June 25, 2007

Seek & Destroy
By Dan Harvey
For The Record
Vol. 19 No. 13 P. 26

The strength of a superpowered molecule provides the muscle behind an effort to destroy cancer cells.

Researchers at the Washington University School of Medicine in St. Louis are harnessing a dreaded source for beneficial purposes—their investigations have generated a unique cancer-fighting strategy that utilizes a protein found in the human immunodeficiency virus (HIV).

This multidisciplinary effort brings together the expertise of cancer and HIV researchers, as well as radiologists. While the work has only recently begun, so far researchers have combined a peptide derived from the HIV-1 TAT (transactivator of transcription) protein—or simply TAT—and a proapoptotic Bcl-2 family member—termed Bim—to induce apoptosis (programmed cell death) in cancer tumors. The hope is that the TAT-Bim combination can be delivered to the tumor site using a positron emission tomography (PET) tracer.

In an article that appeared in the January issue of the Annals of Surgical Oncology, the researchers revealed how this TAT-Bim compound inhibits tumor growth in mouse models and impacts survival. They are now working to combine TAT-Bim with a PET radiotracer for the next stages of the research project.

In the projected larger picture, TAT will essentially perform a transportation function, carrying Bim into cancer cells. Once inside, Bim will then perform a cell-killing function, causing cancer cells to die via apoptosis. Eventually, a PET radiotracer will provide the function of locating the cancer cells.

Induced Apoptosis
In the article, the researchers report how the TAT-Bim compound induced apoptosis in cancer cells and enhanced the cell-killing effects of radiation. Specifically, the researchers set out to determine whether their TAT-Bim construct induced apoptosis in several cancer cell lines (T-cell lymphoma [EL4], pancreatic cancer [Panc-02], and melanoma [B16]) and whether TAT-Bim treatment synergized with radiation.

They reported that TAT-Bim resulted in apoptosis in a dose-dependent fashion in all cell lines. In addition, sublethal levels of irradiation augmented the effects of the TAT-Bim–induced apoptosis. Summarizing their results, the researchers note that after 40 days, 80% of the mice that received TAT-Bim were still alive compared with 20% of control mice that didn’t receive the treatment. The researchers concluded that TAT-Bim demonstrates a cancer therapy strategy involving apoptosis “by antagonizing the endogenous anti-apoptotic machinery.”

Further, they wrote that small peptide therapeutics, when combined with traditional adjuvant therapies such as radiation, “may provide a valuable ‘second hit’ and drive tumor cells into programmed cell death.”

Serendipitous Circumstances
The paper’s lead author, William G. Hawkins, MD, assistant professor of surgery at Washington University School of Medicine, says he embarked on this research direction almost by accident: He and his colleagues were investigating molecules that would compel cancers to die via apoptosis. “Cancer cells don’t respond to the natural signals that induce apoptosis, even though such signals abound,” he explains.

He was pointed toward TAT-Bim when he learned that Richard S. Hotchkiss, MD, professor of anesthesiology, medicine, and surgery and associate professor of molecular biology and pharmacology at Washington University, was looking for proteins that would hinder the apoptotic process in patients with potentially deadly infections. During the course of his own research, Hotchkiss had combined TAT with Bim, but he realized the compound had an effect counter to what he sought—it promoted apoptosis.

Given that knowledge, Hawkins wondered if TAT-Bim could prove an effective weapon against cancer. As a result, Hawkins and Hotchkiss’s labs entered into a research collaboration to investigate the cancer-killing potential of TAT-Bim. (Hotchkiss is a coauthor of the Annals of Surgical Oncology article.) “We wondered if the combined apoptotic and transportation processes would kill cancer cells from the inside,” says Hawkins.

As it turned out, TAT worked effectively to carry apoptotic-inducing molecules into cancer cells. “This gave the cells permission to die, so to speak,” says Hawkins.

Herculean Protein
TAT itself is a fascinating molecule. It doesn’t cause cells to die (that’s the job of Bim in this process); rather, it permeabolizes cells, enabling it to transport substances through cell membranes. Furthermore, TAT alone won’t cause HIV, lead to AIDS, or have any other adverse impact on health. “It’s just the binding protein. By itself, it has no infectious properties,” Hawkins says.

