robust activity of avapritinib, potent and highly ... · the dosing solutions of imatinib,...

Precision Medicine and Imaging

Robust Activity of Avapritinib, Potent andHighly Selective Inhibitor of Mutated KIT,in Patient-derived Xenograft Models ofGastrointestinal Stromal TumorsYemarshet K. Gebreyohannes1, Agnieszka Wozniak1, Madalina-Elena Zhai1,Jasmien Wellens1, Jasmien Cornillie1, Ulla Vanleeuw1, Erica Evans2,Alexandra K. Gardino2, Christoph Lengauer2, Maria Debiec-Rychter3,Raf Sciot4, and Patrick Sch€offski1

Abstract

Purpose: Gastrointestinal stromal tumors (GIST) arecommonly treated with tyrosine kinase inhibitors(TKI). The majority of patients with advanced GISTultimately become resistant to TKI due to acquisition ofsecondary KIT mutations, whereas primary resistance ismainly caused by PDGFRA p.D842V mutation. We testedthe activity of avapritinib, a potent and highly selectiveinhibitor of mutated KIT and PDGFRA, in three patient-derived xenograft (PDX) GIST models carrying differ-ent KIT mutations, with differential sensitivity to stan-dard TKI.

Experimental Design: NMRI nu/nu mice (n ¼ 93) weretransplanted with human GIST xenografts with KIT exon11þ17 (UZLX-GIST9KIT 11þ17), exon 11 (UZLX-GIST3KIT 11),or exon 9 (UZLX-GIST2BKIT 9) mutations, respectively.We compared avapritinib (10 and 30 mg/kg/once daily)versus vehicle, imatinib (50 mg/kg/bid) or regorafenib(30 mg/kg/once daily; UZLX-GIST9KIT 11þ17); avapritinib(10, 30, 100 mg/kg/once daily) versus vehicle or imatinib

[UZLX-GIST3KIT11]; and avapritinib (10, 30, 60 mg/kg/oncedaily) versus vehicle, imatinib (50, 100 mg/kg/twice daily),or sunitinib (40 mg/kg/once daily; UZLX-GIST2BKIT9).

Results: In all models, avapritinib resulted in reduction oftumor volume, significant inhibition of proliferation, andreduced KIT signaling. In two models, avapritinib led toremarkable histologic responses, increase in apoptosis, andinhibition of MAPK-phosphorylation. Avapritinib showedsuperior (UZLX-GIST9KIT 11þ17 and -GIST2BKIT 9) or equal(UZLX-GIST3KIT 11) antitumor activity to the standard doseof imatinib. In UZLX-GIST9KIT 11þ17, the antitumor effects ofavapritinib were significantly better than with imatinib orregorafenib.

Conclusions: Avapritinib has significant antitumoractivity in GIST PDX models characterized by differentKIT mutations and sensitivity to established TKI. Thesedata provide strong support for the ongoing clinical trialswith avapritinib in patients with GIST (NCT02508532,NCT03465722).

IntroductionGastrointestinal stromal tumors (GIST) are the most com-

mon soft tissue sarcomas of the gastrointestinal tract with anannual incidence of 10 to 15 cases per million people (1). Thediscovery that the vast majority of GISTs are driven by activat-ing mutations in KIT or platelet-derived growth factor receptoralpha (PDGFRA) has improved our understanding of themolecular pathogenesis of GIST and led to the successfuldevelopment of targeted therapies for this malignancy (2).Mutations in KIT or PDGFRA lead to a constitutive, ligand-independent activation of kinase activity and their downstreamsignaling cascades, resulting in increased tumor cell prolifera-tion and survival (2). Surgical resection is the only availablecurative treatment for primary, localized, and resectable GIST,yet 40% to 50% of patients will experience recurrent or met-astatic disease during follow-up (3). Furthermore, a subset ofpatients is not eligible for surgical treatment due to anatomiclimitations, general condition, or the presence of synchronous

1Laboratory of Experimental Oncology, Department of Oncology, KULeuven, and Department of General Medical Oncology, University Hospi-tals Leuven, Leuven Cancer Institute, Leuven, Belgium. 2Blueprint Med-icines Corporation, Cambridge, Massachusetts. 3Department of HumanGenetics, KU Leuven and University Hospitals Leuven, Leuven, Belgium.4Department of Pathology, KU Leuven and University Hospitals Leuven,Leuven, Belgium.

Note: Supplementary data for this article are available at Clinical CancerResearch Online (http://clincancerres.aacrjournals.org/).

Y.K. Gebreyohannes and A. Wozniak contributed equally to this article.

Prior presentation: Part of the results of this study have been presented at theAmerican Association for Cancer Research Annual Meeting 2017 (April 1–5, 2017Washington DC; abstract 2081).

Corresponding Author: Agnieszka Wozniak, KU Leuven, Herestraat 49, post815, Leuven 3000, Belgium. Phone: 321-634-1669; Fax: 321-634-6901; E-mail:[email protected]

doi: 10.1158/1078-0432.CCR-18-1858

�2018 American Association for Cancer Research.

ClinicalCancerResearch

www.aacrjournals.org 609

on December 7, 2020. © 2019 American Association for Cancer Research. clincancerres.aacrjournals.org Downloaded from

Published OnlineFirst October 1, 2018; DOI: 10.1158/1078-0432.CCR-18-1858

http://crossmark.crossref.org/dialog/?doi=10.1158/1078-0432.CCR-18-1858&domain=pdf&date_stamp=2018-12-26

http://clincancerres.aacrjournals.org/

metastatic disease (1). The dependence of GIST on mutatedreceptor tyrosine kinase led to the exploration of tyrosine kinaseinhibitors (TKI) for the systemic treatment of this rare but well-characterized malignancy. Imatinib, a small molecule with ATP-mimetic properties, has become the standard first-line treatmentfor patients with locally advanced, recurrent, inoperable, or met-astatic disease (4). Imatinib has tremendously improved thesurvival of patients with GIST with advanced disease and achievesdisease control in approximately 85% of the cases (5). Further-more, the drug was found to extend relapse-free survival andoverall survival when used in the adjuvant setting, after surgery inpatients with high risk of relapse (1). In most patients withmetastatic GIST, however, the duration of response to imatinibis limited. The occurrence of resistance, which ismainly caused bythe acquisition of secondary mutations, leads to progressionduring treatment with imatinib or related compounds. Othersmall-molecule TKIs, such as sunitinib and regorafenib, are usedin patients who are progressing on or are intolerant to imatinib(6). Despite their well-documented clinical activity in imatinib-refractory GIST and their broader activity against a variety ofmolecular targets, progression on these agents typically occursafter a median treatment duration of less than a year (6). To date,there are no established standard treatment alternatives forpatients with GIST after failure on all three approved lines of TKItreatment, but a number of compounds are currently being testedin early clinical trials in patients with refractory tumors. Never-theless, there is still an unmet medical need for novel treatmentapproaches, which should be tested first in preclinical settings.

Secondary mutation in KIT or PDGFRA is likely the mostimportant event leading to TKI resistance. These mutations canoccur in KIT exon 13 and 14, encoding the ATP-binding pocketof the receptor, or in exon 17 and 18, in the kinase activationloop. The latter stabilize the receptor in its active conforma-tion, and the majority of these mutations are known to causeresistance to both imatinib and sunitinib, which are widelyused as first- and second-line agents in the clinic (4). Althoughregorafenib is active against some of these mutated forms, atypical patient receiving this third-line treatment progressesafter a median period of only 4–5 months (7, 8). Apart from

the unsatisfactory efficacy of second- and third-line agents, theirbroader activity against multiple molecular targets leads to off-target toxicity, and many patients do not tolerate sunitinib andregorafenib as well as the first-line standard of care.

