SBS Symposium: Advanced Applications Across Academia, Government & Industry

The SBS Symposium on Biomolecular Screening, held through Oct 18 – 19, 2010 in Research Triangle Park, Durham, NC brought together leading scientists in academia, government and industry to discuss screening applications in drug discovery. Chris Austin opened the symposium with a keynote presentation on the screening applications at the NIH Chemical Genomics Center (NCGC) that focus on small molecule and siRNA screens. NCGC takes a collaborative approach to screening where they provide the screening library and automation and their collaborators provide the drug target and its biological hypothesis for disease involvement. The goal of the screening campaign is not typically to provide leads for optimization, but to find chemical probes for further target validation relative to the disease state.

Bob Hertzberg represented Pharma in the opening session and gave an interesting look at small molecule drug discovery at GSK over the last decade. This was done by providing an overview of key drug discovery initiatives over that period. It began with scaling up HTS infrastructure, following up with the industrialization of the HTS process and driving efficiency, then focusing on solving drug attrition when the leads coming out of the HTS campaigns failed, and ending with the current initiatives focusing on flexibility and return on investment. An interesting point was made concerning drug attrition where the practices of driving small molecule potency to nM – pM levels were found to develop larger, more non-polar molecules that failed in late stage ADME/Tox. Hits with adequate potency and low molecular weight (high ligand efficiency) and low clogP are now favored to progress.

Steven Frye, Director of the Center for Integrative Chemical Biology and Drug Discovery at UNC, Chapel Hill, rounded out the representation by discussing why academia should be involved in drug discovery. There are currently about 80 academic and not-for-profit centers for screening small molecules libraries. Some of these are fee-for-service, but it is likely that the long term sustainability of these centers will be reserved for those that function through collaborative efforts that produce chemical probes for further target validation, rather than those that generate leads for further progression in a drug discovery pipeline.

I believe academic and not-for-profit centers like NCGC provide hope for drug discovery going forward. Pharma is solving some of the problems of adverse pharmacokinetics/pharmacodynamics and toxicity by looking more at parameters like ligand efficiency, but efficacy remains a major source of drug attrition. This is inexorably linked to appropriate target validation. Generating tools like chemical probes for better target validation is just what the doctor ordered!

Robert Edwards, 2010 Nobel Laureate for Medicine

I still recall the wonder of the first "test-tube" baby being announced. Louise Brown was born in 1978. She was conceived by removing an oocyte from the ovaries of her mother, Lesley, then placing the egg in a cell culture dish and exposing it to sperm from father John. The fertilized egg rapidly grew into an embryo of eight cells and was returned for gestation to Lesley. After a normal full-term pregnancy, Louise was born to great fanfare. I recall the event so clearly because Louise and I share a birthday.

Today, more than thirty years later, almost four million people have been born due to In Vitro Fertilization (IVF) and Louise Brown has a child of her own – through natural conception. IVF is an established medical practice that brings joy to the 10% of adults that are infertile. The 2010 Nobel Prize was justly awarded to Robert Edwards for his developmental work in IVF.

Robert Edwards work on IVF started two decades before Louise. In that period, he made important discoveries about fertilization. He clarified how human eggs mature, how different hormones regulate their development, and when the eggs are susceptible to the fertilizing sperm. He also determined the conditions under which sperm is activated and has the capacity to fertilize the egg. But it was his collaboration with Patrick Steptoe, a pioneer in laparoscopy, which enabled Louise’s conception. A laparoscope is an in vivo microscope that can visualize small objects, like an oocyte in an ovary, and remove it to be placed in a “test-tube.” The two cemented their relationship following Louise’s birth by establishing the Bourn Hall Clinic in Cambridge, UK, the world's first centre for IVF therapy – a training ground for gynecologists and cell biologists interested in furthering the clinical use of IVF.

Next Generation BioTek Performers

On October 13th, seven time Tour de France winner and cancer survivor Lance Armstrong arrived in Vermont. Just prior to landing Lance "tweeted" and invited all to come on a public ride. BioTek responded and our Whitney Hanson (GMBC green shirt) of BioTek's R&D team and his 15 month old son made the trek to Waterbury to meet and "ride" with Lance. Lance was greeted to a glorious Vermont fall day and along with ~ 200 other Vermonters started the long climb up to the Sugarbush Ski Resort.

