Axitinib overcomes multiple imatinib resistant cKIT mutations including the gatekeeper mutation T670I in gastrointestinal stromal tumors

Background: cKIT kinase overexpression and gain-of-function mutations are the critical pathogenesis of gastrointestinal stromal tumors (GISTs). Although the multiple kinase inhibitors such as imatinib, sunitinib, and regorafenib have been approved for GISTs, the acquisition of polyclonal secondary resistance mutations in KIT is still a limitation for GIST treatment. Here we explored the KIT inhibitory activity of axitinib in preclinical models and describe initial characterization of its activity in GIST patient-derived primary cells. Methods: The activities of axitinib against mutant KIT were evaluated using protein-based assay and a panel of engineered and GIST-derived cell lines. The binding modes of axitinib-KIT/KIT mutants were analyzed. Four primary cells derived from GIST patients were also used to assess the drug response of axitinib. Results: Axitinib exhibited potent activities against a variety of cKIT associated primary and secondary mutations. It displayed better activity against cKIT wild-type, cKIT V559D/A/G, and L576P primary gain-of-function mutations than imatinib, sunitinib, and regorafenib. In addition, it could inhibit imatinib resistant cKIT T670I and V654A mutants in vitro and in vivo GIST preclinical models. Conclusion: Our results provide the basis for extending the application of axitinib to GISTs patients who are unresponsive or intolerant to the current therapies.

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Molecular Correlates of Imatinib Resistance in Gastrointestinal Stromal Tumors

Journal of Clinical Oncology ◽

10.1200/jco.2006.06.2265 ◽

2006 ◽

Vol 24 (29) ◽

pp. 4764-4774 ◽

Cited By ~ 528

Author(s):

Michael C. Heinrich ◽

Christopher L. Corless ◽

Charles D. Blanke ◽

George D. Demetri ◽

Heikki Joensuu ◽

...

Keyword(s):

Gastrointestinal Stromal Tumors ◽

Molecular Mechanisms ◽

Clonal Evolution ◽

Clinical Problem ◽

Primary Resistance ◽

Imatinib Resistance ◽

Secondary Resistance ◽

Stromal Tumors ◽

Rnai Technology ◽

Imatinib Resistant

Purpose Gastrointestinal stromal tumors (GISTs) commonly harbor oncogenic mutations of the KIT or platelet-derived growth factor alpha (PDGFRA) kinases, which are targets for imatinib. In clinical studies, 75% to 90% of patients with advanced GISTs experience clinical benefit from imatinib. However, imatinib resistance is an increasing clinical problem. Patients and Methods One hundred forty-seven patients with advanced, unresectable GISTs were enrolled onto a randomized, phase II clinical study of imatinib. Specimens from pretreatment and/or imatinib-resistant tumors were analyzed to identify molecular correlates of imatinib resistance. Secondary kinase mutations of KIT or PDGFRA that were identified in imatinib-resistant GISTs were biochemically profiled for imatinib sensitivity. Results Molecular studies were performed using specimens from 10 patients with primary and 33 patients with secondary resistance. Imatinib-resistant tumors had levels of activated KIT that were similar to or greater than those typically found in untreated GISTs. Secondary kinase mutations were rare in GISTs with primary resistance but frequently found in GISTs with secondary resistance (10% v 67%; P = .002). Evidence for clonal evolution and/or polyclonal secondary kinase mutations was seen in three (18.8%) of 16 patients. Secondary kinase mutations were nonrandomly distributed and were associated with decreased imatinib sensitivity compared with typical KIT exon 11 mutations. Using RNAi technology, we demonstrated that imatinib-resistant GIST cells remain dependent on KIT kinase activity for activation of critical downstream signaling pathways. Conclusion Different molecular mechanisms are responsible for primary and secondary imatinib resistance in GISTs. These findings have implications for future approaches to the growing problem of imatinib resistance in patients with advanced GISTs.