Avapritinib (BLU-285, Blueprint Medicines) is an oral, high-ly selective, and potent investigational inhibitor with activityagainst KIT exon 17 activation loop mutants, including KITp.D816V. This mutation is a known driver mutation in syste-mic mastocytosis (9). In vitro, avapritinib disrupts KIT sig-naling as assessed by inhibition of both KIT phosphorylationand activation of downstream proteins such as AKT and STAT3in human mast cell and leukemia cell lines (10). In vivo,avapritinib achieves dose-dependent tumor growth inhibitionin a mouse model of systemic mastocytosis (10). Moreover,avapritinib also inhibits PDGFRA p.D842V (11), the mutationresponsible for one out of five primary gastric GIST, forwhich there is no effective treatment available (12). Avapritinibis currently under investigation in patients with unresectable,treatment-resistant solid tumors including GIST (ClinicalTrials.gov: NCT02508532 and NCT03465722) and in advanced sys-temic mastocytosis (NCT02561988).

In this work, we assessed the preclinical activity of avapri-tinib in vivo, using three patient-derived xenograft (PDX) mod-els of GIST, characterized by diverse KIT mutations and varyingsensitivity to the available standard TKI therapies.

Materials and MethodsXenograft models

For the current project, we transplanted three PDX models,established and fully characterized in the Laboratory of Experi-mental Oncology, KU Leuven (Leuven, Belgium). A total of 93NMRI (nu/nu) mice (Janvier Laboratories) were bilaterallyengrafted with models UZLX-GIST9KIT 11þ17 (KIT: p.P577del;W557LfsX5;D820G), -GIST3KIT 11 (KIT: p.W557_V559delinsF),and -GIST2BKIT 9 (KIT: p.A502_Y503dup), which are known toretain morphologic and molecular features of the original tumorduring passaging. The model characteristics and experimental setup are presented in Table 1. Xenografting of human tumors fromconsenting patientswithGISTwas approved by theMedical EthicsCommittee of the University Hospitals Leuven and the animalexperiments using PDX were approved by the Ethics Committeefor Animal Research, KU Leuven, and performed according to itsguidelines and Belgian regulations.

Drugs, reagents, and experimental designThe dosing solutions of imatinib, sunitinib, and regorafenib

(all from Sequoia Research) were prepared as described earlier(13). Avapritinib, provided by Blueprint Medicines, was dis-solved in 0.5% carboxymethyl cellulose supplemented with 1%Tween 80. The resulting suspension was kept at 4�C protectedfrom light; a fresh suspension was prepared every 3 days.Chemical structures of all drugs used in the study are presentedin Supplementary Fig. S1, the structure of avapritinib has beenpreviously published by Evans and colleagues (11). Whentumor growth had reached a threshold of 500 mm3, mice weretreated with the oral compounds for 16 days by gavaging. Thedoses of 10 and 30 mg/kg avapritinib, tested in all threemodels, were chosen based on previous preclinical in vivo workin a KIT exon 17 mutant mastocytoma model demonstratingdose response activity that was well tolerated in vivo (11). The

Translational Relevance

Advanced gastrointestinal stromal tumors (GIST) are rou-tinely treated with tyrosine kinase inhibitors (TKI). However,over time, the vast majority of patients develop resistance toTKI, mainly due to the acquisition of a secondary mutation inthe activation loop of KIT. Both imatinib and sunitinib areineffective in treating GIST with such mutations. Althoughregorafenib is active against some of these activation loopmutants, in the clinic it achieves a median progression-freesurvival of only 4.5 months. Avapritinib, a novel, potent, andselective inhibitor of KIT and PDGFRA activation loop muta-tions, showed robust in vivo antitumor activity in patient-derived GIST xenografts. Our preclinical findings indicate thatavapritinib could be a relevant treatment for patients withGIST with primary or secondary resistance to approved TKIand support investigation in ongoing clinical trials in patientswith GIST.

Gebreyohannes et al.

Clin Cancer Res; 25(2) January 15, 2019 Clinical Cancer Research610




increased doses of avapritinib, used in models with primarymutations (i.e. UZLX-GIST3KIT 11 and UZLX-GIST2BKIT 9), werechosen based on the biochemical evaluation of avapritinib,which demonstrated IC50 of approximately 0.2–1 nmol/L onKIT exon 17 mutants, 1–2 nmol/L against KIT exon 11 mutants,and >50 nmol/L IC50 against KIT wild-type kinase domain (11)and suggested that potentially higher exposure of avapritinibmight be required for inhibition of GIST tumors driven byprimary KIT exon 9 and 11 mutants. The detailed informationabout treatment groups and doses are presented in Table 1.During the dosing period, tumor volume was measured threetimes per week using a digital caliper and the body weight andgeneral well-being of the animals was followed up daily. At theend of the experiment, mice were sacrificed 2 hours after the lastdose, and tumors were divided with one half snap frozen inliquid nitrogen and one half fixed in 4% buffered formaldehydefor further histopathologic and molecular assessments. Antitu-mor activity was assessed on the basis of the evolution of tumorvolume expressed as the percentage of the normalized baselinevalue. Furthermore, histopathology and Western blotting wereperformed. For each mouse, the bilateral tumors were countedas independent events.

Western blotting and IHC were conducted using the fol-lowing antibodies and reagents: KIT from Dako/Agilent; dis-covered on GIST 1 (DOG1) from Novocastra; phospho-KITY719(pKITY719), phospho-KITY703 (pKITY703), phospho-AKTS473(pAKTS473), AKT, a-tubulin, p42/44 MAPK, phospho-MAPK(pMAPK), 4-E binding protein 1 (4EBP1), phospho-4EBP1(p4EBP1), histone H3 (HH3), phospho-HH3 (pHH3), andcleaved-PARP all from Cell Signaling Technology; Ki67from Thermo Fisher Scientific; EnVisionþ System-HRP and30diaminobenzidine-tetrahydrochloride (DAB), both fromDako/Agilent. For Western blotting, the secondary antibodies,conjugated with horseradish peroxidase, were from Cell Sig-naling Technology and Western Lightning Plus-ECL fromPerkinElmer was used for band visualization.

HistopathologyHematoxylin and eosin (H&E) staining was performed to

evaluate the general tumor morphology, the histologicresponse (HR) to treatment, as well as to count mitotic andapoptotic cells. Stained tissue sections were analyzed using anOlympus CH-30M microscope (Olympus). Representative pic-tures were captured using the Olympus Color View digitalcamera and analyzed with Olympus Cell D imaging software.

