Drug Discovery

High throughput screening is the process where a large chemical library is screened for activity in an assay, typically using 384 or 1536-well plate technology. The Penn Center for Molecular Discovery (PCMD) is one of nine centers participating in the NIH Molecular Libraries Screening Centers Network (MLSCN) to screen the NIH repository for biological activity in assays submitted by scientists around the country.

Miniaturization technology allows scientists to screen thousands of different molecules using tiny wells in plastic plates. But the decreasing size of these wells reached a practical limit of a few microliters. The Diamond laboratory invented a method that doesn’t use wells at all. Molecules are printed like ink as individual nanoliter droplets on a glass microscope slide. An entire library of 100,000 compounds now fits in a slide holder the size of a deck of cards. Using piezo-aerosol delivery methods, Diamond can then add various enzymes, reactants, and fluorescent detectors to each nanoliter reaction. First published in PNAS in 2003, the scale-down saves costs on reagents and robotics needed to move multiple plates around; and, like a printing press, they can print as many copies as they need to use in different screening campaigns.

Diamond’s lab has already put the new HTS process to use. This year, Penn microbiologist Paul Bates identified a target, an enzyme called cathepsin L, required for the SARS (severe acute respiratory syndrome) virus to infect cells. Diamond, while independently screening for enzyme inhibitors, found one called MDL28170 that potently inhibited cathepsin L. Bates then tested the compound back in his biological assay with the SARS virus; sure enough, the molecule prevented the infection processes, giving science a new tool for the development of therapeutics against the SARS virus. (The findings were published in the early August 2005 issue of the PNAS.)

Publications

D. Gosalia, S.L. Diamond. Printing chemical libraries for nanoliter fluid phase reactions. Proc. Natl. Acad. Sci. USA (Track II) 100, 8721 (2003).

Horiuchi KY, Wang Y, Diamond SL, Ma H: Microarrays for the functional analysis of the chemical-kinase interactome. J Biomol Screen 2006, 11:48-56.

Simmons G, Gosalia DN, Rennekamp AJ, Reeves JD, Diamond SL, Bates P: Inhibitors of cathepsin L prevent severe acute respiratory syndrome coronavirus entry. Proc Natl Acad Sci U S A 2005, 102:11876-11881.

H. Ma, K. Y. Horiuchi, Y. Wang, S. A. Kucharewicz, S. L. Diamond. Nanoliter Homogeneous Ultra High Throughput Screening Microarray for Lead Discoveries and IC50 Profiling. Assay & Drug Develop.Tech.. 3:177 (2005).

Shah PP, Myers MC, Beavers MP, Purvis JE, Jing H, Grieser HJ, Sharlow ER, Napper AD, Huryn DM, Cooperman BS, Smith AB, Diamond SL. Kinetic Characterization and Molecular Docking of a Novel, Potent, and Selective Slow-binding Inhibitor of Human Cathepsin L.
Mol Pharmacol. 2008 Apr 10.

Myers MC, Shah PP, Diamond SL, Huryn DM, Smith AB 3rd. Identification and synthesis of a unique thiocarbazate cathepsin L inhibitor.
Bioorg Med Chem Lett. 2008 Jan 1;18(1):210-4. Epub 2007 Nov 1.

de Almeida RA, Burgess D, Shema R, Motlekar N, Napper AD, Diamond SL, Pavitt GD. A Saccharomyces cerevisiae cell-based quantitative beta-galactosidase assay compatible with robotic handling and high-throughput screening. Yeast. 2008 Jan;25(1):71-6.

Myers MC, Napper AD, Motlekar N, Shah PP, Chiu CH, Beavers MP, Diamond SL, Huryn DM, Smith AB 3rd. Identification and characterization of 3-substituted pyrazolyl esters as alternate substrates for cathepsin B: the confounding effects of DTT and cysteine in biological assays. Bioorg Med Chem Lett. 2007 Sep 1;17(17):4761-6. Epub 2007 Jul 5.

Wong EY, Diamond SL. Enzyme microarrays assembled by acoustic dispensing technology. Anal Biochem. 2008 Oct 1;381(1):101-6.

Beavers MP, Myers MC, Shah PP, Purvis JE, Diamond SL, Cooperman BS, Huryn DM, Smith AB 3rd. Molecular docking of cathepsin L inhibitors in the binding site of papain. J Chem Inf Model. 2008 Jul;48(7):1464-72.

Motlekar N, Diamond SL, Napper AD. Evaluation of an orthogonal pooling strategy for rapid high-throughput screening of proteases. Assay Drug Dev Technol. 2008 Jun;6(3):395-405.

Myers MC, Shah PP, Beavers MP, Napper AD, Diamond SL, Smith AB 3rd, Huryn DM. Design, synthesis, and evaluation of inhibitors of cathepsin L: Exploiting a unique thiocarbazate chemotype. Bioorg Med Chem Lett. 2008 Jun 15;18(12):3646-51.

Huang AH, Motlekar NA, Stein A, Diamond SL, Shore EM, Mauck RL.High-throughput screening for modulators of mesenchymal stem cell chondrogenesis. Ann Biomed Eng. 2008 Nov;36(11):1909-21. Epub 2008 Sep 13.

Wong EY, Diamond SL.Advancing Microarray Assembly with Acoustic Dispensing Technology. Anal Chem. 2008 Nov 26.

Sivendran S, Jones V, Sun D, Wang Y, Grzegorzewicz AE, Scherman MS, Napper AD, McCammon JA, Lee RE, Diamond SL, McNeil M. Identification of triazinoindol-benzimidazolones as nanomolar inhibitors of the Mycobacterium tuberculosis enzyme TDP-6-deoxy-d-xylo-4-hexopyranosid-4-ulose 3,5-epimerase (RmlC). Bioorg Med Chem. 2009 Nov 20.

Pagano N, Wong EY, Breiding T, Liu H, Wilbuer A, Bregman H, Shen Q, Diamond SL, and Meggers E. From Imide to Lactam Metallo-Pyridocarbazoles: Distinct Scaffolds for the Design of Selective Protein Kinase Inhibitors. J Org Chem. 2009 Dec 4;74(23):8997-9009.