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Ethyl-2-amino-pyrrole-3-carboxylates are active against imatinib-resistant gastrointestinal stromal tumors in vitro and in vivo

Anti-Cancer Drugs ◽

10.1097/cad.0000000000000753 ◽

2019 ◽

Vol 30 (5) ◽

pp. 475-484

Author(s):

Sergei Boichuk ◽

Aigul Galembikova ◽

Pavel Dunaev ◽

Ekaterina Micheeva ◽

Maria Novikova ◽

...

Keyword(s):

Gastrointestinal Stromal Tumors ◽

Stromal Tumors ◽

Imatinib Resistant

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Inhibition of PI3K and MAPK pathways along with KIT inhibitors as a strategy to overcome drug resistance in gastrointestinal stromal tumors

PLoS ONE ◽

10.1371/journal.pone.0252689 ◽

2021 ◽

Vol 16 (7) ◽

pp. e0252689

Author(s):

Anu Gupta ◽

Shuang Ma ◽

Kepeng Che ◽

Ajaybabu V. Pobbati ◽

Brian P. Rubin

Keyword(s):

Drug Resistance ◽

Tyrosine Kinases ◽

Gastrointestinal Stromal Tumors ◽

Mapk Signaling ◽

Resistance Mutations ◽

Secondary Resistance ◽

Stromal Tumors ◽

Activating Mutations ◽

Mapk Pathways ◽

Induced Apoptosis

Activating mutations in KIT/PDGFRA receptor tyrosine kinases drive gastrointestinal stromal tumors (GIST). KIT/PDGFRA inhibitors, such as imatinib do not evoke an effective cytocidal response, leaving room for quiescence and development of multiple secondary resistance mutations. As the majority of the secondary resistance clones activate PI3K and MAPK pathways, we investigated whether combined targeting of KIT/PI3K/MAPK (KPM) pathways overcomes drug resistance and quiescence in GIST cells. We monitored the proliferation of imatinib–sensitive and–resistant GIST cell lines after treating them with various combinations of drugs to inhibit KPM pathways. Cytocidal response was evaluated through proliferation, apoptosis and colony outgrowth assays. Combined inhibition of KPM signaling pathways using a KPM inhibitor cocktail decreased the survival of drug-resistant GIST cells and dramatically reduced their proliferation. Downstream pathway analysis showed that the residual PI3K/MAPK signaling observed after KIT inhibitor treatment plays a role in mediating quiescence and drug resistance. The KPM inhibitor cocktail with sunitinib or regorafenib effectively induced apoptosis and prevented colony outgrowth after long-term drug removal, suggesting that it can be used as an effective strategy against quiescence and drug resistance in metastatic GIST.

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Activity of AT13387, a novel, non-ansamycin inhibitor of heat shock protein 90, against gastrointestinal stromal tumors (GIST).

Journal of Clinical Oncology ◽

10.1200/jco.2013.31.4_suppl.105 ◽

2013 ◽

Vol 31 (4_suppl) ◽

pp. 105-105 ◽

Cited By ~ 2

Author(s):

Daruka Mahadevan ◽

Geoffrey Shapiro ◽

Sandra E. Kurtin ◽

James M. Cleary ◽

John F. Lyons ◽

...

Keyword(s):

