A Precious Jewel in Sweet Home Alabama

Alabama. the heart of dixie, the Beautiful State, and the state whose license plate reads "Stars Fell on Alabama" is also home to a star player in anticancer drug development. Situated near the University of Alabama, Birmingham, the nonprofit Southern Research Institute has an enviable track record in discovering and developing anticancer drugs. With five Food & Drug Administration-approved drugs already in use, one in FDA fast-track status, and four more in clinical trials, Southern, as the institute is sometimes referred to, is shooting for the moon.

Created in 1941, Southern's first big project was working out a method for homogenizing peanut butter for the National Peanut Council. The institute also helped create the mechanism for sleeper sofas and invented a cigarette-smoking machine for use in nicotine and tar inhalation studies. Today, the institute includes labs that focus on automotive and aerospace engineering and environmental technologies. But its greatest success has been in developing anticancer drugs.

"Our record is significantly better than average, and we're proud of it," says John A. Secrist III, vice president of Southern's Drug Discovery Division. In fact, the institute can be credited with creating the first protocol for combination chemotherapy--that is, treatment using more than one anticancer drug.

"Back in the early days of cancer research, our scientists were involved in the initial evaluations of combination chemotherapy in mouse models," Secrist says. "We established the principle that combination chemotherapy is superior to single-agent chemotherapy in mouse models."

With funding from the National Institutes of Health's National Cancer Institute as well as from contracts with various pharmaceutical companies, current research focuses on creating better anticancer agents, as well as drugs to treat malaria, tuberculosis, herpes, and SARS (severe acute respiratory syndrome). A high-throughput platform and a diverse chemical compound library are what make this work possible. "Our database contains a lot of materials that were known pharmacophores," says Robert C. Reynolds, Southern's manager of medicinal chemistry and director of drug development activities. "We're finding that a lot of them have activity in a wide variety of targets, and the goal is to make them selective for particular targets."

Southern's success can also be attributed to numerous collaborations with researchers in academia and in industry, including the biopharmaceutical company Gilead Sciences; Cornell University; Pennsylvania State University College of Medicine, Hershey; and the University of Alabama, Birmingham. "We're starting to reach out, primarily to the academic community because a lot of them don't have access to a high-throughput platform," Reynolds says. "We're making deals with those universities where we acquire their biological targets, put them on our platform, help adapt them to high-throughput assays, and apply what chemical diversity we have to them."

According to Arthur D. Broom, a professor of medicinal chemistry at the University of Utah, Salt Lake City, Southern is successful because of its people. "Over the years, they have demonstrated a remarkable ability to attract world-class scientists," he says. "I think it is safe to say that Southern Research Institute is the leading private research institute in the country--probably in the world--in the field of drug discovery and development, and that that success is attributable in large part to the enlightened leadership and focus shared by the late John A. Montgomery [a key player in the development of the first four approved anticancer drugs from Southern] and Jack Secrist."

ALTHOUGH MUCH of the success can be ascribed to the focus on compound classes with special activity and on the teams of chemists and biologists who work so well together, "we've been lucky," Secrist quips. "Never underestimate luck."

Among Southern's achievements, born from hard work and maybe a bit of luck, are five FDA-approved anticancer drugs: amifostine, carmustine, dacarbazine, fludarabine, and lomustine. "As far as we know, no other organization out there has discovered or developed that many cancer drugs on its own," says Rhonda S. Jung, Southern's director of marketing communications and public relations. The institute has also evaluated 80% of all available cancer drugs at some point in their development. "If someone is getting a cancer drug out there, odds are at some point that drug was tested at Southern Research Institute," Jung says.

So just how does the institute develop and test all of these drugs? It all starts with the chemical repository. Southern stores 10,000 different compounds. The trick is finding compounds that have selective biological activity. "In the last five years, we've been focusing on marrying chemical diversity and biological diversity in various ways to continue to move forward in drug discovery," Reynolds says. They do this by choosing a subset of their chemical compounds and screening that against a biological target, such as an enzyme. The compounds are carefully selected from the library to minimize the time and number of chemicals used. For example, only compounds that will potentially fit the active site of the biological target--as can be gleaned from crystal structures--are tested.

