The chemotherapeutic and chemopreventative effects of d-limonene, a monoterpene found in the essential oils of citrus fruits, spices, and herbs, have been studied extensively in spontaneous and chemically induced rodent tumors . and usually prove fatal . The development of secondary tumors is a sequential process, commonly referred to as a metastatic cascade, and failure to complete Disopyramide any one step prevents metastasis . During the metastatic cascade, primary tumor cells digest their surrounding extracellular matrix, migrate through interstitial spaces, and enter blood or lymphatic vessels where they are carried to distant organs. Once lodged in the target organs, these cells migrate into the interstitial spaces and continue to grow to develop a secondary tumor, or metastasis . Thus, the migration and invasion of cancer cells provides many potential targets for therapeutic intervention. Most anticancer drugs target the hyperproliferation of metastatic cells. While many of these drugs are efficacious in treating the beginning stages of cancer, none are curative for metastatic disease. Any delay in the diagnosis also renders many of these drugs ineffective . In addition, antiproliferation compounds cause many adverse side effects, including nausea, vomiting, suppressed immune system, and hair loss. It is, therefore, paramount that alternative therapies be developed to treat a greater scope of the disease with less disturbance to the wellbeing of the patient. Even though tumor cell migration is a hallmark feature of metastasis, since 1978, fewer than 100 compounds that have some capacity to inhibit tumor cell migration have been reported; however, thousands of compounds have yet to be tested. To address the need for a more comprehensive screen of possible antimigration compounds, we recently, developed an automated high-throughput cytotoxicity/cell migration assay suitable for screening large numbers of samples . During the development of our automated assay, we applied cell proliferation and migration assays to screen drugs, already known to have anticancer effects, for noncytotoxic antimigration properties. This report focuses on the effects of d-limonene and its derivatives. Disopyramide The chemotherapeutic and chemopreventative effects of d-limonene, a monoterpene found in the essential oils of citrus fruits, spices, and herbs, have been studied extensively in spontaneous and chemically induced rodent tumors . Limonene serves as a precursor to other oxygenated monocyclic monoterpenes such as carveol, carvone, menthol, perillyl alcohol (POH), and perillyl aldehyde . Due to success in tumor regression in various rodent cancer models, clinical testing of the cancer chemotherapeutic activity of these compounds is in progress, including Disopyramide Phase I clinical trials of POH in breast cancer patients [7, 8]. D-limonene and its derivatives disrupt isoprenylation of members of the Ras family of G proteins by geranylgeranyl transferases in some instances , though the effects of this disruption on downstream cellular behaviors are only now being elucidated. Thus much of the focus far has been on the inhibition of the cell cycle machinery and the induction of apoptosis . The effects of these compounds on migration, especially in breast cells, are unreported to date. We report Disopyramide here that POH inhibits migration of both malignant and nonmalignant human breast cells, and that migration inhibiting doses also prove cytotoxic in malignant cells. These results suggest that subtoxic doses of POH may PITX2 act as a preventative treatment for breast cancer. MATERIALS AND METHODS Materials MCF-10A and MDA-MB 435 cells were purchased from American Type Culture Collection. Fibronectin, epidermal growth.