<b>Screening Marches On</b><br/><br/>

Screening Marches On

by Mike May

Every thing moves faster these days, or so hopes nearly everyone in pharma or biotech. Much of that increase in speed comes from high-throughput screening (HTS), which combines automation, robotics, and other technologies to run many compounds quickly through a series of tests. In the therapeutic business, these tests often involve how a compound interacts with a disease target. Deborah Hartman, vice president, lead generation in the discovery enabling capabilities and sciences group at AstraZeneca Pharmaceuticals says, “We consider HTS to be a powerful engine for lead generation. It’s used to find novel compounds at promising targets.”

Roche scientist Ralph Garippa using automated cellular imaging in the search for new drug compounds effective against cancer.

From outside pharma and biotech, those processes finding more potential compounds and sites to attack with them don’t seem to be providing the expected returns. As Berta Strulovici, research vice president, head of automatic biotech at Merck Research Laboratories, says, “The media is always asking: ‘If industry is investing in this technology, why doesn’t pharma have more drugs?’” It seems like a fair enough question, especially as biotech and pharma tout not only HTS but sometimes even so-called ultra-HTS. Still, maybe it’s the wrong question.

As Strulovici explains, “HTS is only a process. You now have a much more efficient and cost-effective process at the very early stage of drug discovery, but the rest of the pipeline is still inefficient.” So HTS does make part of drug discovery run better and faster, but there are many other steps along the way.

Others in industry agree. Ralph Garippa, research leader, Roche discovery technologies and cell-based HTS/HCS, says, “It was an unrealistic expectation that everything would come tumbling down the pike right now, because all phases of the drug-discovery process were not revolutionized simultaneously and to the degree that HTS was.” Still, biotech and pharma companies continue to improve HTS and expand its applications.

Still-better screening

First, keep in mind that HTS itself remains an active field. Scientists and companies continue to develop better approaches to screening. Strulovici says, “What distinguishes us is that we have employed technologies to miniaturize in every type of assay so that we can screen our large collection versus every target in a time- and cost-effective manner.” Merck’s collection includes nearly two million compounds. She adds, “Most companies use only selected portions of their libraries versus targets, which does not give them the advantage of finding something outside the box.” To screen so many compounds against a target, Strulovici and her colleagues run all types of assays in 3,456- and 1,536-well plates. “Within a maximum of 10 days,” says Strulovici, “we can perform a screen versus the entire library. This is completely prohibitive if done in 96- or 384-well plates. It would take much longer, and the cost would be prohibitive.”

AstraZeneca uses acoustic nanoliter dispensing to make assay-ready plates.

Moreover, better screening does not necessarily depend on better technology. At AstraZeneca, for example, Hartman says the process improved because of better company organization. “We have fully integrated HTS in cross-disciplinary, lead-generation teams within research areas,” she says. “The teams work together though an internal network to share best practices and jointly assess new technology.”

On top of organizational changes, AstraZeneca is also employing new technologies, such as acoustic nanoliter dispensing. With this system, a scientist can order a set of compounds to test, and then robotic systems retrieve the compounds and make assay-ready plates. Combining this method of dispensing compounds with wells containing cryopreserved cells simplifies cell-based assays, says Hartman. “The end result is increased efficiency and a dramatic reduction of the amount of compound used.” She adds, “We often get better data quality because of the accuracy of the dispensing.”

Some companies also combine existing technologies in new ways. A joint venture of Applied Biosystems and MDS Sciex, for instance, produced the FlashQuant workstation, which uses MALDI ionization as the front end for triple-quadrupole mass spectrometry (MS). “The core feature is about speed,” says Tamara Bond, director of pharma business at Applied Biosystems. According to tests by Applied Biosystems and MDS Sciex, FlashQuant runs 25-fold faster for quantifying small molecules, as compared with the fastest commercial system that uses liquid chromatography as the front end of triple-quadrupole MS. Bond says, “This system makes sample introduction work like a microarray or plate reader, loading many spots at once.” She adds, “This was not possible before on triple quads for small molecules.”