The HR was graded by assessing the magnitude of necrosis,myxoid degeneration, and/or fibrosis using a previouslydescribed grading system: grade 1 (0%–10% of tumor area),grade 2 (>10% and �50%), grade 3 (>50% and �90%), andgrade 4 (>90%; refs. 14, 15). Moreover, IHC was performed forKIT and DOG1, Ki67, and pHH3 staining was used to assessproliferation, cleaved-PARP to quantify apoptosis and pMAPKto evaluate KIT pathway activity. Proliferation and apoptosiswere assessed by counting the number of mitotic and apoptoticcells on H&E–stained slides and the IHC analysis was alsobased on counting positive cells. Both evaluations were per-formed in 10 high-power fields (HPF) at 400-fold magnifica-tion. The Ki67-labeling index was calculated as an averagepercentage of Ki67-stained nuclei in 5 digital images taken at400-fold magnification. KIT pathway inhibition was evaluatedby grading the intensity of the pMAPK staining as well as thepercentage of tumor area showing positivity, as described inSupplementary Table S1.

Western blot analysisTo determine the effect of the different treatments on the

KIT signaling pathway, Western blotting was performedas described previously (16). Densitometry was done usingthe AIDA software (Raytest) to do semiquantitation of thephospho-protein levels (17).

Statistical analysisThe comparison between tumor volumes on day 1 versus day

16 was done using the Wilcoxon matched pair test (WMP).Comparisons between different treatment groups were doneusing the Mann–Whitney U test (MWU). A value of P < 0.05 wasdefined as statistically significant. STATISTICA version 13 (DellInc.) was used for all calculations.

ResultsTumor volume assessment

After the 16-day treatment period, vehicle-treated tumorsfrom all models showed a steady and statistically significantincrease in relative tumor volume (216% of the baselinevolume for UZLX-GIST9KIT 11þ17, 293% for -GIST3KIT 11, and172% for -GIST2BKIT 9; P < 0.05 for all, WMP; Fig. 1). Consis-tent with previous reports, no significant difference wasobserved in the UZLX-GIST9KIT 11þ17 model between the rel-ative tumor volume of the vehicle- and imatinib-treated tumors

Table 1. Detailed description of xenograft models and experimental set-up

Model name UZLX-GIST9KIT 11þ17 UZLX-GIST3KIT 11 UZLX-GIST2BKIT 9

Model characteristicsKIT mutation Exon 11: p.P577del;W557LfsX5; exon 17: D820G Exon 11: p.W557_V559delinsF Exon 9: p.A502_Y503dupSensitivity to imatinib in vivo Resistant Sensitive Dose-dependent sensitivity

Treatment groupsControl Vehiclea (n ¼ 7) Vehiclea (n ¼ 6) Vehiclea (n ¼ 4)Imatinib 50 mg/kg/bid (n ¼ 7) 50 mg/kg/bid (n ¼ 5) 50 mg/kg/bid (n ¼ 4)

n/a n/a 100 mg/kg/bid (n ¼ 4)Sunitinib n/a n/a 40 mg/kg/qd (n ¼ 4)Regorafenib 30 mg/kg/qd (n ¼ 7) n/a n/aAvapritinib 10 mg/kg/qd (n ¼ 6) 10 mg/kg/qd (n ¼ 6) 10 mg/kg/qd (n ¼ 5)

30 mg/kg/qd (n ¼ 6) 30 mg/kg/qd (n ¼ 6) 30 mg/kg/qd (n ¼ 5)n/a n/a 60 mg/kg/qd (n ¼ 5)n/a 100 mg/kg/qd (n ¼ 6) n/a

Abbreviations: bid, twice daily; n, number of mice; n/a, not applicable; qd, once daily.a0.5% carbomethyl cellulose with 1% Tween 80.

Activity of Avapritinib in GIST Xenografts

www.aacrjournals.org Clin Cancer Res; 25(2) January 15, 2019 611




(13), whereas regorafenib resulted in modest tumor regression(82% of baseline, P ¼ 0.02 WMP). As expected, treatment withimatinib led to a regression in tumor volume (to 32% ofbaseline) in UZLX-GIST3KIT 11, confirming the imatinib sensi-tivity described previously (18). In contrast, UZLX-GIST2BKIT 9

tumors treated with the standard dose of imatinib (50 mg/kg)grew significantly. In this model, doubling the dose of imatinibled to tumor stabilization owing to the dose-dependent sensi-tivity to imatinib previously observed in this model and in linewith the known behavior of KIT exon 9–mutated GIST in theclinic (1). Sunitinib caused significant tumor shrinkage in thismodel (Fig. 1).

In all three xenograft models, avapritinib (10 mg/kg) resultedin tumor volume stabilization compared with the baseline value.This effect was comparable to the effects induced by the higherdose of imatinib in UZLX-GIST2BKIT 9 (100 mg/kg) or to regor-afenib inUZLX-GIST9KIT 11þ17 (Fig. 1). Remarkably, at the dose of30 mg/kg, avapritinib treatment resulted in substantial tumorregression as compared with baseline in two of the testedmodels,to 27% in UZLX-GIST9KIT 11þ17 (P ¼ 0.005) and to 26% in-GIST3KIT 11 (P ¼ 0.008, both WMP), and tumor volume stabi-lization (90%) in UZLX-GIST2BKIT 9 (P ¼ 0.08, WMP). Similarly,in UZLX-GIST3KIT 11, higher dose of avapritinib (100 mg/kg), ledto a significant tumor regression to 26% of baseline value (P ¼0.005, WMP), which was similar to the effect of imatinib in thismodel (Fig. 1B). In addition, inUZLX-GIST2BKIT 9, avapritinib at adose of 60 mg/kg led to tumor shrinkage, which was signifi-cantly better than imatinib (at both doses) and comparablewith sunitinib (Fig. 1C). Taken together, avapritinib induced

remarkable and dose-dependent effects on tumor volume in allthree models.

During the course of this study, the treatment with avapritinibwas well tolerated, and mice had a stable body weight withinethically acceptable limits (Supplementary Fig. S2). We didobserve a yellowish skin discoloration in all mice treated with100mg/kg of avapritinib, although it did not have any impact onthe well-being of animals.

Histopathologic evaluationIn all three models, vehicle-treated tumors showed spindle cell

morphology and diffuse KIT and DOG1 immunopositivity (Sup-plementary Fig. S3). These characteristics resembled featuresobserved in the original patient samples used for xenografting,as well as those found in previous passages, proving stablemorphology of the models. In addition, KIT mutational analysisof ex-mouse tumor samples confirmed the presence of mutationsas seen in the patient biopsy.

HR was assessed on H&E–stained slides by evaluating themagnitude of the necrosis, fibrosis, and myxoid degeneration inthe tumor tissue, induced by different treatments (Fig. 2A). Onlyminimal (grade 1) HR was observed in UZLX-GIST9KIT 11þ17

tumors treated with imatinib. Regorafenib induced grade 2 HRin 36% of these tumorsmainly through the induction of necrosis.As expected, and in line with our prior observations, imatinibcaused grade 2 or higher HR in all treated UZLX-GIST3KIT 11

tumors, with 67% of the xenografts showing grade 3 HR. In theUZLX-GIST2BKIT 9 model, all tumors treated with imatinib (bothat standard and higher dose) showed grade 1 HR.

In two out of three models, avapritinib resulted in remark-able HR. In UZLX-GIST9KIT 11þ17, 30 mg/kg avapritinibinduced grade 2 and grade 3 HR in 60% of the tumors. InUZLX-GIST3KIT 11, 10 mg/kg avapritinib induced grade 2 HR inthe vast majority (80%) of the tumors. Moreover, the higherdoses (30 and 100 mg/kg) led to grade 3 and grade 4 HR in thismodel. This effect was slightly more pronounced thanin tumors treated with imatinib. Interestingly, in bothUZLX-GIST9KIT 11þ17 and -GIST3KIT 11, the HR observed withavapritinib was characterized mainly by the induction of myx-oid degeneration, which is a typical response pattern observedin tumors collected from patients with GIST who responded tothe treatment with imatinib in the clinic (ref. 19; Fig. 2A).

