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Structural Insights into the ATP Binding Pocket of the Anaplastic Lymphoma Kinase by Site-Directed Mutagenesis, Inhibitor Binding Analysis, and Homology Modeling

Supporting Info

Rosalind H. Gunby,^† Shaheen Ahmed,^‡ Roberta Sottocornola,^† Marc Gasser,^‡ Sara Redaelli,^† Luca Mologni,^† Carmen J Tartari,^†^§ Valentina Belloni,^† Carlo Gambacorti-Passerini,*^† and Leonardo Scapozza^‡

Department of Clinical Medicine, Via Cadore 48, University of Milano-Bicocca, Monza, 20052, Italy, Pharmaceutical Biochemistry Group, School of Pharmaceutical Sciences, University of Geneva, University of Lausanne, Quai Ernest-Ansermet 30, CH-1211 Geneva 4, Switzerland, Department of Experimental Oncology, Istituto Nazionale dei Tumori, Via Venezian 1, Milan, 20133, Italy, and Department of Oncology, McGill University, Montreal, Canada

J. Med. Chem., 2006, 49 (19), pp 5759–5768

DOI: 10.1021/jm060380k

Publication Date (Web): August 23, 2006

†

University of Milano-Bicocca.

Authors contributed equally.

‡

University of Geneva.

Istituto Nazionale dei Tumori.

To whom correspondence should be addressed: Tel.: +39 0264488059. Fax: +39 0264488363. E-mail: carlo.gambacorti@unimib.it.

McGill University.

Abstract

Anaplastic lymphoma kinase (ALK) is a valid target for anticancer therapy; however, potent ALK inhibitors suitable for clinical use are lacking. Because the majority of described kinase inhibitors bind in the ATP pocket of the kinase domain, we have characterized this pocket in ALK using site-directed mutagenesis, inhibition studies, and molecular modeling. Mutation of the gatekeeper residue, a key structural determinant influencing inhibitor binding, rendered the fusion protein, NPM/ALK, sensitive to inhibition by SKI-606 in the nanomolar range, while PD173955 inhibited the NPM/ALK mutant at micromolar concentrations. In contrast, both wild type and mutant NPM/ALK were insensitive to imatinib. Computer modeling indicated that docking solutions obtained with a homology model representing the intermediate conformation of the ALK kinase domain reflected closely experimental data. The good agreement between experimental and virtual results indicate that the ALK molecular models described here are useful tools for the rational design of ALK selective inhibitors. In addition, 4-phenylamino-quinoline compounds may have potential as templates for ALK inhibitors.

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