So-called Lipinski rules are also applied. The Lipinski rules specify a set of five criteria--including hydrophobicity and molecular weight--that a compound should meet before it is investigated further as a potential drug component. Biologically active compounds that meet the Lipinski criteria have an increased probability of having the other properties desired of a drug.

The selected compounds are screened for biological activity by subjecting them to high-throughput assays. The targets are placed on a platform and, as in an assembly line, they are moved through as various chemicals are added.

"Each platform holds 180 [96-well] plates, and each well contains a chemical," says Thomas M. Fletcher III, Southern's manager for high-throughput screening. "The robot moves the plate to a liquid handler, which adds a buffer and then moves the plate to another liquid handler, which adds the enzyme," he says. "We can test 80,000 different chemicals in two to four hours this way."

The setup contains a multiple Beckman Coulter robotics platform and a PerkinElmer liquid-handling platform. These platforms automate initial plate setup for compound testing and perform all other assay requirements including microplate lid removal, plate shaking and sealing, and supply of disposable tips. After the assay, plate readers give results for fluorescence, luminescence, a variety of different radioactivities, or absorbance.

In this way, Southern has screened more than 20 million different compounds in the past three years. These compounds are derived from their own library and from those of other companies. Only a small fraction of the compounds screened have biological activity. Southern has tested 10,000 potential anticancer compounds in cell culture in the past 40 years, and of these, only a fraction have been successful enough to move forward to in vivo testing in immune-deficient mice.

THE INSTITUTE houses around 6,000 mice infected with 100 different human tumor lines. "Our ultimate goal is to cure the mice of the disease," says William R. Waud, director of cancer therapeutics and immunology. Once a drug passes in vivo testing, it goes to preclinical toxicology trials, in which the drug is tested in a rodent and a nonrodent model, usually a rat and a beagle dog--an FDA requirement. Once it passes these preclinical trials, the drug moves to human clinical trials. Because of limited funding, Southern licenses its drugs to companies prior to the preclinical toxicology trials.

In the past, these drugs were often alkylating agents, compounds that prevent DNA replication. More recently, Southern has focused on nucleosides, compounds that act by interfering with DNA synthesis. Nucleosides attack cancer cells by targeting one or more enzymes and can, theoretically, target multiple sites along biosynthetic pathways leading to DNA synthesis. The downside is that they also can inhibit these same enzymes in normal cells, resulting in toxic effects.

"Using a nucleoside is not as glamorous as picking some new enzyme, blocking it, and showing it has effects," Secrist says. "It's not as focused an attack, but it gives you multiple places where mechanistic influence can occur, and you get selectivity through the right combination of effects on various biological targets. So it's a very useful way to develop anticancer drugs."

A new nucleoside drug candidate discovered by Southern and being developed further by two companies may be more precise in its function than previous nucleoside drugs. Currently under evaluation by FDA, the drug, called clofarabine, would be used to treat leukemias. According to William B. Parker, a biochemist at Southern, "The clinical results have been good, and we are hoping for a positive answer from FDA by the end of the year."

Clofarabine is a nucleoside analog. When phosphorylated, natural nucleosides become nucleotides, which are the building blocks of DNA. Similarly, when clofarabine is phosphorylated, it is converted to clofarabine triphosphate, which is an analog of deoxyadenosine triphosphate (dATP), a building block for DNA synthesis. During DNA chain elongation, DNA polymerases will incorporate clofarabine triphosphate into the DNA just as they would dATP. Because clofarabine triphosphate is structurally different from dATP, once it is incorporated into the DNA, it slows the ability of the polymerases to add more nucleotides. In this way, clofarabine triphosphate effectively interferes with subsequent addition of nucleotides in cancerous cells.

Clofarabine triphosphate also inhibits ribonucleotide reductase, an enzyme involved in the metabolism of nucleotides. The combination of these two actions contributes to clofarabine's good antitumor activity.

Southern Research Institute's involvement in cancer therapeutics gives it a noble cause. "It's really an extremely rewarding job," Secrist says. "It's great to go home and tell my wife that we helped someone."