biomedical Engineering, University of Michigan, Ann Arbor, MI 48109, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4893-1587","authenticated-orcid":false,"given":"Jiaqi","family":"Shi","sequence":"additional","affiliation":[{"name":"Department of Pathology, University of Michigan, Ann Arbor, MI 48109, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-6366-8206","authenticated-orcid":false,"given":"Lili","family":"Zhao","sequence":"additional","affiliation":[{"name":"Department of Pathology, University of Michigan, Ann Arbor, MI 48109, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9577-7626","authenticated-orcid":false,"given":"Clifford S.","family":"Cho","sequence":"additional","affiliation":[{"name":"Department of Surgery, University of Michigan, Ann Arbor, MI 48109, USA"},{"name":"Department of Cell Biology, Van Andel Institute, Grand Rapids, MI 49503, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Anutosh","family":"Ganguly","sequence":"additional","affiliation":[{"name":"Department of Surgery, University of Michigan, Ann Arbor, MI 48109, USA"},{"name":"Department of Cell Biology, Van Andel Institute, Grand Rapids, MI 49503, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0262-3044","authenticated-orcid":false,"given":"Zhen","family":"Xu","sequence":"additional","affiliation":[{"name":"Department of Biomedical Engineering, University of Michigan, Ann Arbor, MI 48109, USA"},{"name":"Department of Radiology, University of Michigan, Ann Arbor, MI 48109, USA"},{"name":"Department of Neurosurgery, University of Michigan, Ann Arbor, MI 48109, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2025,11,26]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"e233051","DOI":"10.1148\/radiol.233051","article-title":"The #HOPE4LIVER Single-Arm Pivotal Trial for Histotripsy of Primary and Metastatic Liver Tumors","volume":"312","author":"Wiggermann","year":"2024","journal-title":"Radiology"},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"259","DOI":"10.1007\/s00270-022-03309-6","article-title":"A Multi-Centre, Single Arm, Non-Randomized, Prospective European Trial to Evaluate the Safety and Efficacy of the HistoSonics System in the Treatment of Primary and Metastatic Liver Cancers (#HOPE4LIVER)","volume":"46","author":"Wah","year":"2023","journal-title":"Cardiovasc. Interv. Radiol."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"2466","DOI":"10.1016\/j.ultrasmedbio.2016.05.018","article-title":"Visualizing the Histotripsy Process: Bubble Cloud\u2013Cancer Cell Interactions in a Tissue-Mimicking Environment","volume":"42","author":"Vlaisavljevich","year":"2016","journal-title":"Ultrasound Med. Biol."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"145","DOI":"10.3109\/02656736.2015.1007538","article-title":"Histotripsy Methods in Mechanical Disintegration of Tissue: Towards Clinical Applications","volume":"31","author":"Khokhlova","year":"2015","journal-title":"Int. J. Hyperth."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"9830304","DOI":"10.34133\/2020\/9830304","article-title":"Effects of Histotripsy on Local Tumor Progression in an in Vivo Orthotopic Rodent Liver Tumor Model","volume":"2020","author":"Worlikar","year":"2020","journal-title":"BME Front."},{"key":"ref_6","first-page":"e29620","article-title":"A Review of the Abscopal Effect in the Era of Immunotherapy","volume":"14","author":"Nabrinsky","year":"2022","journal-title":"Cureus"},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"234","DOI":"10.1259\/0007-1285-26-305-234","article-title":"Whole Body Irradiation\u2014Radiobiology or Medicine?","volume":"26","author":"Mole","year":"1953","journal-title":"Br. J. Radiol."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"270","DOI":"10.1038\/s41586-019-1170-y","article-title":"CD8+ T Cells Regulate Tumour Ferroptosis during Cancer Immunotherapy","volume":"569","author":"Wang","year":"2019","journal-title":"Nature"},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"110","DOI":"10.1186\/s13045-020-00946-7","article-title":"Ferroptosis, Necroptosis, and Pyroptosis in Anticancer Immunity","volume":"13","author":"Tang","year":"2020","journal-title":"J. Hematol. Oncol."