Histoplasty Modification of the Tumor Microenvironment in a Murine Preclinical Model of Breast Cancer

Purpose To develop a non-invasive therapeutic approach able to alter the biophysical organization and physiology of the extracellular matrix in breast cancer. Materials and methods In a 4T1 murine model of breast cancer, histoplasty treatment with a proprietary 700-kHz multielement therapy transducer using a coaxially aligned ultrasound imaging probe was used to target the center of an ex vivo tumor and deliver sub-ablative acoustic energy. Tumor collagen morphology was qualitatively evaluated pre- and post-histoplasty with second harmonic generation. Separately, mice bearing bilateral 4T1 tumors (n=4; 8 total tumors) were intravenously injected with liposomal doxorubicin. The right flank tumor was histoplasty treated, and tumors were fluorescently imaged to detect Doxil uptake following histoplasty treatment. Next, 4T1 tumor-bearing mice were randomized into 2 treatment groups (sham vs histoplasty, n=3/group). 48 hours following sham/histoplasty treatment, tumors were harvested and analyzed via flow cytometry. Results Histoplasty significantly increased (p=0.0023) liposomal doxorubicin diffusion into 4T1 tumors compared to untreated tumors (2.12 vs 1.66 fold increase over control). Flow cytometry on histoplasty treated tumors (n=3) demonstrated a significant increase in tumor macrophage frequency (42% of CD45 vs 33%, p<0.05) and a significant decrease in myeloid derived suppressive cell frequency (7.1% of CD45 vs 10.3%, p<0.05). Histoplasty treated tumors demonstrated increased CD8+ (5.1 % of CD45 vs 3.1%, p=0.117) and CD4+ (14.1% of CD45 vs 11.8%, p=0.075) T cell frequency. Conclusion Histoplasty is a non-ablative focused ultrasound approach to non-invasively modify the tumor ECM, increase chemotherapeutic uptake, and alter the tumor immune microenvironment.