Phase I ◽

Kinase Inhibitor ◽

Heat Shock Protein 90 ◽

Tumor Growth Inhibition ◽

Resistance Mutations ◽

Hsp90 Inhibition ◽

Secondary Resistance ◽

Imatinib Resistant

105 Background: AT13387 is a second-generation potent, novel non-ansamycin HSP90 inhibitor (Kd 0.71nM). The majority of GIST tumors are characterized by activating mutations of c-KIT, an HSP90 client protein. Secondary resistance mutations within c-KIT limit clinical responses to TKIs. The dependence of c-KIT and its mutated forms on HSP90 suggests that HSP90 inhibition may be a valuable treatment option for imatinib-sensitive and resistant clones. In vitro, AT13387 inhibited the proliferation of imatinib-sensitive (GIST882, GIST-T1) and imatinib-resistant (GIST430, GIST48) cell lines. In vivo, AT13387 demonstrated anti-tumor activity in the imatinib-sensitive (GIST-PSW) and imatinib-resistant (GIST430) xenograft models. Induction of HSP70, depletion of phospho-c-KIT and inhibition of c-KIT signaling were observed in both models. Combination treatment of imatinib and AT13387 in the GIST430 model was well tolerated and significantly enhanced tumor growth inhibition over either monotherapies. Methods: In a completed phase I study, AT13387 was administered IV over 1 hour twice weekly or weekly of a 28-day cycle in a standard 3+3 dose-escalation design. The primary endpoint was to determine the MTD; secondary endpoints included PK, PD, safety and tolerability. Results: The PK exposures were dose-dependent and linear. AT13387 was well tolerated on both schedules. DLTs included primarily G2 AEs of GI toxicities, fatigue and infusion site reactions. The once weekly RP2D was determined to be 260 mg/m2. HSP70 induction was 2–7 fold at higher doses. A total of 7 GIST subjects were enrolled. An objective and durable PR was observed in one subject and 2 SDs at 8, 7 and 11 months, respectively. The PR subject demonstrated molecular resistance to kinase inhibitor treatment in the c-KIT gene prior to initiation of AT13387 therapy in two resected lesions by harboring the same activating c-KIT deletion in exon 11 and two separate TKI resistance mutations in exon 17. Conclusions: Overall, these results suggest AT13387 is a promising agent in GIST, including TKI-resistant c-Kit positive GIST. AT13387 is currently being evaluated in combination with imatinib in an ongoing phase I/II study. Clinical trial information: NCT01294202.

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A Novel Kindred with Familial Gastrointestinal Stromal Tumors Caused by a Rare KIT Germline Mutation (N655K): Clinico-Pathological Presentation and TKI Sensitivity

Journal of Personalized Medicine ◽

10.3390/jpm10040234 ◽

2020 ◽

Vol 10 (4) ◽

pp. 234

Author(s):

Mara Fornasarig ◽

Daniela Gasparotto ◽

Luisa Foltran ◽

Michele Campigotto ◽

Sara Lombardi ◽

...

Keyword(s):

Germline Mutation ◽

Gastrointestinal Stromal Tumors ◽

Kinase Inhibitors ◽

Age Of Onset ◽

Mesenchymal Tumors ◽

Kit Mutation ◽

Stromal Tumors ◽

Activating Mutations ◽

Imatinib Resistant

Gastrointestinal stromal tumors (GISTs), the most common mesenchymal tumors of the gastrointestinal tract, are characterized by activating mutations in KIT or PDGFRA genes. The vast majority of GISTs are sporadic, but rare hereditary forms have been reported, often featuring multifocality and younger age of onset. We here report the identification of a novel kindred affected by familial GIST caused by a KIT germline mutation in exon 13 (N655K). No family affected by hereditary GIST due to this KIT variant has been reported in literature so far. We were able to track the mutation in three members of the family (proband, mother, and second-degree cousin), all affected by multiple GISTs. Due to its rarity, the N655K variant is poorly characterized. We conducted in vitro drug sensitivity assays that indicated that most tyrosine kinase inhibitors (TKIs) currently included in the therapeutic armamentarium for GISTs have a limited inhibitory activity toward this mutation. However, when compared to a classical imatinib-resistant KIT mutation (T670I), N655K was slightly more sensitive to imatinib, and encouraging responses were observed with last-generation TKIs.

