PLoS One. 2013 Aug 27;8(8):e73607.

IMD-0354 targets breast cancer stem cells: a novel approach for an adjuvant to chemotherapy to prevent multidrug resistance in a murine model.

Azucena Gomez-Cabrero1*, Wolfgang Wrasidlo2, Ralph A. Reisfeld1

1 Department of Immunology and Microbial Sciences, The Scripps Research Institute, La Jolla, California, United States of America, 2 Moores Cancer Center, University of California San Diego, San Diego, California, United States of America

 

ABSTRACT

Although early detection of breast cancer improved in recent years, prognosis of patients with late stage breast cancer remains poor, mostly due to development of multidrug resistance (MDR) followed by tumor recurrence. Cancer stem cells (CSCs), with higher drug efflux capability and other stem cell-like properties, are concentrated in a side population (SP) of cells, which were proposed to be responsible for MDR and tumor repopulation that cause patients to succumb to breast cancer. Therefore, targeting of CSCs as an adjuvant to chemotherapy should be able to provide a more effective treatment of this disease. Here, we used IMD-0354, an inhibitor of NF-κB, identified for targeting CSCs, in a combination therapy with doxorubicin encapsulated in targeted nanoparticles. IMD-0354 did target CSCs, evidenced by a decrease in the SP, demonstrated by the inhibition of the following: dye/drug efflux, reduction in ABC transporters as well as in colony formation in soft agar and low attachment plates. Decrease of stem-like gene expression of Oct4, Nanog and Sox2, and apoptosis resistance related to the Survivin gene also was observed after treatment with this compound. In addition, IMD-0354 targeted non-CSCs as indicated by reducing viability and increasing apoptosis. Targeted drug delivery, achieved with a legumain inhibitor, proved to enhance drug delivery under hypoxia, a hallmark of the tumor microenvironment, but not under normoxia. Together, this allowed a safe, non-toxic delivery of both anticancer agents to the tumor microenvironment of mice bearing syngeneic metastatic breast cancer. Targeting both bulk tumor cells with a chemotherapeutic agent and CSCs with IMD-0354 should be able to reduce MDR. This could eventually result in decreasing tumor recurrences and/or improve the outcome of metastatic disease.

 

SUPPLEMENTARY

Breast cancer is the second most commonly diagnosed malignancy among women, and is second in cancer related deaths. Treatment of breast cancer with radio- and/or chemotherapy frequently leads to multiple drug resistance (MDR) and tumor recurrence, in addition to other side effects. For this reason, more efficient therapies are urgently needed.

We developed a novel delivery system that allows the encapsulation of chemotherapeutic drugs in lipid nanoparticles (Figure 1). This approach minimizes side effects from the free drug in circulation and in healthy organs. In addition, our lipid nanoparticles display an external ligand that allows them and their cargo (a chemotherapy drug) to specifically direct the treatment to the tumor and its microenvironment. In this way, we minimize liver toxicity observed for other non-targeted lipid nanoparticles, and maximize the therapeutic drug concentration in the tumor. We successfully applied such targeted nanoparticles to deliver two different drugs in an orthotopic syngeneic mouse model of breast cancer, showing efficacy and safety.

To address the tumor recurrence problem from which many cancer patients suffer, we focused on cancer stem cells. These are a small subpopulation of cells, found in most solid tumors, which present multi drug resistance and high potency. For this reason, cancer stem cells are believed to be a major factor responsible for tumor recurrence and a very attractive target for cancer treatment.

Our focus for this study was three fold: (1) to specifically target cancer stem cells, (2) to minimize toxicity from drugs, and (3) to simplify delivery of more than one compound by encapsulation in efficient safe targeting nanoparticles.

We found IMD-0354 to target cancer stem cells in a previous screening. This compound has anti-inflammatory properties and inhibits the NF-κB pathway. The commonly used in vivo administration for this drug is intraperitoneal injection, which is neither common nor convenient for treatment of breast cancer patients. Its loading into specifically targeted nanoparticles could easily solve such problems.

We also found that IMD-0354 was able to target, not only cancer stem cells, but also bulk tumor cells in vitro. This compound also increased the cytotoxic effect of a common chemotherapeutic drug such as doxorubicin (Dox), which we already successfully tested with our targeted nanoparticles.

More important, our strategy was able to significantly reduce tumor progression in a syngenic mouse model of aggressive breast cancer. We were also able to (1) use our targeting nanoparticles to deliver more than one drug in a safe and specific way to the tumor and surrounding tissues, and (2) target both cancer stem cells and bulk tumor cells at the same time.

Our findings represent several potential benefits in the biomedical field. The use of targeted nanoparticles reduces toxicity of the drugs to healthy organs, concentrates the therapeutic compounds in tumor cells and in the tumor microenvironment, and consolidates pharmacokinetics and pharmacodynamics of combination therapies. In addition, our approach allows for lower dosing of the chemotherapeutic agent since less compound is lost by kidney filtration, liver detoxification, decomposition in the blood stream and off-target organ retention. It also allows for a higher local concentration of the compound in the tumor microenvironment, which translates into a higher therapeutic efficiency.

Since this strategy showed excellent efficacy in a very aggressive model of breast cancer, it is likely that one could achieve this in other, more common and less aggressive, breast cancer cases? These results gave us the confidence and motivation to keep working with these novel targeted nanoparticles and expanding their applications.

Concurrent results showed a very similar efficacy in a xenograft model of human lung cancer, which we recently published in the Journal of Cancer Science and Therapy. It is indeed most encouraging that our strategy of targeting bulk and cancer stem cells with drugs encapsulated in tumor targeted nanoparticles showed excellent efficacy in aggressively growing tumors, in the presence of metastasis. The fact that this proved possible in two different types of cancer, with both human and murine cancer cells, and in both immunocompetent and immunocompromised models makes us most encouraged about the broad spectrum of applicability of this strategy to treat not only breast cancer, but also many other types of solid tumors.

Azucena Gomez-Cabrero pic1Figure 1. Structure of targeting nanoparticles.

 

IMG_2811b

Authors: Dr. Azucena Gomez-Cabrero (right), Prof. Ralph A. Reisfeld (center) and Dr. Wolfgang Wrasidlo (left).

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