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F18-PET Detection of Breast Cancer Micrometastasis by CXCR4 Antagonist
The chemokine receptor, CXCR4 has shown to be one of critical factors for breast cancer metastasis. Its ligand, stromal cell derived factor –1 (SDF-1) is expressed in common destinations of breast cancer metastasis, including lymph node, lung, liver, and bone marrow. CXCR4 expression is low in normal breast tissues and high in malignant tumors. We identified a synthetic 14-mer peptide that inhibits CXCR4/SDF-1 mediated invasion with high specificity. Positron Emission Tomography (PET) offers target-specific imaging with a great sensitivity. In oncology, PET is the only technology that can reliably diagnose tumor from post surgical changes or radiation necrosis. Our CXCR4 antagonist is proven to be an excellent imaging probe to detect CXCR4 proteins in tissue culture cells and paraffin-embedded patient samples. Since CXCR4 is required for breast cancer metastasis, it can be used to detect metastasis because all metastasized tumors should have high expression of CXCR4. Our objective is to use CXCR4 antagonist as an imaging probe to detect primary tumors with high metastatic potential and their metastasis by non-invasive F18-PET. We have developed an animal model for bioluminescence imaging. This model will allow us to validate PET imaging data. We will make CXCR4 antagonist with aminooxy acetyl (Aoa) group attached to the N-terminal. Then, we will generate 4-[18F]fluorobenzaldehyde ([18F]FB-CHO) by using a method published by Poethko et al. (2004). Aoa and [18F]FB-CHO will be mixed to generate F18-labeled CXCR4 antagonist. The reaction mixture will be purified by RP-HPLC to eliminate unreacted materials. The method developed by Poethko et al. is ideal to label very short (<5 mers) peptides with 18F. We are currently modifying this method to optimize labeling of slightly longer peptides with disulfide bond (10 - 14 mers). Once our technique is established, the same technique can be used to label other peptides designed to target different surface receptors. Following the both images, animals will be sacrificed and their organs will be harvested for immunohistochemistry and RT-PCR to validate that CXCR4 antagonist is a specific imaging probe to detect CXCR4-positive tumors in vivo. By combining specificity of the antagonists for CXCR4 and the superior signal-to-noise ratio of 18F-PET, we can detect micrometastasis in any part of the body non-invasively. The successful outcome of our study can be readily translated into the clinic and will benefit cancer patients tremendously through the early detection of breast cancer metastasis.
Breast cancer is the second leading cause of death for women in the United States. The original site of the cancer in the breast, the ¡°primary¡± cancer can be resected for cure. Breast cancer spreads in a predictable fashion, the first site of spread is to the lymph nodes adjacent to the breast, then lung, liver, and bone marrows. Cancer cells located in the lymph node are the sign that the cancer has potentially spread out of the primary site. For this reason the lymph glands are removed surgically at the time of resection of the primary in order to determine if the cancer has migrated out from the original site of the cancer in the breast. The reason why breast cancer first spreads to the adjacent lymph glands and lung is not known. In recent studies a receptor on the cell called CXCR4 has been shown to be increased in metastatic cancer, and not in non-metastatic cancer or normal tissue from breast. This receptor is of particular interest since it has been described as directing the migration and homing of white blood cells to the lymphatic system, and in particular the lymph nodes.
Our preliminary data on the detection of CXCR4 show that CXCR4 expression is low in normal breast tissues but a highly significant presence in those individuals with cancer in the lymph nodes. If a means of limiting the expression of CXCR4 could be found, it might be possible to limit the ability of the cancer cells to migrate to the lymph nodes and lung, the early stage in the metastatic spread of the cancer. We identified such drug that is protein-base. Our published data on the invasion assay of breast cancer cells show that our CXCR4 antagonist is potent inhibitor against CXCR4-mediated invasion with superior specificity over CXCR4 antibody in our tissue culture model system. Furthermore, our animal data indicate that CXCR4 antagonist can block metastasis completely. Our objective is to develop non-invasive, highly sensitive imaging method to detect micrometastasis of breast cancer that can be ultimately used in the clinic.
Our study may lead us to a clinical trial that can benefit breast cancer patients through early detection and prevention of cancer metastasis.