One of TAT’s most interesting—and, in this case, beneficial—characteristics is its remarkable strength. Small yet powerful, the TAT molecule can transport other relatively large molecules up to 1,000 times its own size. Hawkins indicates that researchers don’t understand how it accomplishes this. “TAT helps the HIV virus bind to and go inside cells, but we’re not really sure how and why it can drag an entire virus into a cell,” he says. “Not many molecules are capable of doing something like that.”

Providing an analogy, Hawkins remarks that TAT is so strong, it can drag the microscopic equivalent of a bus. “So, you can hook something as big as an antibody onto this molecule, and it will drag it across the cell membrane,” he adds. “It is a very unique little peptide.”

Enter Radiology
Its one limitation, as far as the researchers are concerned, is that it’s not selective. It will take itself inside all cells, not just cancer cells, and this presented a substantial consideration. “We would expect that, with this first-generation compound, there might be some toxicities, as it could affect normal cells and tissues,” explains Hawkins. “We wanted to do better.”

That is how Robert H. Mach, PhD, professor of radiology at Washington University School of Medicine, came into the story, explains Hawkins. Mach has been imaging cancer and working to develop novel tracers, used with PET, that selectively bind to cancer cells. Hawkins and his research colleagues will work to combine their molecules, which can enter and kill cancer cells, with tracer molecules that can seek out and target cancer. This will further augment the anticancer properties of TAT-Bim, as well as other proteins in development, as it will specifically target cancer cells and not impact healthy cells.

“By working with Dr. Mach, we are trying to take advantage of the research he has already done, in terms of developing a way to better target cancer,” says Hawkins. “He has been trying to find molecules up-regulated in cancers that he can create radiotracers for so that he can find cancers at much higher concentrations.”

“For the past 10 years, I’ve been working with Kenneth Wheeler, PhD, from the Wake Forest University School of Medicine, researching a protein to act as a biomarker of cell proliferation,” Mach says. “The function of this unique protein is not well understood, and we are among the few people looking at it.”

The receptor, he says, behaves much like Ki-67, a protein used in vitro as a gold standard for measuring cell proliferation. “With this receptor, we are able to get images that measure the proliferative status of a tumor,” says Mach. “We’ve published a series of papers that have validated it as a receptor-based biomarker for proliferation, which is a much different strategy from what other people are doing.”

“This could bring the whole process together,” says Hawkins. “We could use the radiotracers that bind to cancer cells in combination with virus particles that allow these things to go inside the cell, along with our peptides that allow cancer cells to complete the death process. In essence, we will be able to find the cancer, thanks to the PET radiotracer, and then get inside the cancer, thanks to the viral protein, and then we could kill the cancers, thanks to the work in our lab—much of which was pioneered by Dr. Hotchkiss, who was also instrumental in moving this collaboration forward.”

Looking Ahead
Hawkins indicates that his direction of research combined with Mach’s presents tantalizing potential. First, more cancers could be imaged more selectively. Second, proapoptotic peptides could be delivered directly into cancer cells.

Furthermore, Hawkins wants to combine apoptosis-inducing proteins such as TAT-Bim with cancer-fighting therapies, such as chemotherapy and radiation therapy, to increase the apoptotic signals in cancer cells. “We could use the same technologies to improve the effectiveness of chemotherapy and enhance novel therapies to make them more selective to cancer cells,” he says.

There are research implications, too, according to Hawkins. “We could use these to screen large numbers of compounds to see which ones—once inside a cell and selectively bound to cancer—will have less toxicity,” he explains. “So, there are a lot of potential uses, both from an imaging and cancer standpoint.”

Moreover, researchers could eventually use TAT in a manner more consistent with what Hotchkiss had originally envisioned—that is, the prevention of cell death, particularly in cases involving massive trauma, sepsis, and infections. “From the apoptotic standpoint, one could think of ways to prevent cells from dying. We don’t have to restrict ourselves to proapoptotic molecules; we can use antiapoptotic molecules,” says Hawkins.

Only the Beginning
Hawkins and Mach caution that the research in TAT-Bim is only getting started. The radiotracer element will truly come into play during the next steps in this ongoing work. At the same time, the research represents a promising approach to cancer therapy. Continued multidisciplinary research is expected to result in knowledge that could lead to new cancer-fighting strategies.

— Dan Harvey is a freelance writer based in Wilmington, Del.