Heat-Sensitive Liposome Aids Chemotherapy Delivery to Tumors

A tiny fat-based spherical structure called a liposome, modified at Duke to be sensitive to mild heating, can triple the amount of an anti-cancer drug delivered to tumors in mice compared to other liposome-based drug-delivery methods, Duke Medical Center researchers reported Thursday.

"Earlier this year we saw an improved therapeutic effect in animals, but didn't know exactly why," said Garheng Kong, lead author of the new study findings published in the Dec. 15 issue of Cancer Research. "Now we've shown that it's due to increasing the concentration of the drug in the tumor compared to the other treatments tested."

Because delivering anti-cancer drugs into tumors is one of the major hurdles in advancing potential drugs from the cell culture dish to clinical trials, this discovery could pave the way to improve the success of chemotherapy in humans, the researchers said.

"There are countless drugs that are very effective at killing cancer cells in the laboratory, but they act very differently in a living system - many just won't go where you want them to go," said principal investigator Mark Dewhirst, director of Duke Comprehensive Cancer Center's Hyperthermia Program. "We're able to see an enhanced anti-tumor effect in mice with this new liposome because we're getting more drug into the tumor. This new liposome may eventually be applicable in cases where traditional chemotherapy is not effective."

While human studies are more than a year away, the results of the scientists' studies in mice differentiate the therapeutic importance of heat, liposomes and drug release - and the interaction of these factors - by measuring amounts of the chemotherapy drug doxorubicin delivered to tumors by 10 different treatments. The study was funded by the National Institutes of Health, the Department of Defense, an NIH Medical Scientist Training Grant and Celsion Corp., Columbia, Md.

Drugs enclosed in standard liposomes, which aren't sensitive to temperature, are less able to escape blood vessels and enter normal tissues than free drugs. Thus, using standard liposomes to carry anti-cancer drugs generally reduces the toxic side effects of chemotherapy, but usually doesn't improve efficacy since standard liposomes don't deliver more drug to tumors.

So, in an effort to improve drug delivery to tumors, liposomes sensitive to temperatures above 42 degrees Celsius (108 degrees F) have been created. Heat-sensitive liposomes "melt" to varying degrees when heated, allowing the drug inside to escape.

"Only tumor blood vessels have gaps in them that allow the passive leaking of liposomes," explained Dewhirst, professor of radiation oncology. "With heat there are more gaps and therefore more liposomes enter the tumor. However, if we use a liposome sensitive to slight heating, not only do we get more liposome in the tumor, we also get more of the drug released. Essentially we increase the functional concentration of drug in the tumor."

Compared to other thermosensitive liposomes, the patented Duke liposome releases its cargo much faster - 50 percent in just 30 seconds - and at clinically obtainable temperatures of 39 to 40 degrees C (about 102 to 104 degrees F).

"This liposome is like a soccer ball with stitches - when heated, its stitches become leaky and the drug that is trapped inside will come out, and in this invention, will come out rapidly," explained the liposome's inventor, David Needham, professor of mechanical engineering and materials science at Duke's Pratt School of Engineering who co-authored the study. "After it goes through the heated area it seals up, and when it comes around again to the tumor it releases again."

This "low-temperature-sensitive" liposome and initial tumor growth experiments were reported in the March 1 issue of Cancer Research. Now, the scientists report they can increase drug delivery to the tumor and still protect normal tissues by using this liposome to carry the drug.

"Even if we heated normal tissue in addition to the tumor, it doesn't appear from our other work that the liposomes would come out of the blood vessels. Only tumor vessels were made 'leakier' by heating," said Dewhirst, referring to research the team published in the Aug. 15 issue of Cancer Research.

In the new study, the researchers compared mice whose implanted tumors were heated to either 34 degrees C (about 93 degrees F, which is skin temperature) or 42 degrees C, and who were injected with one of three liposome-encapsulated forms of doxorubicin or with free doxorubicin or no doxorubicin. The three liposomes were a standard non-thermosensitive liposome, a traditional temperature-sensitive liposome triggered at 42 to 45 degrees C and Duke's low-heat liposome.

At 34 degrees C, injecting free doxorubicin, a potent and clinically used chemotherapy drug, did not significantly slow tumor growth in mice compared to controls, the scientists reported. Only encapsulating the drug in the non-thermosensitive liposome gave a drug-related therapeutic effect at this temperature.

However, heating the tumor at 42 degrees C for an hour slowed tumor growth with or without doxorubicin, indicative of hyperthermia's own ability to kill cells. At 42 degrees C, all the liposome formulations were significantly better than the control or doxorubicin groups, but the Duke-engineered liposome was by far the most effective treatment, cutting the tumor growth rate in half and causing long-term complete regression of tumors in at least two-thirds of the mice, the researchers reported.

Through their analysis, the researchers concluded that adequate and rapid release of the encapsulated drug in the tumor or in tumor blood vessels is the most important factor for therapeutic success. The Duke liposome allowed drug concentration in the tumor to exceed what appears to be a threshold amount needed for effective therapy, they added.

"Total doxorubicin levels in the mouse tumors were increased with a non-thermosensitive liposome and with a traditional thermosensitive liposome plus heat, but it appears that the heat-triggered and rapid release of the drug by the low-temperature sensitive liposome is instrumental in achieving sufficient concentrations of drug to give a larger therapeutic effect," said Kong, an M.D./Ph.D. candidate in biomedical engineering at Duke.

The researchers add that while water baths were used to warm each mouse's tumor for these experiments, human patients receive hyperthermia treatment using specially designed microwave equipment. During the last 15 years, the Hyperthermia Program at Duke has worked to develop, characterize and improve clinical use of hyperthermia as a method to improve cancer treatment, said Dewhirst.

"The best part of these liposome results is that heating tumor areas to 40 to 43 degrees can be achieved today with modern hyperthermia equipment, without adverse side effects from heating," said Dewhirst.

Celsion Corp., which has licensed the rights for commercial development of the liposome from Duke, will decide when and where future human studies are done, assuming current and future lab work at Duke and elsewhere uphold the potential of the technique.

Other co-authors on the paper are Gopal Anyarambhatla, William Petros, Rod Braun and Dr. Michael Colvin, all of Duke.

Written by Joanna Downer.