A variety of issues prevented the use of MALDI as an MS front end for small molecules. “When looking at small molecules, you had lots of matrix interference,” says Bond. The MALDI matrix on the plates caused the noise. “You couldn’t see the compound,” she says, “and were not able to quantify it.” To solve that problem, scientists at Applied Biosystems and MDS Sciex identified a noninterferring matrix.

A scientist in a Merck laboratory sets up an assay.

“The FlashQuant is really focused at first on the ADME space,” says Bond. ADME absorption, distribution, metabolism, and excretion is used to profile compounds in drug discovery. It determines if a compound has the right characteristics to be a good drug. By doing this faster, FlashQuant can weed out bad compounds sooner.

Other companies also aim HTS at ADME, and other stages along the drug-development pipeline. Hartman says, “We use HTS technologies to profile large selections of compounds in ADME. It is also being applied to safety and toxicity assays.” She adds, “This helps to identify compounds with safety or toxicity liability, and to find it early. Then, these compounds can be removed or further developed to repair the problems. This pushes better leads into clinical trials.”

Beyond screening

Now that biotech and pharma can screen many compounds quickly, these companies want to speed up the steps that follow. In many cases, this means getting more from a screen than just a hit that says a compound interacted with a well in a plate. “We put in technology that can take initial hits,” says Strulovici, “and eliminate off-target effects and eliminate various artifacts, like assay-related artifacts.” Merck scientists also want more content behind every experiment. Strulovici says that adding technologies like microcalorimetry and cell microscopy can “add content to hits. So by the time we give data to therapeutic areas, they know which compounds have value to them.” Overall, she says, “HTS will push through the pipeline.”

That is already happening. At Roche, says Garippa, “We also use HTS for other strategies, such as derisking compounds before going to animals.” For example, Roche scientists might run an asthma or diabetes compound in a HERG-channel assay. If the compound blocks this channel, it can prevent a patient’s heart from working properly. “We use this kind of HTS to eliminate these kinds of toxicity issues even before scaling up for animals, and certainly before humans,” says Garippa.

In addition to getting better compounds, companies also want better targets, and HTS can help here, too. For example, scientists at Merck run full genome–scale siRNA screens to identify new targets. “We knock down genes, one by one, to find new targets and pathways,” says Strulovici. Garippa says that Roche’s scientists also use siRNA to screen for better targets. He says, “With siRNA, we can go in and knock down many of these new-drug targets, and see how it changes phenotypes of cells carrying those proteins.”

Spreading the success

Although some outside pharma might think that HTS fell below its expected promise, it’s too soon to tell. “Today, we see that a significant portion of AstraZeneca compounds in the pipeline are based on HTS starting points. It is bearing fruit today,” says Hartman.

HTS results also appear in other aspects of making new therapeutics. Garippa points out that quality control of the newest assays often originates in HTS. “We now have common standards for quality controls on biochemistry and cell-based assays that we didn’t have before,” he says. He also mentions that HTS improves steps in lead optimization. “The process is more sophisticated now,” he says. “Instead of just isolating a compound as an inhibitor or activator, we can see how it changes the kinetics of a process.”

Some of the HTS advances also come from spreading collaborations. For example, Merck works with Gideon Dreyfuss of the University of Pennsylvania School of Medicine. He identified a source of relevant cells originating from a patient with spinal muscular atrophy, and Merck scientists then ran screens all for free to identify compounds that interact with the given target within those cells. This will help Dreyfuss develop compounds to fight spinal muscular atrophy. Although Merck will be providing Dreyfuss with free screening results and lead compounds, Merck gets something too. “We learn,” says Strulovici. “Sometimes, we don’t have the knowledge to work on every target, but we could partner with academics who have this strength in their labs.”

In fact, high-throughput processes should be seen as a collaborative effort. In particular, HTS should be thought of as one soldier in an army against disease, not as a single white knight saving the drug business. As Strulovici says, “The time when people thought they could be isolated geniuses and could solve all of the world’s problems are gone.” The same goes for technology.