In all models, tumors in the vehicle-treated group showedhigh mitotic activity with an average of �45 mitotic figures in10 HPF. Compared with vehicle, all doses of avapritinib led to asignificant reduction of proliferation in all three xenograftmodels (Fig. 2B; Table 2). Remarkably, in the imatinib-resistantmodel UZLX-GIST9KIT 11þ17, both 10 and 30 mg/kg avapritinibinhibited tumor proliferation significantly better than theother tested agents. The antiproliferative effect of avapritinibwas comparable with imatinib in the imatinib-sensitive model,UZLX-GIST3KIT 11. In UZLX-GIST2BKIT 9, higher doses (30 and60 mg/kg) of avapritinib inhibited the proliferation significant-ly better than imatinib. Moreover, avapritinib at 60 mg/kghad the same antiproliferative effect as sunitinib. These find-ings were confirmed by Western blotting of pHH3 and byIHC for pHH3 and Ki67 (Fig. 2B; Supplementary Fig. S4A;Table 2). The expression level of pHH3 showed a near-com-plete absence in avapritinib-treated tumors at both 10 and 30mg/kg doses in UZLX-GIST9KIT 11þ17, a marked decrease in

Figure 1.

Evolution of tumor volume during the treatment, presented as relativetumor volume (% change compared with normalized, baseline value) � SDin UZLX-GIST9KIT 11þ17 (A), -GIST3KIT 11 (B), and –GIST2BKIT 9 (C).






100%80%60%40%20%

0%

Vehi

cle

Vehi

cle

Vehi

cle

Sun

itini

b

Imat

inib

Imat

inib

(50

mg/

kg)

Imat

inib

(100

mg/

kg)

Imat

inib

Reg

oraf

enib

Avap

ritin

ib (1

0 m

g/kg

)

Avap

ritin

ib (3

0 m

g/kg

)

Vehi

cle

VehicleVehicle

Sunitinib

VehicleImatinib

Imatinib

Regorafenib

Vehi

cle

Vehi

cle

Sun

itini

b

Imat

inib

Imat

inib

Imat

inib

(50

mg/

kg)

Imat

inib

(100

mg/

kg)

Reg

oraf

enib

Avap

ritin

ib (1

0 m

g/kg

)

Avapritinib (mg/kg) Avapritinib (mg/kg) Imatinib (mg/kg) Avapritinib (mg/kg)

Avap

ritin

ib (1

0 m

g/kg

)

Avap

ritin

ib (3

0 m

g/kg

)

Avap

ritin

ib (1

0 m

g/kg

)

Avap

ritin

ib (3

0 m

g/kg

)

Avap

ritin

ib (6

0 m

g/kg

)

Avap

ritin

ib (1

00 m

g/kg

)

Avap

ritin

ib (3

0 m

g/kg

)

Avap

ritin

ib (1

0 m

g/kg

)

Avap

ritin

ib (3

0 m

g/kg

)

Avap

ritin

ib (1

00 m

g/kg

)

Avap

ritin

ib (1

0 m

g/kg

)

Avap

ritin

ib (3

0 m

g/kg

)

Avap

ritin

ib (6

0 m

g/kg

)

Garde 1 Garde 2 Garde 3 Garde 4

UZLX-GIST9KIT 11+17

UZLX-GIST9KIT 11+17

UZL

X-G

IST9

KIT

11+

17U

ZLX

-GIS

T3K

IT 1

1U

ZLX

-GIS

T2B

KIT

9

UZLX-GIST3KIT 11

UZLX-GIST3KIT 11

UZLX-GIST2BKIT 9

UZLX-GIST2BKIT 9

Vehicle

Vehicle

Vehicle

Imatinib

Imatinib (50 mg/kg) Imatinib (100 mg/kg)

Imatinib

Regorafenib Avapritinib (10 mg/kg)

Avapritinib (10 mg/kg) Avapritinib (30 mg/kg) Avapritinib (100 mg/kg)

Avapritinib (10 mg/kg) Avapritinib (30 mg/kg) Avapritinib (60 mg/kg)

Avapritinib (30 mg/kg)

Sunitinib

80

60

40

20

0

Mea

n nu

mbe

r of m

itose

s pe

r 10

HP

F

6961

71 76

13 0 1 1 0

4735

19

0

23

82

10 1030 30 50 100100 10 30 60

pHH3

HH3

A

B

C

D

Figure 2.

A, Assessment of HR. B, Representative images of H&E staining after treatment (200� magnification). C, Mean number of mitotic cells in 10 HPF grouped bytreatment. D, Assessment of pHH3 expression by Western blotting.






-GIST3KIT 11, and a dose-dependent inhibition in -GIST2BKIT 9

(Fig. 2B).In two out of three models, avapritinib had significant

proapoptotic activity. In UZLX-GIST9KIT 11þ17, avapritinib(30 mg/kg) induced a significant increase in apoptosis (3.4-foldincrease compared with vehicle-treated tumors, P < 0.001,MWU). This was comparable with the effects of regorafenib(3-fold increase; Table 2). In UZLX-GIST3KIT 11, all doses ofavapritinib led to a significant and dose-dependent increase ofapoptotic activity compared with the vehicle-treated tumors.However, the difference in the induction of apoptosis betweenthe 30 and 100 mg/kg avapritinib or in comparison with imati-nib was not statistically significant. Of note, in this model, themajority of the tumor cells in actively treated tumors were re-placed by myxoid matrix, therefore counting apoptotic cellscould only be done in areas with remaining viable cells, whichmay have had an impact on the reliability of the analysis.

Evaluation of KIT signalingWestern blot analysis showed that KIT and its downstream

signaling proteins were expressed and activated in vehicle-treated tumors from all three models, as expected (Fig. 3A).In UZLX-GIST9KIT 11þ17, avapritinib (30 mg/kg) inhibitedphosphorylation of KITY703 as well as its downstream compo-nents, AKT and MAPK, and to a lesser extent, the phosphory-lation of 4EBP1. Moreover, in UZLX-GIST3KIT 11, all treatmentsinhibited the phosphorylation of KIT as well as the downstreamsignaling proteins. In addition, the expression of total forms ofthe proteins was lower in imatinib- and avapritinib-treated tu-mors (30 and 100 mg/kg) in comparison with vehicle-treated,which was most likely related to the substantial decrease of thecellularity in the response to treatment. Similarly, expressionof KIT was found to be lower in UZLX-GIST9KIT 11þ17 tumorstreated with 30 mg/kg avapritinib as compared with thevehicle-treated group (Fig. 3A). In UZLX-GIST2BKIT 9, phos-

phorylation of KITY703 and downstream proteins was inhib-ited by all treatments, with remarkable inhibition resultingfrom treatment with sunitinib and avapritinib at dose of60 mg/kg (Fig. 3A and B). Even though avapritinib did notinhibit phospho-KIT completely in this model (Fig. 3A), thephospho-KIT expression, normalized against the total form ofKIT, is significantly lower when compared with the vehicle-treated tumors (Fig. 3B). In UZLX-GIST2BKIT 9, sunitinibshowed increased ability to decrease phospho-KIT, howeverits downstream inhibitory effect was mainly through the inhi-bition of phospho-AKT and not phospho-MAPK pathway, asis seen with other effective GIST agents (Fig. 3).

