1. ¹Division of Hematology/Oncology, Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Children's Hospital Boston, Boston, MA, USA
2. ²Department of Cell Biology, Institute for Genome Sciences and Policy, Duke University, Durham, NC, USA
3. ³Department of Medical Oncology, Dana Farber Cancer Institute, Boston, MA, USA
4. ⁴MD Anderson Cancer Center, Houston, TX, USA
5. ⁵Division of Hematology/Oncology, Oregon Health and Science University Cancer Institute, Portland, OR, USA
6. ⁶Department of Genetics, Harvard Medical School, Boston, MA, USA

Correspondence: Associate Professor GQ Daley, The Daley Laboratory, Division of Hematology/Oncology, Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Children's Hospital Boston, Karp Family Research Building 7214, 300 Longwood Avenue, Boston, MA 02115, USA. E-mail: george.daley@childrens.harvard.edu; Assistant Professor J Zhu, Department of Cell Biology, Institute for Genome Sciences and Policy and Duke University, 2353A CIEMAS, 101 Science Drive, Durham, NC 27708, USA. E-mail: jun.zhu@duke.edu

⁷These two authors equally contributed to the study.

Received 31 May 2007; Revised 26 June 2007; Accepted 1 July 2007; Published online 6 August 2007.

Abstract

Resistance to molecularly targeted chemotherapy, and the development of novel agents that are active against resistant forms of target proteins create the need for a sensitive and quantitative assay to monitor drug-resistant mutations in patients to guide treatment and assess response. Here, we describe an application of the polymerase colony (polony) method to identify and quantify known point mutations in the BCR-ABL oncogene in patients with chronic myelogenous leukemia who evolve resistance to ABL kinase inhibitors. The assay can detect mutations with a sensitivity of 10^-4, quantify the burden of drug-resistant cells, and simultaneously monitor the dynamics of several coexisting mutations. As a proof of concept, we analysed blood samples from three patients undergoing therapy with ABL kinase inhibitors and found that the patients' response to therapy correlated with our molecular monitoring. We were also able to detect mutations emerging in patients long before clinical relapse. Therefore, the polony assay could be applied to a larger patient sample to assess the utility of early mutation detection in patient-specific treatment decisions. Finally, this methodology could be a valuable research tool to shed light on the natural behavior of mutations pre-existing kinase inhibitors therapy and either disappearing over time or slowly taking over.