ORIGINAL ARTICLE The low frequency of clinical resistance to PDGFR inhibitors in myeloid neoplasms with abnormalities of PDGFRA might be related to the limited repertoire of possible PDGFRA kinase domain mutations in vitro N von Bubnoff 1,4 , SP Gorantla 1,4 , RA Engh 2 , TM Oliveira 2 , S Tho¨ne 3 ,EA ˚ berg 2 , C Peschel 1 and J Duyster 1 1 III. Medizinische Klinik und Poliklinik, Klinikum rechts der Isar, Technische Universita¨t Mu¨nchen, Mu¨nchen, Germany; 2 The Norwegian Structural Biology Centre, Departments of Chemistry and Pharmacy, University of Troms, Troms, Norway and 3 Institute of Experimental Cancer Reserach, Comprehensive Cancer Center Ulm and University Hospital Ulm, Ulm, Germany Myeloproliferation with prominent eosinophilia is asso- ciated with rearrangements of PDGFR-A or -B. The most common rearrangement is FIP1L1-PDGFRA (FP). The majority of patients with PDGFR-rearranged myelopro- liferation respond to treatment with imatinib. In contrast to BCR–ABL-positive chronic myelogenous leukemia, only few cases of imatinib resistance and mutations of the FP kinase domain have been described so far. We hypothesized that the number of critical residues mediating imatinib resistance in FP in contrast to BCR–ABL might be limited. We performed an established systematic and comprehensive in vitro resistance screen to determine the pattern and frequency of possible TKI resistance mutations in FP. We identified 27 different FP kinase domain mutations including 25 novel variants, which attenuated response to imatinib, nilotinib or sorafenib. However, the majority of these exchanges did not confer complete inhibitor resis- tance. At clinically achievable drug concentrations, FP/ T674I predominated with imatinib, whereas with nilotinib and sorafenib, FP/D842V and the compound mutation T674I þ T874I became prevalent. Our results suggest that the PDGFR kinase domain contains a limited number of residues where exchanges critically interfere with binding of and inhibition by available PDGFR kinase inhibitors at achievable concentrations, which might explain the low frequency of imatinib resistance in this patient population. In addition, these findings would help to select the appropriate second-line drug in cases of imatinib-resistant disease and may be translated to other neoplasms driven by activated forms of PDGFR-A or -B. Oncogene (2011) 30, 933–943; doi:10.1038/onc.2010.476; published online 25 October 2010 Keywords: myeloid neoplasms; FIP1L1-PDGFRA; signaling therapies; drug resistance Introduction Idiopathic hypereosinophilic syndrome is characterized by sustained and unexplained eosinophilia exceeding 1.5  10 9 per liter (Chusid et al., 1975). A subset of patients with primary eosinophilia fall into the WHO (World Health Organization) category of myeloid neoplasms associated with eosinophilia and abnormal- ities of PDGFRA, PDGFRB or FGFR1 (Cross and Reiter, 2008; Gotlib and Cools, 2008; Tefferi and Vardiman, 2008). Myeloid neoplasms associated with rearrangements of PDGFRA, PDGFRB and FGFR1 are considered to be very rare entities (Gotlib and Cools, 2008). The most common molecular feature is FIP1L1-PDGFRA (FP), generated by a small inter- stitial deletion at chromosome 4q12 (Cools et al., 2003a). The resulting fusion gives rise to constitutive PDGFRA kinase activity and transforming potential (Cools et al., 2003a). The ABL, KIT and PDGFR small-molecule kinase inhibitor imatinib mesylate was reported to induce remissions in cases of idiopathic hypereosinophilic syndrome (Gleich et al., 2002). Later, molecular responses were ascribed to expression of FP (Cools et al., 2003a), but rapid and durable responses were also seen in myeloproliferation positive for numerous other rearrangements of PDGFR-A or -B (Cross and Reiter, 2008). In chronic myelogenous leukemia (CML) and Ph þ ALL, clinical resistance to imatinib is associated with the emergence of point mutations within the ABL kinase domain that obviate binding of the drug. The first exchange reported was BCR–ABL/T315I (Gorre et al., 2001), generating resistance to imatinib, nilotinib and dasatinib. In the meantime, more than 70 different BCR–ABL exchanges that confer clinical imatinib resistance have been identified in patients (Apperley, 2007). In contrast, only seven cases of FP-positive myeloproliferation with acquired imatinib resistance due to a point mutation in the PDGFRA kinase domain have been reported so far (Cools et al., 2003a; von Bubnoff et al., 2005a; Ohnishi et al., 2006; Gotlib and Cools, 2008; Simon et al., 2008; Lierman et al., 2009; Score et al., 2009). Strikingly, in six cases, either a FP/T674I (corresponding to T315I in cABL) or a Received 10 February 2010; revised 2 July 2010; accepted 8 September 2010; published online 25 October 2010 Correspondence: Dr N von Bubnoff, III. Medizinische Klinik und Poliklinik, Klinikum rechts der Isar, Technische Universita¨t Mu¨ nchen, Ismaningerstrae 22, Mu¨nchen, Bavaria 81675, Germany. E-mail: n.bubnoff@lrz.tum.de 4 These authors contributed equally to this work. Oncogene (2011) 30, 933–943 & 2011 Macmillan Publishers Limited All rights reserved 0950-9232/11 www.nature.com/onc