Subsequently, we also performed histopathologic assess-ment of KIT signaling using pMAPK immunostaining. Theevaluation was based on both the staining intensity and per-centage of tumor area showing positivity (Supplementary TableS1). In the two models (UZLX-GIST9KIT 11þ17 and -GIST3KIT 11),we observed a strong to very strong MAPK phosphorylationin the majority of vehicle-treated tumors, whereas inUZLX-GIST2BKIT 9, staining was variable with focal positivity.In UZLX-GIST9KIT 11þ17, treatment with imatinib or regorafenibdid not affect MAPK phosphorylation, but in the majority oftumors treated with avapritinib, there was almost completeinhibition of phosphorylation, independent of a dose (Fig. 4).In UZLX-GIST3KIT 11, all active treatments reduced the MAPKphosphorylation substantially. In UZLX-GIST2BKIT 9, however,only a slight inhibition of phosphorylation was observed inavapritinib-treated tumors in comparison with the vehicle-treated group.

DiscussionPrimary and acquired resistance to treatment with established

TKI represents the ultimate challenge in the clinical setting ofGIST, as there is no approved therapy for those who progress

Table 2. Assessment of proliferation and apoptotic activity in GIST

UZLX-GIST9KIT 11þ17 UZLX-GIST3KIT 11 UZLX-GIST2BKIT 9

pHH3 Ki67 pHH3 Ki67 pHH3 Ki67

Proliferative activity Imatinib (50 mg/kg) ¼ ¼ ###a ###a ¼ ¼Imatinib (100 mg/kg) n/a n/a n/a n/a #2.5a #2.1aSunitinib n/a n/a n/a n/a ###a ###aRegorafenib ¼ ¼ n/a n/a n/a n/aAvapritinib (10 mg/kg) #14.4a ###a ###a ###a #1.3a #1.9aAvapritinib (30 mg/kg) ###a ###a ###a ###a #3.4a #3.9aAvapritinib (60 mg/kg) n/a n/a n/a n/a #13.9a #7.9aAvapritinib (100 mg/kg) n/a n/a ###a ###a n/a n/a

H&E Cleaved PARP H&E Cleaved PARP H&E Cleaved PARPApoptotic activity Imatinib (50 mg/kg) "2.1a ¼ "6.5a "6.8a ¼ ¼

Imatinib (100 mg/kg) n/a n/a n/a n/a #6.8a #2.4aSunitinib n/a n/a n/a n/a #3.7 #1.3aRegorafenib "3.0� "2.6�

n/a n/a n/a n/aAvapritinib (10 mg/kg) ¼ #2.6a "4.5a "2.3a ¼ ¼Avapritinib (30 mg/kg) "3.4a "3.0a "5.7a "17.1a #2.3a #1.9aAvapritinib (60 mg/kg) n/a n/a n/a n/a #2.6a #1.9aAvapritinib (100 mg/kg) n/a n/a "8.7a "18.1a n/a n/a

NOTE: Values are presented as fold change in comparison with the vehicle-treated tumors.Abbreviations: pHH3, phospho-histone H3.aP < 0.05 compared with the vehicle-treated by Mann–Whitney U test.#Decrease.###>50-fold decrease."Increase.¼No significant difference.






on agents with regulatory approval for treatment of patientswith GIST. In this study, we were able to document a robustantitumor activity of avapritinib, a potent and selective KITinhibitor, in three GIST PDX models characterized by differentKIT mutations and varying sensitivity to standard TKI. All threexenografts utilized in this study were previously validated forpreclinical testing of novel anti-GIST therapies (17, 20).

In the presented experiments, imatinib led to a significanttumor regression in UZLX-GIST3KIT 11 and a dose-dependenteffect on tumor volume in UZLX-GIST2BKIT 9, as already observedin multiple studies, confirming once again the stable biologicalbehavior of thismodel (13, 17, 18). These findings are also in linewith the behavior of such tumors in the clinic; patients withKIT exon 9 mutations respond better to higher dose of imatinib,as confirmed by a clinical meta-analysis (21). Furthermore, inUZLX-GIST2BKIT 9, sunitinib caused tumor regression, which isconsistent with others' observations that KIT exon 9 mutationsbenefit more from the therapy with sunitinib than with otheragents (22). Taken together, these data demonstrate that ourxenograft models exactly mirror the clinical situation and canprovide further useful guidance for clinical evaluation of noveltherapeutic approaches for GIST.

Avapritinib was designed to potently and selectively inhibitKIT exon 17 mutations (11). In vitro biochemical activity forthe KIT p.D816V–mutated receptor was achieved with anIC50 ¼ 0.27 nmol/L (11). This mutation is known to becausative for systemic mastocytosis, and is found in the vastmajority of patients with this systemic disorder (10). Further-more, avapritinib inhibits proliferation both in vitro and in vivoin leukemia models that harbor the KIT exon 17 p.N822Kmutation (23). As expected, in our PDX model with KITexon 11 and 17 mutations (UZLX-GIST9KIT 11þ17 with KIT:p.P577del; W557LfsX5; D820G), avapritinib showed a benefi-cial effect on tumor volume as well as on proliferation, whichmost likely was caused by inhibition of KIT signaling. Ofnote, the secondary p.D820G exon 17 KIT mutation, presentin our UZLX-GIST9KIT 11þ17 model, has been reported in severalTKI-resistant GIST cases with the incidence similar to othermutations affecting kinase activation loop domain of thereceptor (24–27). In our UZLX-GIST9KIT 11þ17

–resistant model,the avapritinib dose of 10 mg/kg affected pathway activation,which resulted in a remarkable decrease of proliferation, how-ever this dose led only to tumor stabilization and limited HR.On the other hand, the higher dose of the investigational agent

VehicleImatinib

VehicleVehicle

SunitinibImatinib

RegorafenibAvapritinib (mg/kg) Avapritinib (mg/kg) Avapritinib (mg/kg)Imatinib (mg/kg)10 10 30 50 10 30 6010010030

UZLX-GIST9KIT 11+17 UZLX-GIST3KIT 11 UZLX-GIST2BKIT 9

KIT

pKITY719

pKITY703

pAKTS473

AKT

MAPKpMAPK

4EBP1p4EBP1

Tubulin

Rel

ativ

e in

tens

ity v

alue

s

50%

0%

−50%

−100%

50%

0%

−50%

−100%

50%

0%

−50%

−100%pKITY703 pKITY703 pKITY703

pKITY719 pKITY719 pKITY719pAKT pAKT pAKT

pMAPK pMAPKpMAPK

p4EBP1 p4EBP1p4EBP1

Imatinib ImatinibImatinib (50 mg/kg) Imatinib (100 mg/kg)

Regorafenib Avapritinib (10 mg/kg) Avapritinib (30 mg/kg) Avapritinib (10 mg/kg) Avapritinib (10 mg/kg) Avapritinib (30 mg/kg) Avapritinib (60 mg/kg)Avapritinib (30 mg/kg) Avapritinib (100 mg/kg)Sunitinib

A

B

Figure 3.

KIT signaling pathway. A, Assessment of the effect of treatments in different xenograft models. B, Densitometric assessment of phospho-protein forms in KITsignaling pathway.






