Research

Identification of secondary targets of N-containing bisphosphonates in mammalian cells via parallel competition analysis of the barcoded yeast deletion collection

Nicoletta Bivi^1,2 , Milena Romanello¹ , Richard Harrison³ , Ian Clarke² , David C Hoyle² , Luigi Moro⁴ , Fulvia Ortolani⁵ , Antonella Bonetti⁵ , Franco Quadrifoglio¹ , Gianluca Tell^1* and Daniela Delneri^2*

¹  Department of Biomedical Sciences and Technologies, University of Udine, Piazzale Kolbe, 33100, Udine, Italy

²  Faculty of Life Science, University of Manchester, Oxford Road, M13 9PT, Manchester, UK

³  School of Biological Sciences, Institute of Evolutionary Biology, King's Buildings, West Mains Road, Edinburgh EH9 3JT, UK

⁴  The Center for the Study of Metabolic Bone Diseases, via Vittorio Veneto, 34170, Gorizia, Italy

⁵  Department of Medical and Morphological Research, University of Udine, Piazzale Kolbe, 33100, Udine, Italy

author email corresponding author email* Contributed equally

Genome Biology 2009, 10:R93doi:10.1186/gb-2009-10-9-r93

Published:	10 September 2009

Subject areas: Cancer, Drug discovery, Medicine

Abstract

Background

Nitrogen-containing bisphosphonates are the elected drugs for the treatment of diseases in which excessive bone resorption occurs, for example, osteoporosis and cancer-induced bone diseases. The only known target of nitrogen-containing bisphosphonates is farnesyl pyrophosphate synthase, which ensures prenylation of prosurvival proteins, such as Ras. However, it is likely that the action of nitrogen-containing bisphosphonates involves additional unknown mechanisms. To identify novel targets of nitrogen-containing bisphosphonates, we used a genome-wide high-throughput screening in which 5,936 Saccharomyces cerevisiae heterozygote barcoded mutants were grown competitively in the presence of sub-lethal doses of three nitrogen-containing bisphosphonates (risedronate, alendronate and ibandronate). Strains carrying deletions in genes encoding potential drug targets show a variation of the intensity of their corresponding barcodes on the hybridization array over the time.

Results

With this approach, we identified novel targets of nitrogen-containing bisphosphonates, such as tubulin cofactor B and ASK/DBF4 (Activator of S-phase kinase). The up-regulation of tubulin cofactor B may explain some previously unknown effects of nitrogen-containing bisphosphonates on microtubule dynamics and organization. As nitrogen-containing bisphosphonates induce extensive DNA damage, we also document the role of DBF4 as a key player in nitrogen-containing bisphosphonate-induced cytotoxicity, thus explaining the effects on the cell-cycle.

Conclusions

The dataset obtained from the yeast screen was validated in a mammalian system, allowing the discovery of new biological processes involved in the cellular response to nitrogen-containing bisphosphonates and opening up opportunities for development of new anticancer drugs.