Among them, acquired secondary KIT mutation is the most commonly observed etiology [5], [6]. Based on the results of two clinical trials, the current standard Temsirolimus supplier of care for IM-refractory GISTs is SU [7], [8]. However, genotype analysis showed that patients with secondary KIT mutation involving activation-loop domain have poor PFS and overall survival (OS) [7], [9]. In nowadays, SU remains the standard of care for IM-refractory GISTs regardless the status of their secondary KIT mutation. Clinically, some patients with secondary KIT mutation involving activation-loop domain experienced rapid disease after switch their treatment from IM to SU, as shown in Fig. 1. Figure 1 Rapid progression of IM-resistant tumor after SU treatment.

In the past few years, several commercially available TKIs, for example, nilotinib, dasatinib and sorafenib, are under clinical investigation for IM/SU-resistant GISTs. Nilotinib is designed based on the structure of IM and shows higher affinity to the ATP-binding site of ABL kinase to overcome IM-resistant chronic myeloid leukemia (CML) and also selectively inhibits KIT and PDGFR [10]. Dasatinib, an oral TKI for both BCR-ABL and Src family, is also a second-line drug for patients with IM-resistant CML and able to inhibit the activation of exon 11Val560Asp or exon 17Asp816Val KIT mutants [11]. Sorafenib is a multi-target inhibitor actively against BRAF, vascular endothelial growth factor receptor 2/3, PDGFR, and KITTrp557_Lys558del/Thr670Ile mutant expressed in Ba/F3 system and also has the activity to suppress the growth of GIST with KIT exon 11 fragment deletion in xenograft mouse model [12]�C[14].

Besides, recent data suggested that all three agents exhibit varied degree of activity against IM/SU-resistant GISTs with a disease-control rate (DCR, complete or partial response plus stable disease for 6 months or more) ranging from 25 to 70%. Unfortunately, genotype study has rarely been reported in these clinical studies. Whether individual KIT TKI preferntially against certain genotype of IM/SU-resistant GISTs remain largely unknown. This issue is clinically important because it may not only be useful in selecting optimal treatment for IM/SU-resistant GISTs but also help to identify potentially more active second-line treatment for IM-resistant GISTs with secondary activation-loop mutation, for which SU showed little activity.

In this study, we used Carfilzomib an in vitro cell-based drug screening platform, which consists of a series of COS-1 cells expressing KIT cDNA constructs encoding mutant exon 9Ala502_Tyr503insAlaTyr, 11Val555_Leu576del, 11Val560Asp, 13Val654Ala, 14Thr670Ile, 17Asp820Gly, and 17Asn822Lys either alone or in combination to mimicking the common KIT mutations observed in GISTs, to study the potential activity of several commercially available KIT TKIs at their achievable serum steady-state concentration.