(30 mg/kg) caused a striking tumor volume shrinkage (to 27%of baseline) and an impressive HR with complete absence ofproliferative activity. This observation is similar to findings byEvans and colleagues where avapritinib potently inhibited themouse equivalent of KIT p.D816Y–driven tumor growth in vivoin a dose-dependent manner (11). Moreover, in a GIST PDXmodel derived from a refractory GIST tumor that harborsKIT exon 11/17 (p.K557_K558del; Y823D), avapritinib againled to tumor regression from the baseline value (11). Interest-ingly, in our experiments, the pattern of response observed inUZLX-GIST9KIT 11þ17 tumors treated with 30 mg/kg was char-acterized by myxoid degeneration; a phenomenon whereviable tumor cells are replaced by an amorphous collagenousmatrix containing only a few scattered cells. Myxoid degener-ation is described frequently as a feature characteristic forGIST responding to the treatment with imatinib, both inpreclinical and clinical settings (18, 19). Although in thisresistant model, the effect of 30 mg/kg avapritinib on thetumor volume and histologic features was significantly better

than what was achieved with 10 mg/kg, both treatment groupsexposed to this novel TKI had better responses than imatinib-or regorafenib-treated tumors. The antitumor activity of ava-pritinib is currently being evaluated in GIST (NCT02508532and NCT03465722; ref. 28). Of note, the maximum tolerateddose of avapritinib in patients is 400 mg/day whereas the doseof 300 mg/day is being evaluated in the currently ongoingphase III trial (NCT03465722). The animal equivalent dose,calculated on the basis of the body surface (29), is respectively82 and 61.5 mg/kg, which falls within the range of doses testedin this study.

Subsequently, we evaluated avapritinib in two additionalPDX models with a primary mutation in KIT exon 11(UZLX-GIST3KIT 11, imatinib sensitive) or exon 9 (UZLX-GIST2BKIT 9, dose-dependent sensitivity to imatinib, sensitiveto sunitinib). We observed a significant inhibition of prolifer-ation in these models at all doses tested, suggesting broaderinhibitory capacity of avapritinib against different KIT muta-tions outside of the activation loop region (11). In the UZLX-

100%

80%

60%

40%

20%

0%

Vehi

cle

Imat

inib

Reg

oraf

enib

Regorafenib

Avap

ritin

ib (1

0 m

g/kg

)

Avap

ritin

ib (3

0 m

g/kg

)


Avapritinib (30 mg/kg) Avapritinib (100 mg/kg)



Vehi

cle

Vehi

cle

Imat

inib

Sun

itini

b

Avap

ritin

ib (1

0 m

g/kg

)

Imat

inib

(50

mg/

kg)

Imat

inib

(100

mg/

kg)

Avap

ritin

ib (3

0 m

g/kg

)

Avap

ritin

ib (1

00 m

g/kg

)

Avap

ritin

ib (1

0 m

g/kg

)

Avap

ritin

ib (3

0 m

g/kg

)

Avap

ritin

ib (6

0 m

g/kg

)

UZLX-GIST9KIT 11+17

UZL

X-G

IST9

KIT

11+

17

UZLX-GIST3KIT 11

UZL

X-G

IST3

KIT

11

UZLX-GIST2BKIT 9

UZL

X-G

IST2

BK

IT 9

Per

cent

of t

umor

s

Negative (-) Very weak (±) Weak (+) Strong (++) Very strong (+++)

Vehicle

Vehicle

Vehicle

Imatinib

Imatinib

Imatinib (50 mg/kg) Imatinib (100 mg/kg) Avapritinib (10 mg/kg) Avapritinib (30 mg/kg) Avapritinib (60 mg/kg)Sunitinib

A

B

Figure 4.

A, Evaluation of pMAPK positivity based on the intensity and percent tumor area showing pMAPK positivity. Grading system is presented in SupplementaryTable S1. B, Representative images of pMAPK immunostaining of the different treatment groups at 200�.






GIST3KIT 11 model, we found a pronounced effect on tumorvolume and histologic responses at 30 and 100 mg/kg, and thisobservation was similar to the effect caused by imatinib. On theother hand, avapritinib in UZLX-GIST2BKIT 9 was better thanimatinib, but only the dose of 60 mg/kg led to a similar efficacyas achieved with sunitinib. This observation suggests thatavapritinib could also be effective in GIST with primary muta-tions, however higher in vivo concentrations are required forexon 9 antitumor activity. Our studies in mice suggest avapri-tinib is well tolerated at these higher concentrations, butclinical data will be required to fully assess the activity ofavapritinib in patients with KIT exon 9–driven tumors Theongoing phase I clinical trial with avapritinib accepts patientswho have failed two or more agents and likely have accumu-lated secondary resistance mutations, and patients withPDGFRA p.D842 mutant–driven GIST independent of priorlines of treatment. Interestingly, it is already known that ava-pritinib inhibits the activity of PDGFRA p.D842V–mutatedreceptor in vitro (11); the in vivo evaluation is hampered bythe lack of relevant models with this mutation.

In our study, we found only a moderate proapoptotic effectin UZLX-GIST3KIT 11 and -GIST9KIT 11þ17 (at a dose of 30 mg/kg),in part explained by the presence of myxoid degeneration as aresponse to avapritinib in these models. The reduced cellularitypotentially leads to an underestimation of the proapoptoticeffects of this treatment in our models. This speculation is sup-ported by the observation of increased apoptosis in tumorsharvested after 8 days of treatment with avapritinib, when thehistologic response was not as pronounced as on day 16 (11). Theproapoptotic effect of avapritinib is likely induced by inhibitionof KIT signaling. This observation matches previous findingsreported by our group, where potent KIT signaling inhibitionresulted in an increased apoptosis (16, 17). Moreover, Evans andcolleagues showed proapoptotic activity of avapritinib in amousemastocytoma cell linewith KIT exon 17 substitution p.D814Y, theequivalent of human p.D816Y mutant (10, 23). On the otherhand, in the UZLX-GIST2BKIT 9, neither of the experimentaltreatments led to an increase of apoptotic activity after 16 daysof treatment. A trend toward a decline of apoptosis was observedwith increasing dose of avapritinib. Similar observations werepreviously reported in this model, where treatments that led toinhibition of KIT signaling resulted in a decrease of apoptoticactivity (17, 20). This effect could be a consequence of the variableexpression of signaling molecules due to specific KIT genotype asreported by several groups (30–32).

Treatment with avapritinib was well tolerated in our nudemice. Although we observed yellow skin discoloration in micetreated with the highest avapritinib dose tested (100 mg/kg), itdid not impact the animals' well-being. We also did not findany macroscopic or microscopic changes in their organs,including the liver. The skin color change may be due to astronger systemic inhibition of KIT, disrupting its physiologicfunction in follicular melanocytes and or impairing melano-genesis (33). Skin depigmentation as a result of strong KITinhibition was previously reported by Kim and colleagues inthe evaluation of another potent KIT inhibitor, PLX3397 andwas reported in metastatic renal cell carcinoma, treated withTKI sorafenib (34, 35).

