PROJECT SUMMARY Misdiagnosis is prevalent in younger women with dense breast tissue receiving breast cancer screening, resulting in missed cancers, needless follow-up testing, anxiety, and medical costs. Compared to mammography, magnetic resonance imaging (MRI) detects more breast cancers but still suffers from high false positive rates due to the conventional contrast agents used, e.g., gadolinium (Gd)-chelates. Our long-term goal is to develop novel, safe contrast agents for early detection of breast cancer that reduce the false positives and false negatives of breast MRI. The poor performance of Gd-chelates results from their lack of targeting and constant MRI signal. By remaining active, Gd-chelates produce high background signal in normal tissues and highlight both benign and malignant tumors. To address our long-term goal, we have developed Nano-, Encapsulated Manganese Oxide (NEMO) particles that will provide superior replacements for Gd-chelates. Our preliminary data shows that NEMO particle specificity is achieved by adding peptide targeting to underglycosylated mucin-1, which is overexpressed exclusively on breast cancer cells. NEMO particles provide a unique pH-switchable signal that is only activated upon internalization in acidic tumor cell endosomes (pH 5). No MRI signal is produced at pH of the blood (pH 7.4) or tumor extracellular space (pH 6.5). Our in vivo studies demonstrate that NEMO particles are safely tolerated in mice and exhibit a stronger signal than Gd-chelates. Currently, no high throughput method exists for testing new MRI contrast agents that predicts in vivo performance. The goals of the current project are to develop an innovative portable apparatus to enable evaluation of the sensitivity, specificity, and safety profile of NEMO particles vs. Gd-chelates using MRI and optical imaging of 3D microfluidic tumor models. Our central hypothesis is that NEMO particles will elicit low toxicity, specifically label breast cancer cells, and yield higher MRI contrast compared to Gd-chelates in 3D microfluidic tumor models and in mice with breast cancer. Our hypothesis will be tested with two aims: 1) Evaluate NEMO particle vs. Gd-chelate MRI contrast in 3D microfluidic tumor models and mice. 2) Evaluate toxicity and distribution of NEMO particles in 3D microfluidic tumor models and mice. This project is innovative because NEMO particles uniquely respond to endosomal pH to generate contrast only inside breast cancer cells to provide a simple binary readout (benign “OFF”, malignancy “ON”). Previously developed pH-sensitive MRI contrast agents respond to the acidic extracellular space, which is similar in benign and malignant tumors. We will also create a novel apparatus to enable MRI of 3D microfluidic tumor models for the first time. The proposed research is significant because we will demonstrate that NEMO particles have superior specificity, signal strength, and safety compared to Gd-chelates. This R15 award will offer cutting- edge training to undergraduates in nanomaterials and medical imaging research at West Virginia University. Over 3 years, 6 undergraduates pursuing engineering or biomedical sciences degrees will be recruited, trained, and assessed. This work will lead to further preclinical development and clinical trials of NEMO particles.

PROJECT NARRATIVE The proposed research is relevant to public health because misdiagnosis is prevalent in younger women with dense breast tissue receiving breast cancer screening, which causes missed cancers, needless follow up testing, anxiety, and increased medical costs. We seek to be the first to characterize an innovative imaging agent by complementary MRI and optical imaging within a micro-device that mimics the environment of breast cancer. This new MRI contrast agent is expected to expedite, simplify, and enhance the accuracy of breast cancer diagnosis in the clinic to improve patient outcome and reduce wasted medical dollars.