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"e000200","DOI":"10.1136\/jitc-2019-000200","article-title":"Non-Thermal Histotripsy Tumor Ablation Promotes Abscopal Immune Responses That Enhance Cancer Immunotherapy","volume":"8","author":"Qu","year":"2020","journal-title":"J. Immunother. Cancer"},{"key":"ref_11","doi-asserted-by":"crossref","unstructured":"Pepple, A.L., Guy, J.L., McGinnis, R., Felsted, A.E., Song, B., Hubbard, R., Worlikar, T., Garavaglia, H., Dib, J., and Chao, H. (2023). Spatiotemporal Local and Abscopal Cell Death and Immune Responses to Histotripsy Focused Ultrasound Tumor Ablation. Front. Immunol., 14.","DOI":"10.3389\/fimmu.2023.1012799"},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"3001","DOI":"10.1109\/TUFFC.2021.3100267","article-title":"Liver Histotripsy Mediated Abscopal Effect\u2014Case Report","volume":"68","author":"Ziemlewicz","year":"2021","journal-title":"IEEE Trans. Ultrason. Ferroelectr. Freq. Control"},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"742","DOI":"10.1161\/CIRCULATIONAHA.109.889071","article-title":"Noninvasive Creation of an Atrial Septal Defect by Histotripsy in a Canine Model","volume":"121","author":"Xu","year":"2010","journal-title":"Circulation"},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"2091","DOI":"10.1109\/TUFFC.2007.504","article-title":"High Speed Imaging of Bubble Clouds Generated in Pulsed Ultrasound Cavitational Therapy\u2014Histotripsy","volume":"54","author":"Xu","year":"2007","journal-title":"IEEE Trans. Ultrason. Ferroelectr. Freq. Control"},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"102000","DOI":"10.1016\/j.gassur.2025.102000","article-title":"The First International Experience with Histotripsy: A Safety Analysis of 230 Cases","volume":"29","author":"Wehrle","year":"2025","journal-title":"J. Gastrointest. Surg."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"1056","DOI":"10.1016\/j.ultrasmedbio.2018.10.035","article-title":"For Whom the Bubble Grows: Physical Principles of Bubble Nucleation and Dynamics in Histotripsy Ultrasound Therapy","volume":"45","author":"Bader","year":"2019","journal-title":"Ultrasound Med. Biol."},{"key":"ref_17","doi-asserted-by":"crossref","unstructured":"Tang, S., McGinnis, R., Cao, Z., Baker, J.R., Xu, Z., and Wang, S. (2025). Ultrasound-Guided Histotripsy Triggers the Release of Tumor-Associated Antigens from Breast Cancers. Cancers, 17.","DOI":"10.3390\/cancers17020183"},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"909","DOI":"10.1016\/j.ultrasmedbio.2025.02.005","article-title":"Acoustic Cavitation Emissions Predict Near-Complete\/Complete Histotripsy Treatment in Soft Tissues","volume":"51","author":"Haskell","year":"2025","journal-title":"Ultrasound Med. Biol."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"148","DOI":"10.1007\/BF00300234","article-title":"Determination of Subcutaneous Tumor Size in Athymic (Nude) Mice","volume":"24","author":"Tomayko","year":"1989","journal-title":"Cancer Chemother. Pharmacol."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"229","DOI":"10.1002\/jso.2930310402","article-title":"Tumor Measurement in the Nude Mouse","volume":"31","author":"Euhus","year":"1986","journal-title":"J. Surg. Oncol."},{"key":"ref_21","unstructured":"Kassambara, A., Kosinski, M., Biecek, P., and Fabian, S. (2025, August 17). Survminer: Drawing Survival Curves Using \u201cGgplot2\u201d. Available online: https:\/\/cran.r-project.org\/web\/packages\/survminer\/survminer.pdf."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"5089","DOI":"10.1158\/0008-5472.CAN-20-0342","article-title":"Generalized Additive Mixed Modeling of Longitudinal Tumor Growth Reduces Bias and Improves Decision Making in Translational Oncology","volume":"80","author":"Forrest","year":"2020","journal-title":"Cancer Res."},{"key":"ref_23","doi-asserted-by":"crossref","unstructured":"Wood, S.N. (2017). Generalized Additive Models: An Introduction with R, Chapman and Hall\/CRC. [2nd ed.].","DOI":"10.1201\/9781315370279"},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"71","DOI":"10.1038\/nrc1256","article-title":"Tumour-Educated Macrophages Promote Tumour Progression and Metastasis","volume":"4","author":"Pollard","year":"2004","journal-title":"Nat. Rev. Cancer"},{"key":"ref_25","first-page":"5","article-title":"Tumor-Associated Macrophages in Neoplastic Progression: A Paradigm for the in Vivo Function of Chemokines","volume":"71","author":"Mantovani","year":"1994","journal-title":"Lab. Investig."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"e155022","DOI":"10.1172\/jci.insight.155022","article-title":"CD206+ Tumor-Associated Macrophages Cross-Present Tumor Antigen and Drive Antitumor Immunity","volume":"7","author":"Modak","year":"2022","journal-title":"JCI Insight"},{"key":"ref_27","doi-asserted-by":"crossref","unstructured":"Fujimura, T., Kambayashi, Y., Fujisawa, Y., Hidaka, T., and Aiba, S. (2018). Tumor-Associated Macrophages: Therapeutic Targets for Skin Cancer. Front. Oncol., 8.","DOI":"10.3389\/fonc.2018.00003"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"145","DOI":"10.1186\/s13046-025-03377-9","article-title":"Tumor-Associated Macrophages Remodel the Suppressive Tumor Immune Microenvironment and Targeted Therapy for Immunotherapy","volume":"44","author":"Yang","year":"2025","journal-title":"J. Exp. Clin. Cancer Res."