As a selective inhibitor of KIT and PDGFRA p.D842V muta-tions, avapritinib is less likely to cause significant off targettreatment-related toxicity at efficacious doses, in contrast to

multitargeted agents currently being used in the clinical setting.Agents such as sunitinib, regorafenib, dasatinib, or ponatinibshow effectiveness in some genotypes of resistant GIST, butdose-limiting toxicities, arising from the simultaneous block-age of several kinases, translates into higher toxicity and limitthe clinical usefulness of some of these agents (36, 37).Although very preliminary data from the ongoing phase Iclinical trial suggests a favorable safety profile (28), the highspecificity of avapritinib can in theory increase the risk of rapiddevelopment of secondary resistance to this compound. GIST isknown for its clinical inter- and intratumoral heterogeneity interms of the mutational profile, and it is likely that some clonesof this disease may lead to further progression on treatmentwith the novel agent. In this context, it is noteworthy that wesaw broad activity in a variety of GIST genotype in our mice.

In conclusion, we provide in vivo evidence that the novel TKIavapritinib has significant antitumor activity in GIST PDXmodels. Our results demonstrate that in KIT exon 11þ17double mutated GIST, this inhibitor is more active than estab-lished standard treatments. Moreover, in imatinib-sensitivemodels with primary KIT mutations, avapritinib shows a sim-ilar or even higher level of activity in comparison with imatinib.In all models tested, the pharmacologic antiproliferative effecton tumor volume was mainly achieved through a markedinhibition of KIT signaling. Our data strongly support thescientific rationale of the ongoing exploration of avapritinibin GIST and provide arguments for exploration of the novelcompound in the clinic in both imatinib-sensitive and TKI-resistant genotypes of this mesenchymal malignancy. Theresults seen in our mouse PDXs and the early findings reportedfrom the ongoing clinical trial in patients with GIST suggest thatavapritinib has the potential to become an important treatmentoption for this orphan malignancy.

Disclosure of Potential Conflicts of InterestE. Evans, A.K. Gardino, and C. Lengauer have ownership interests (including

patents) in BlueprintMedicines. Nopotential conflicts of interest were disclosedby the other authors.

Authors' ContributionsConception and design: A. Wozniak, E. Evans, C. Lengauer, P. Sch€offskiDevelopment of methodology: Y.K Gebreyohannes, A. Wozniak, J. Wellens,M. Debiec-Rychter, P. Sch€offskiAcquisition of data (provided animals, acquired and managed patients,provided facilities, etc.): Y.K Gebreyohannes, A. Wozniak, M.-E. Zhai,J. Wellens, J. Cornillie, R. Sciot, P. Sch€offskiAnalysis and interpretation of data (e.g., statistical analysis, biostati-stics, computational analysis): Y.K Gebreyohannes, A. Wozniak, M.-E. Zhai,E. Evans, C. Lengauer, R. Sciot, P. Sch€offskiWriting, review, and/or revision of the manuscript: Y.K Gebreyohannes,A. Wozniak, M.-E. Zhai, J. Cornillie, E. Evans, A.K. Gardino, C. Lengauer, M.Debiec-Rychter, R. Sciot, P. Sch€offskiAdministrative, technical, or material support (i.e., reporting or organiz-ing data, constructing databases): J. Wellens, U. Vanleeuw, M. Debiec-Rychter,P. Sch€offskiStudy supervision: E. Evans, C. Lengauer, P. Sch€offski

The costs of publication of this article were defrayed in part by thepayment of page charges. This article must therefore be hereby markedadvertisement in accordance with 18 U.S.C. Section 1734 solely to indicatethis fact.

Received June 13, 2018; revisedAugust 9, 2018; accepted September 25, 2018;published first October 1, 2018.






References1. Joensuu H, Hohenberger P, Corless CL. Gastrointestinal stromal tumour.

Lancet 2013;382:973–83.2. Corless CL, Barnett CM, Heinrich MC. Gastrointestinal stromal tumours:

origin and molecular oncology. Nat Rev Cancer 2011;11:865–78.3. Rutkowski P, Hompes D. Combined therapy of gastrointestinal stromal

tumors. Surg Oncol Clin N Am 2016;25:735–59.4. Nishida T, Blay JY, Hirota S, Kitagawa Y, Kang YK. The standard diagnosis,

treatment, and follow-up of gastrointestinal stromal tumors based onguidelines. Gastric Cancer 2016;19:3–14.

5. Demetri GD, von Mehren M, Blanke CD, Van den Abbeele AD,Eisenberg B, Roberts PJ, et al. Efficacy and safety of imatinib mesylatein advanced gastrointestinal stromal tumors. N Engl J Med 2002;347:472–80.

6. Metaxas Y, Oikonomopoulos G, Pentheroudakis G. Update on clinicalresearch and state of the art management of patients with advancedsarcomas and GIST. ESMO Open 2016;1:e000065.

7. George S, Wang Q, Heinrich MC, Corless CL, Zhu M, Butrynski JE, et al.Efficacy and safety of regorafenib in patients with metastatic and/orunresectable GI stromal tumor after failure of imatinib and sunitinib: amulticenter phase II trial. J Clin Oncol 2012;30:2401–7.

8. Demetri GD, Reichardt P, Kang YK, Blay JY, Rutkowski P, Gelderblom H,et al. Efficacy and safety of regorafenib for advanced gastrointestinalstromal tumours after failure of imatinib and sunitinib (GRID): aninternational, multicentre, randomised, placebo-controlled, phase 3 trial.Lancet 2013;381:295–302.

9. Abid A, Malone MA, Curci K. Mastocytosis. Prim Care 2016;43:505–18.10. Evans EK, Hodous BL, Gardino A, Zhu J, Shutes A, Davis A, et al. First

selective KIT D816V inhibitor for patients with systemic mastocytosis.Blood 2014;124:3217.

11. Evans EK, Gardino A, Kim JL, Hodous BL, Shutes A, Davis A, et al.A precision therapy against cancers driven by KIT/PDGFRA mutations.Sci Transl Med 2017;9:pii:eaao1690.

12. Wozniak A, Rutkowski P, Sch€offski P, Ray-Coquard I Hostein I, SchildhausHU, et al. Tumor genotype is an independent prognostic factor in primarygastrointestinal stromal tumors of gastric origin: a European multicenteranalysis based on ConticaGIST. Clin Cancer Res 2014;20:6105–16.

13. Van Looy T, Gebreyohannes YK, Wozniak A, Cornillie J, Wellens J, Li H,et al. Characterization and assessment of the sensitivity and resistanceof a newly established human gastrointestinal stromal tumour xeno-graft model to treatment with tyrosine kinase inhibitors. Clin SarcomaRes 2014;4:10.

14. Antonescu CR, Besmer P, Guo T, Arkun K, Hom G, Koryotowski B, et al.Acquired resistance to imatinib in gastrointestinal stromal tumor occursthrough secondary gene mutation. Clin Cancer Res 2005;11:4182–90.

15. Agaram NP, Besmer P, Wong GC, Guo T, Socci ND, Maki RG, et al.Pathologic and molecular heterogeneity in imatinib-stable or imatinib-responsive gastrointestinal stromal tumors. Clin Cancer Res 2007;13:170–81.

16. Floris G, Sciot R, Wozniak A, Van Looy T, Wellens J, Faa G, et al. The novelHSP90 inhibitor, IPI-493, is highly effective in human gastrostrointestinalstromal tumor xenografts carrying heterogeneous KIT mutations. ClinCancer Res 2011;17:5604–14.