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Preclinical development of HQP1351, a multikinase inhibitor targeting a broad spectrum of mutant KIT kinases, for the treatment of imatinib-resistant gastrointestinal stromal tumors

Cell & Bioscience ◽

10.1186/s13578-019-0351-6 ◽

2019 ◽

Vol 9 (1) ◽

Author(s):

Xuechao Liu ◽

Guangfeng Wang ◽

Xianglei Yan ◽

Haibo Qiu ◽

Ping Min ◽

...

Keyword(s):

Cell Cycle ◽

Clinical Trials ◽

Cell Migration ◽

Cell Lines ◽

Gastrointestinal Stromal Tumors ◽

Migration And Invasion ◽

Stromal Tumors ◽

Imatinib Resistant

Abstract Background Imatinib shows limited efficacy in patients with gastrointestinal stromal tumors (GISTs) carrying secondary KIT mutations. HQP1351, an orally bioavailable multikinase BCR-ABL inhibitor, is currently in clinical trials for the treatment of T315I mutant chronic myelogenous leukemia (CML), but the potential application in imatinib-resistant GISTs carrying secondary KIT mutations has not been explored. Methods The binding activities of HQP1351 with native or mutant KIT were first analyzed. Imatinib-sensitive GIST T1 and imatinib-resistant GIST 430 cells were employed to test the in vitro antiproliferative activity. Colony formation assay, cell migration assay and cell invasion assay were performed to evaluate the clonogenic, migration and invasion ability respectively. Flow cytometry and western blot analysis were used to detect cell apoptosis, cell cycle and signaling pathway. In vivo antitumor activity was evaluated in mouse xenograft models derived from GIST cell lines. Results HQP1351 potently inhibited both wild-type and mutant KIT kinases. In both imatinib-resistant and sensitive GIST cell lines, HQP1351 exhibited more potent or equivalent antiproliferative activity compared with ponatinib, a third generation BCR-ABL and KIT inhibitor. HQP1351 led to more profound inhibition of cell colony formation, cell migration and invasion, cell cycle arrest and cell apoptosis than ponatinib. Furthermore, HQP1351 also inhibited p-KIT, p-AKT, p-ERK1/2, and p-STAT3 to a higher extent than ponatinib. Finally, in xenograft tumor models derived from imatinib-resistant GIST cancer cell lines, HQP1351 exhibited antitumor activity superior to ponatinib. Conclusions Collectively, our in vitro and in vivo results suggest that the therapeutic application of HQP1351 in imatinib-resistant GIST patients deserves further investigation in clinical trials.

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Surgical interventions for focal progression of gastrointestinal stromal tumors under imatinib therapy

Journal of Clinical Oncology ◽

10.1200/jco.2006.24.18_suppl.9548 ◽

2006 ◽

Vol 24 (18_suppl) ◽

pp. 9548-9548

Author(s):

T. Nishida ◽

J. Hasegawa ◽

A. Nishitani ◽

T. Takahashi ◽

T. Kanda ◽

...

Keyword(s):