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"4411","DOI":"10.4049\/jimmunol.1101717","article-title":"Cutting Edge: Expression of XCR1 Defines Mouse Lymphoid-Tissue Resident and Migratory Dendritic Cells of the CD8\u03b1+ Type","volume":"187","author":"Crozat","year":"2011","journal-title":"J. Immunol."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"5516","DOI":"10.4049\/jimmunol.161.10.5516","article-title":"Eliciting T Cell Immunity Against Poorly Immunogenic Tumors by Immunization with Dendritic Cell-Tumor Fusion Vaccines1","volume":"161","author":"Wang","year":"1998","journal-title":"J. Immunol."},{"key":"ref_31","doi-asserted-by":"crossref","unstructured":"Zhang, Y., Lin, Z., Wan, Y., Cai, H., Deng, L., and Li, R. (2019). The Immunogenicity and Anti-Tumor Efficacy of a Rationally Designed Neoantigen Vaccine for B16F10 Mouse Melanoma. Front. Immunol., 10.","DOI":"10.3389\/fimmu.2019.02472"},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"881","DOI":"10.1038\/s41591-024-03418-4","article-title":"Personalized, Autologous Neoantigen-Specific T Cell Therapy in Metastatic Melanoma: A Phase 1 Trial","volume":"31","author":"Borgers","year":"2025","journal-title":"Nat. Med."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"1647","DOI":"10.7150\/thno.92089","article-title":"Focused Ultrasound Ablation of Melanoma with Boiling Histotripsy Yields Abscopal Tumor Control and Antigen-Dependent Dendritic Cell Activation","volume":"14","author":"Thim","year":"2024","journal-title":"Theranostics"},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"477","DOI":"10.1097\/01.cji.0000436722.46675.4a","article-title":"Immunogenicity of Murine Solid Tumor Models as a Defining Feature of In Vivo Behavior and Response to Immunotherapy","volume":"36","author":"Lechner","year":"2013","journal-title":"J. Immunother."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"255","DOI":"10.1006\/jsre.2000.6075","article-title":"Comparison of Intradermal and Subcutaneous Injections in Lymphatic Mapping","volume":"96","author":"Kersey","year":"2001","journal-title":"J. Surg. Res."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"2284","DOI":"10.1007\/s00268-007-9278-x","article-title":"Minimally Invasive Ablative Therapies for Invasive Breast Carcinomas: An Overview of Current Literature","volume":"31","author":"Mali","year":"2007","journal-title":"World J. Surg."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"1088","DOI":"10.1158\/1535-7163.MCT-24-0715","article-title":"Histotripsy-Focused Ultrasound Treatment Abrogates Tumor Hypoxia Responses and Stimulates Antitumor Immune Responses in Melanoma","volume":"24","author":"Song","year":"2025","journal-title":"Mol. Cancer Ther."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"350","DOI":"10.1007\/s13671-018-0234-9","article-title":"Immunology of Wound Healing","volume":"7","author":"Ellis","year":"2018","journal-title":"Curr. Derm. Rep."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"1566","DOI":"10.3892\/mmr.2014.2914","article-title":"Ki67 Is a Promising Molecular Target in the Diagnosis of Cancer (Review)","volume":"11","author":"Li","year":"2015","journal-title":"Mol. Med. Rep."},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"307","DOI":"10.1007\/s00262-004-0593-x","article-title":"Interaction of PD-L1 on Tumor Cells with PD-1 on Tumor-Specific T Cells as a Mechanism of Immune Evasion: Implications for Tumor Immunotherapy","volume":"54","author":"Blank","year":"2005","journal-title":"Cancer Immunol. Immunother."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"647","DOI":"10.1084\/jem.173.3.647","article-title":"Interferon Gamma and Tumor Necrosis Factor Have a Role in Tumor Regressions Mediated by Murine CD8+ Tumor-Infiltrating Lymphocytes","volume":"173","author":"Barth","year":"1991","journal-title":"J. Exp. Med."},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"31","DOI":"10.1034\/j.1600-065X.2003.00044.x","article-title":"Granzyme B: A Natural Born Killer","volume":"193","author":"Lord","year":"2003","journal-title":"Immunol. Rev."}],"container-title":["Cancers"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2072-6694\/17\/23\/3773\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,11,27]],"date-time":"2025-11-27T12:02:49Z","timestamp":1764244969000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2072-6694\/17\/23\/3773"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,11,26]]},"references-count":42,"journal-issue":{"issue":"23","published-online":{"date-parts":[[2025,12]]}},"alternative-id":["cancers17233773"],"URL":"https:\/\/doi.org\/10.3390\/cancers17233773","relation":{},"ISSN":["2072-6694"],"issn-type":[{"value":"2072-6694","type":"electronic"}],"subject":[],"published":{"date-parts":[[2025,11,26]]}}}