Alginate-derived nanovolume cytochrome P450 microarrays for high-throughput inhibition assays

BIOT 26

Sumitra Meena Sukumaran, sukums@rpi.edu1, Moo-Yeal Lee, leem2@rpi.edu2, Douglas S Clark, clark@cchem.berkeley.edu3, and Jonathan S. Dordick, dordick@rpi.edu1. (1) Department of Chemical and Biological Engineering, Rensselaer Polytechnic Institute, 110 8th Street, Troy, NY 12180, (2) Solidus Biosciences Inc, Troy, NY 12180, (3) Department of Chemical Engineering, University of California at Berkeley, 497 Tan Hall, Berkeley, CA 94720
Cytochrome P450 inhibition represents a safety hurdle and a major expense in drug discovery. Previous in vitro assay technologies include well-plates and microfluidic platforms; however, these are not truly high-throughput, nanovolume assays.

We have developed a solid-phase, fluorescence-based microarray for high-throughput P450 inhibition assays. P450s immobilized in 10 nl alginate spots reproducibly exhibited reactivity comparable to that in solution, and increased storage stability and linear reaction time. We show that a single 1,080 or 2,856-spot array can simultaneously provide IC50 values for nine potential inhibitors with multiple P450s. IC50 values comparable to those in solution were obtained for P450 isoforms 3A4 and 2C9 with a number of compounds, including inhibitors like ketoconazole and sulphaphenazole. In addition, the P450 microarrays were interfaced to a wide-field, whole-slide imaging system to rapidly generate IC50 and Ki values. These advances represent steps toward an automated, high-throughput metabolism and toxicology assay directed towards personalized medicine.

 

Upstream Processing: Advances in Biocatalysis
8:00 AM-11:00 AM, Sunday, August 19, 2007 BCEC -- 107C, Oral

Division of Biochemical Technology

The 234th ACS National Meeting, Boston, MA, August 19-23, 2007