Gastrointestinal Stromal Tumors ◽

Bile Leak ◽

Imatinib Therapy ◽

Secondary Resistance ◽

Surgical Interventions ◽

Stromal Tumors ◽

Imatinib Resistant ◽

Chronic Therapy ◽

Hospital Deaths

9548 Background: Although Imatinib has shown high activity oin advanced gastrointestinal stromal tumors (GIST), secondary resistance appears as focal or systemic progressions during chronic therapy. At present, there are limited therapeutic options for Imatinib-resistant GIST. This retrospective study examines the safety and short-term outcomes of surgical interventions for focal progressions under Imatinib. Patients and Methods: Between Jan. 2002 and May 2005, 16 patients (pts) with focal progressions of secondarily-resistant GIST (Male:Female, 12:4; median age, 61 years) underwent surgical interventions including resection (N=13), radiofrequency ablation (RFA) (n=2), and their combination (n=1). Doses of Imatinib were 400 (n=10) or 600 (n=6) mg/day. Results: Postoperative complications including liver abscess (n=1), minor bile leak (n=1), wound infection (n=1) and transient ileus (n=1) were occurred to 4 patients, who recovered from them within a few weeks. There was no in-hospital deaths. Median time to progression (TTP) was 5.5 months. Only one patient died of the of disease, 16 months after the resection, and the remaining 15 pts are alive, with a median follow-up of 13 months. Pts with total eradication of resistant lesions (n=7) had longer TTP than those with incomplete (n=9) (p<0.05). Total eradication could be performed in patients with a smaller number (P=0.014) and size (P=0.018) of resistant lesions. Overall survival after Imatinib therapy was 100% at 1 year and 93% at 3 years, with a median follow-up of 39 months. Conclusions: These data suggest that surgical interventions for focal progressions of secondarily-resistant GIST under Imatinib may be safe and that total eradication of resistant lesions may result in a survival benefits under conditions of limited treatment modality. No significant financial relationships to disclose.

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Use of ponatinib to inhibit kinase mutations associated with drug-resistant gastrointestinal stromal tumors (GIST).

Journal of Clinical Oncology ◽

10.1200/jco.2013.31.15_suppl.10509 ◽

2013 ◽

Vol 31 (15_suppl) ◽

pp. 10509-10509 ◽

Cited By ~ 1

Author(s):

Michael C. Heinrich ◽

Jonathan A. Fletcher ◽

Rana Anjum ◽

Cesar Serrano-Garcia ◽

Sadanand Vodala ◽

...

Keyword(s):

Cell Lines ◽

Gastrointestinal Stromal Tumors ◽

Kinase Activity ◽

Binding Pocket ◽

Drug Resistant ◽

Line Treatment ◽

Resistance Mutations ◽

Secondary Resistance ◽

Stromal Tumors ◽

Exon 11

10509 Background: Ponatinib (PO) is a multi-targeted tyrosine kinase inhibitor with potent pan-BCR-ABL activity that has recently been approved for treatment of CML and Ph+ ALL. PO also inhibits the kinase activity of KIT. Approximately 80% of gastrointestinal stromal tumors (GIST) contain primary activating KIT mutations, the majority of which cluster in exon 11. Imatinib (IM) is approved for the 1st line treatment of GIST; however, patients frequently relapse due to the acquisition of secondary resistance mutations located in either the KIT ATP-binding pocket or the activation (A) loop. Sunitinib (SU) is approved for 2nd line treatment of GIST but does not effectively inhibit A-loop mutants. Here we explored the activity of PO against major primary and secondary KIT mutants found in GIST. Methods: The drug sensitivity of KIT mutants was determined using engineered Ba/F3 cells harboring mutant forms of KIT exon 11 with or without ATP binding pocket or A-loop mutations. The abilities of PO, IM, SU, and regorafenib (RE) to inhibit viability and/or KIT kinase activity were compared using this system as well as an isogenic CHO cell system. We also profiled these same drugs using a panel of GIST cell lines, including cell lines with IM-resistant secondary KIT mutations. Results: In all in vitro systems, PO potently inhibited KIT exon 11 mutant kinases, with an IC50 of < 30 nM. PO also potently inhibited a range of secondary KIT mutants, including multiple A-loop mutant kinases. PO induced substantial tumor regression in Ba/F3 tumor models expressing a KIT exon 11 mutant with or without an A-loop mutation (D816H). Using GIST cell lines, PO inhibited the viability of those harboring primary KIT exon 11 and secondary resistance mutations more effectively than IM, SU, and RE. Importantly, in patients dosed once daily with 45 mg ponatinib, plasma concentrations achieved are predicted to lead to inhibition of all KIT mutants tested with the possible exception of V654A. Conclusions: PO potently inhibits the majority of clinically relevant KIT mutant kinases and has a broader spectrum of activity compared to IM, SU, or RE. Based on these data, a phase 2 study of PO in drug-resistant GIST is being initiated.

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