When Whitney is not focused on his microplate R&D efforts, he hits the roads. He is a road racing veteran in Vermont and this summer won a criterium and took top three on several other occasions. He also participated in the national class Green Mountain Stage Race and took 8th in sprint points. Joining him in escorting Lance was Peter Weith, a bit farther behind.

By Peter Weith, BioTek's Vice President of Marketing, Sales & Service

iPSCs Take a Huge Step Towards Clinical Application

Last week Harvard researchers proved that it is not only possible to create pluripotent stem cells from ordinary skin cells rather than using controversial human embryos, but without viral delivery of the critical genes required to induce the transformation. Rossi et al published the use of modified mRNA constructs to introduce the requisite gene products into common cell types to induce cellular reprogramming to a pluripotent state. The newly induced pluripotent cells have been coined RNA-induced pluripotent stem cells (RiPSCs). Furthermore, the work of Rossi and colleagues show increased efficiency of iPSC derivation by 3 to 4 orders of magnitude significantly reducing low reprogramming efficiency hurdles towards clinical application. Moreover, concerns associated with the possibility of tumor growth, as was seen with gene therapy techniques using viral vectors, will not be a factor as mRNA has no known association with genetic integration into host cells. The significance of this breakthrough was apparent as the Harvard Stem Cell Institute immediately adopted the technique as the method of choice for stem cell line development moving forward. Such advancement can be seen as instrumental to developing, safe and efficient manner, donor specific somatic cell derived iPSCs for use in areas such as basic research, disease modeling, regenerative medicine and eventually clinical applications.

This development couldn’t come at a more critical juncture as the U.S. Government is battling with two scientists who sued to stop critical research dollars from being distributed by the NIH to support stem cell research. Although a Federal Appeals judgment lifted the injunction barring funding, disruptions have already occurred in many research laboratories. The importance of the research being conducted in this area is nearly universally understood by the scientific community and prominent political figures in international communities. In fact, the U.S. Justice Department recently argued that “Congress never intended for the Dickey-Wicker rules” to prevent all of the various research being conducted with embryonic stem cells. The rules were created to insure that new cell lines were created from eligible donors and in an ethical manner, as championed by the President of the United States.

In support of the continued funding of stem cell research though federal grant funding, Senator Specter, (D-Penn.) introduced legislation on Sept. 13, S.3766-Stem Cell Research Advancement Act of 2010. The purpose: To amend the Public Health Service Act to provide for human stem cell research, including human embryonic stem cell research, and for other purposes. The act can be seen as a mechanism to proactively summons Congress to determine the fate of stem cell research funding on the Senate floor that will otherwise be determined in the courts. Let’s hope Congress is not so distracted in an election season to overlook the critical nature of stem cell research.

Live-cell Assay to Interrogate GPCRs

In a previous post we discussed the advantages associated with luminescent assays mentioning a recently developed biosensor utilized in a live-cell assay format. The ability to either stably or transiently transform the biosensor into a cell line allows investigation of both endogenous and over-expressed GPCR targets. More recently we have demonstrated the ability to miniaturize the assay to use as little as 1,000 cells in a 384-well format to monitor the response of the endogenous β-adrenergic receptor in HEK293 cells (Figure 1).

















The assay conditions were further optimize to determine the optimal reagent equilibration incubation period and both agonist and antagonist incubation time. The Synergy H4 (BioTek Instruments, INC.) was used to measure the luminescence signal generated to determine the EC50 and IC50 of several agonists and an antagonist (Figure 2). The data was presented at ELRIG and MIPTEC conferences earlier this month.















Further efforts demonstrate that the automation of the assay, using the MultiFlo and Precision instruments, will be of particular importance for secondary screening efforts that employ the use of serial dilution of compounds that registered as hits in a screening campaign necessary to construct dose-response curves and determine potency. Assay performance as well as the pharmacology of several compounds were investigated and compared to those determined when manual methods were employed. Two new application notes will be available describing this work on the BioTek website in October.