17. Floris G, Wozniak A, Sciot R, Li H, Friedman L, Van Looy T, et al. A potentcombination of the novel PI3K Inhibitor, GDC-0941, with imatinib ingastrointestinal stromal tumor xenografts: long-lasting responses aftertreatment withdrawal. Clin Cancer Res 2013;19:620–30.

18. Van Looy T, Wozniak A, Floris G, Sciot R, Li H, Wellens J, et al.Phosphoinositide 3-kinase inhibitors combined with imatinib inpatient-derived xenograft models of gastrointestinal stromal tumors:rationale and efficacy. Clin Cancer Res 2014;20:6071–82.

19. Sciot R, Debiec-Rychter M. GIST under imatinib therapy. Semin DiagnPathol 2006;23:84–90.

20. Gebreyohannes YK, Sch€offski P, Van Looy T, Wellens J, Vreys L, Cornillie J,et al. Cabozantinib is active against human gastrointestinal stromaltumor xenografts carrying different KIT mutations. Mol Cancer Ther2016;15:2845–52.

21. Gastrointestinal Stromal Tumor Meta-Analysis Group (MetaGIST).Com-parison of two doses of imatinib for the treatment of unresectable ormetastatic gastrointestinal stromal tumors: a meta-analysis of 1,640patients. J Clin Oncol 2010;28:1247–53.

22. Reichardt P, Demetri GD, Gelderblom H, Rutkowski P, Im SA, GuptaS, et al. Correlation of KIT and PDGFRA mutational status withclinical benefit in patients with gastrointestinal stromal tumor treatedwith sunitinib in a worldwide treatment-use trial. BMC Cancer2016;16:22.

23. Evans EK, Hodous BL, Gardino A, Davis A, Zhu J, et al. BLU-285, a potentand selective inhibitor for hematologic malignancies with KIT exon 17mutations. Blood 2015;126:568.

24. Heinrich MC, Corless CL, Demetri GD, Blanke CD, von Mehren M,Joensuu H, et al. Kinase mutations and imatinib response in patientswith metastatic gastrointestinal stromal tumor. J Clin Oncol 2003;21:4342–9.

25. Wardelmann E, Merkelbach-Bruse S, Pauls K, Thomas N, SchildhausHU, Heinicke T, et al. Polyclonal evolution of multiple secondary KITmutations in gastrointestinal stromal tumors under treatment withimatinib mesylate. Clin Cancer Res 2006;12:1743–9.

26. Liegl B, Kepten I, Le C, Zhu M, Demetri GD, Heinrich MC, et al. Hetero-geneity of kinase inhibitor resistance mechanisms in GIST. J Pathol2008;216:64–74.

27. Heinrich MC, Maki RG, Corless CL, Antonescu CR, Harlow A, Griffith D,et al. Primary and secondary kinase genotypes correlate with the biologicaland clinical activity of sunitinib in imatinib-resistant gastrointestinalstromal tumor. J Clin Oncol 2008;26:5352–9.

28. Heinrich MC, Jones R, Schoffski P, Bauer S, von Mehren M, Eskens F,et al. Preliminary safety and activity in a first-in-human phase 1 study ofBLU-285, a potent, highly-selective inhibitor of KIT and PDGFR alphaactivation loop mutants in advanced gastrointestinal stromal tumor(GIST). Eur J Cancer 2016;68:abstract 6LBA.

29. Nair AB, Jacob S. A simple practice guide for dose conversion betweenanimals and human. J Basic Clin Pharm 2016;7:27–31.

30. Landuyt B, Prenen H, Debiec-Rychter M, Sciot R, de Bruijn EA, vanOosterom AT. Differential protein expression profile in gastrointestinalstromal tumors. Amino Acids 2004;27:335–7.

31. Antonescu CR, Viale A, Sarran L, Tschernyavsky SJ, Gonen M, Segal NH,et al. Gene expression in gastrointestinal stromal tumors is distin-guished by KIT genotype and anatomic site. Clin Cancer Res 2004;10:3282–90.

32. Duensing A, Medeiros F, McConarty B, Joseph NE, Panigrahy D, Singer S,et al. Mechanisms of oncogenic KIT signal transduction in primary gas-trointestinal stromal tumors (GISTs). Oncogene 2004;23:3999–4006.

33. D'Mello SAN, Finlay GJ, Baguley BC, Askarian-Amiri ME. Signaling path-ways in melanogenesis. Int J Mol Sci 2016;17:1144.

34. Kim TS, Cavnar MJ, Cohen NA, Sorenson EC, Greer JB, Seifert AM, et al.Increased KIT inhibition enhances therapeutic efficacy in gastrointestinalstromal tumor. Clin Cancer Res 2014;20:2350–62.

35. Dasanu CA, Alexandrescu DT, Dutcher J. Yellow skin discoloration asso-ciated with sorafenib use for treatment of metastatic renal cell carcinoma.South Med J 2007;100:328–30.

36. Ashman LK, Griffith R. Therapeutic targeting of c-KIT in cancer. ExpertOpin Investig Drugs 2013;22:103–15.

37. Garner AP, Gozgit JM, Anjum R, Vodala S, Schrock A, Zhou T, et al.Ponatinib inhibits polyclonal drug-resistant KIT oncoproteins andshows therapeutic potential in heavily pretreated gastrointestinal stromaltumor (GIST) patients. Clin Cancer Res 2014;20:5745–55.






2019;25:609-618. Published OnlineFirst October 1, 2018.Clin Cancer Res Yemarshet K. Gebreyohannes, Agnieszka Wozniak, Madalina-Elena Zhai, et al. Gastrointestinal Stromal Tumorsof Mutated KIT, in Patient-derived Xenograft Models of Robust Activity of Avapritinib, Potent and Highly Selective Inhibitor

Updated version

10.1158/1078-0432.CCR-18-1858doi:

Access the most recent version of this article at:

Material

Supplementary

http://clincancerres.aacrjournals.org/content/suppl/2018/09/29/1078-0432.CCR-18-1858.DC1

Access the most recent supplemental material at:

Cited articles

http://clincancerres.aacrjournals.org/content/25/2/609.full#ref-list-1

This article cites 36 articles, 17 of which you can access for free at:

Citing articles

http://clincancerres.aacrjournals.org/content/25/2/609.full#related-urls

This article has been cited by 3 HighWire-hosted articles. Access the articles at:

E-mail alerts related to this article or journal.Sign up to receive free email-alerts

Subscriptions

Reprints and

[email protected]

To order reprints of this article or to subscribe to the journal, contact the AACR Publications Department at

Permissions

Rightslink site. Click on "Request Permissions" which will take you to the Copyright Clearance Center's (CCC)

.http://clincancerres.aacrjournals.org/content/25/2/609To request permission to re-use all or part of this article, use this link



http://clincancerres.aacrjournals.org/lookup/doi/10.1158/1078-0432.CCR-18-1858

http://clincancerres.aacrjournals.org/content/suppl/2018/09/29/1078-0432.CCR-18-1858.DC1

http://clincancerres.aacrjournals.org/content/25/2/609.full#ref-list-1

http://clincancerres.aacrjournals.org/content/25/2/609.full#related-urls

http://clincancerres.aacrjournals.org/cgi/alerts

mailto:[email protected]

http://clincancerres.aacrjournals.org/content/25/2/609


robust activity of avapritinib, potent and highly ... · the dosing solutions of imatinib,...

Documents