Investigators at the Purdue University are making great strides in understanding intracellular mechanisms of delivery and release of drugs targeting neoplastic cells. The research, to be published in the upcoming issue of the Proceedings of the National Academy of Sciences, was conducted by the group of Dr. Philip Low, a professor of chemistry, and founder of Purdue spin-off Endocyte Inc.
Interest in how drugs are released after they enter their targeted cell led Low and his team to develop a color-coded method to visualize the cellular mechanisms. Jun Yang, a postdoctoral research associate in Low’s research group, together with Ji-Xin Cheng, an assistant professor in the Department of Biomedical Engineering, and his graduate student Hongtao Cheng, developed this method using a technique called fluorescence resonance energy transfer imaging.
“The drug turns from red to green when it is released inside the cell, clearly illuminating the process,” Yang said. “This is the first optical method to be developed to monitor this release. The main promise of this method is that it does not damage the cells being studied. Therefore, we are able to observe the process under true physiological conditions and watch it right as it is happening…”
In targeted drug therapy, drugs are linked to molecules that are used in excess by pathologic cells, for example a required nutrient, in order to transport drugs directly to the targeted cells while avoiding significant delivery of the toxic drug to normal cells. A commonly used agent, referred to as a ligand, is the vitamin folic acid. Cancer cells need folic acid to grow and divide and, therefore, have developed abundant receptors to capture it. These receptors are largely absent in normal cells. This means folic acid, and the drug linked to it, are attracted to the pathologic cells and are harmless to healthy cells, Low said.
Low led the team that discovered this folate-targeted treatment method in 1991 and the receptor-targeted technology is proprietary to Endocyte…
Yang examined receptor endocytosis, the process by which cells absorb materials — such as a drug attached to folic acid — that have been captured at special sites, called receptors, on the cell surface. The compound is then broken down and processed, releasing the drug.
One of the key mechanisms of this breakdown is disulfide reduction, which involves the breaking of chemical bonds. It was thought that disulfide reduction relied on the movement of the material along microtubules, hollow tubelike structures, and fusion with special digestive-enzyme containing compartments within the cell called lysosomes. However, the research showed that disulfide reduction occurred even when such components were removed from the process.
By inactivating different cellular components, Yang discovered which components are essential to the disulfide reduction process.
“It was surprising to learn that many other components of the cell, aside from those previously assumed to be responsible, were capable of releasing the drug from folic acid,” Yang said. “This significantly increases the opportunity for the drug to be released. For instance, we used to believe it had to get to a specific location to be released, and now we know it can happen almost anywhere during endocytosis.”
The mechanisms, locations and cellular components involved in the release of drugs within a cell had been under debate for several years, Low said.
“This is the definitive statement on how drugs are released within a cell,” he said. “We will use this knowledge to develop better receptor-targeted drug therapies to treat cancer and other diseases.”
Low and Yang worked with scientists from the Department of Chemistry, Weldon School of Biomedical Engineering and Endocyte, and used facilities at the Oncological Sciences Center, part of the Purdue Cancer Center, and Bindley Bioscience Center at Purdue’s Discovery Park.
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