bio sphere
Microarrays for drug screening D
When the researchers exposed avid Sabatini, Brent Stockthese cells to 70 synthetic comwell, and Steve Bailey at the pounds, 4 of the 980 combinaWhitehead Institute for Biomedtions of compound, siRNA, and ical Research have developed easycell type proved lethal. Well-plate to-use microarrays that can test experiments confirmed this effect the effects of small molecules on for one of those combinations. mammalian cells. The new techBailey says that the next goal nology uses microscope slides, is to decrease the diffusion of which greatly reduce the amount the chemicals so that the spots of sample needed, instead of mido not intermingle. But John crowell plates (Proc. Natl. Acad. Weinstein at the National CanSci. U.S.A. 2004, 101, 16,144– cer Institute says, “The issues 16,149). “The goal of this project of controlling diffusion and of was to develop a higher-throughHuman lung cancer cells were seeded onto an array containquantitation—other than in a put screening method that is com- ing the toxic compound PAO in the center spot. Cell nuclei are relative sense—may be limitapatible with mammalian cells,” stained blue and actin filaments green. Scale bar = 100 µm. tions of the technology fundasays Stockwell. (Adapted with permission. Copyright 2004 National Academy of mentally.” Weinstein also wonThe array is a microscope slide Sciences, U.S.A.) ders whether various compounds printed with ~200-µm-diam dots diffuse out of the polymer at different (PAO) or only DMSO. Cells grew on of poly-(D),(L)-lactide/glycolide. This rates, which would impede comparisons the control spots but not on the dots polymer slowly and consistently releases of their effects on a given cell type. releasing PAO. The response was dosea chemical load and partly dissolves in Kit Lam at the University of Califorand distance-dependent, and the array dimethyl sulfoxide (DMSO), the solvent nia, Davis, says that his group and anothslowly released PAO for at least 10 days. used in most chemical libraries. A comer group developed a cell-based microasAlso, the assay worked with several types mercial arrayer overprints each dot with say about 10 years ago by immobilizing ~20 nL of a chemical dissolved in DMSO. of cancer cells and toxic compounds in one-bead one-compound libraries in soft addition to PAO. A layer of cells is poured over the dried agar together with living cells (Mol. DiTo examine a nonlethal phenotype, slides, and after a given time, the reversity 1996, 2, 57–63). “The main difthe researchers printed rapamycin onto searchers use robotic immunofluoresference between Bailey’s library [and polymer dots and incubated the slides cence microscopy to observe the cells. ours] is that his is spatially addressable, with lung cancer cells. Rapamycin inhib“You could have a slide with 5000 chemicals on it just sitting in the freezer. its the signaling pathway that triggers the but ours is not and therefore requires decoding of the positive beads,” Lam says. phosphorylation of a ribosomal protein. When you’re ready to use it, you just “However, with our method, we can easStaining with a fluorescent antibody put it in a dish and grow cells on it,” ily generate and screen an array of hunshowed that the cells on the rapamycin Bailey says. The amount of compound dreds of thousands to millions of differdots had lost their phosphorylation signeeded per spot is only one-hundredth nal. “There’s no reason to think that you ent compound-beads in a short time.” to one-thousandth of that required for a The developers of the slide-based couldn’t [use these arrays to] study any well-based assay. “So, if you had a miarray hope to see their technology comsignaling event with an antibody-based croliter of some rare natural product, mercialized. “A biologist would be very stain,” Stockwell says. you would have enough to use it thouexcited about the possibility of buying a The researchers then used the array sands and thousands of times,” says microscope slide that carries all 1500 method to screen compounds with unBailey. drugs approved in the United States or known cellular effects. They transfected The researchers tested the array by 5000 biologically active compounds,” small interfering RNAs (siRNAs) into seeding human lung cancer cells on top Stockwell says. a lung cancer and HeLa cells to inactivate of dots impregnated with either the cy—Linda Sage certain oncogenes and tumor suppressors. totoxic compound phenylarsine oxide © 2005 AMERICAN CHEMICAL SOCIETY
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