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Most gastrointestinal stromal tumors (GIST) harbor mutated receptor tyrosine kinase (RTK) KIT/PDGFRA, which provides an attractive therapeutic target. However, a majority of GISTs ultimately develop resistance to KIT/PDGFRA inhibitor imatinib, multiple therapeutic targets will be identified as a reasonable strategy in imatinib-resistant GISTs. Biological mechanisms of non-RTK activated CDC42 associated kinase 1 (ACK1) are still unclear, which has been found to be activated in GISTs. In the current report, ACK1 overexpression is demonstrated in GIST cell lines and biopsies. RNA-seq analysis and immunoblotting show that ACK1 expression is dependent on imatinib treatment time in GIST-T1 cell line. The colocalization/complex of KIT and ACK1 in GIST cells are observed, and ACK1 activation is in a partially KIT and CDC42 dependent manner. Treatment with a specific ACK1 inhibitor AIM-100 or ACK1 siRNA, mildly suppresses cell viability, but markedly inhibits cell migration in imatinib sensitive and in imatinib resistant GIST cell lines, which is associated with inactivation of PI3K/AKT/mTOR and RAF/MAPK signaling pathways, and inhibition of epithelial-mesenchymal transition, evidencing upregulation of E-cadherin and downregulation of ZEB1, N-cadherin, vimentin, snail, and/or β-catenin after treatment with AIM-100 or ACK1/CDC42 shRNAs. Combination inhibition of ACK1 and KIT results in additive effects of anti-proliferation and pro-apoptosis as well as cell cycle arrest, and inhibition of invasiveness and migration in vitro and in vivo, compared to either intervention alone through dephosphorylation of KIT downstream intermediates (AKT, S6, and MAPK). Our data suggest that co-targeting of ACK1 and KIT might be a novel therapeutic strategy in imatinib-resistant GIST., Competing Interests: Declaration of competing interest The authors declare that they have no conflicts of interest associated with the contents of this article., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Chronic myeloid leukemia (CML) is a hematological disease, and imatinib (IM) resistance represents a major problem for its clinical treatment. In the present study, the role of tribbles pseudokinase 2 (TRIB2) in IM resistance of CML and the possible mechanism were investigated. It was found that TRIB2 was highly expressed in IM‑resistant patients with CML through the Oncomine database and this conclusion was confirmed using reverse transcription‑quantitative PCR and western blot experiments. Knockdown of TRIB2 was found to increase the drug sensitivity of KG cells to IM using Cell‑Counting Kit‑8 (CCK‑8) assays, and the low‑expression TRIB2 mice were further found to be more sensitive to the IM and have a higher survival rate in leukemia model mice. Moreover, using western blot and luciferase experiments, it was found that TRIB2 could regulate c‑Fos through the ERK signaling pathway, and c‑Fos suppressed the transcriptional activity and the expression of miR‑33a‑5p. Further investigation identified that the binding site for c‑Fos to function on miR‑33a‑5p was the ‑958‑965 region. Finally, CCK‑8 assays and western blot experiments demonstrated that miR‑33a‑5p could inhibit the proliferation of KG cells and reduce IM resistance by suppressing the expression of HMGA2. In conclusion, it was demonstrated that TRIB2 regulates miR‑33a‑5p to reverse IM resistance in CML, which may help identify novel targets and therapeutic strategies for the clinical treatment of IM resistance.

Purpose: Imatinib is the standard treatment for unresectable and metastatic GIST. In the late stages, patients undergoing imatinib show drug resistance. Surgical intervention has been occasionally performed for resistant lesions. However, the clinical significance of such intervention remains unclear., Methods: Between 2006 and 2015, 37 patients were diagnosed with imatinib-resistant GISTs. We performed surgical intervention only for localized resistant lesions. We retrospectively investigated the background characteristics, data on surgical intervention and subsequent treatment, progression-free survival (PFS), and overall survival (OS)., Results: Eighteen patients diagnosed with localized resistance received surgical intervention (S-group) and 19 patients diagnosed with generalized resistance were received other TKIs (M-group). In S-group, no serious complications occurred, and all patients restarted imatinib after resection. The median PFS was 14.5 months. Five patients underwent surgical intervention multiple times followed by the continuation of imatinib, and the median duration of imatinib continuation was 22.2 months. Second-line TKIs were administered to 93% of the patients and the dose-intensity and outcome were similar in both groups. The median OS was 47.2 months after surgery., Conclusions: Surgical intervention could be performed safely and therefore could be followed by the continuation of TKI therapy. Surgical intervention based on the appropriate criteria of resistance might thus be useful for imatinib-resistant GISTs., (© 2021. Springer Nature Singapore Pte Ltd.)

Imatinib-resistance is a significant concern for Bcr-Abl-positive chronic myelogenous leukemia (CML) treatment. Emodin, the predominant compound of traditional medicine rhubarb, was reported to inhibit the multidrug resistance by downregulating P-glycoprotein of K562/ADM cells with overexpression of P-glycoprotein in our previous studies. In the present study, we found that emodin can be a potential inhibitor for the imatinib-resistance in K562/G01 cells which are the imatinib-resistant subcellular line of human chronic myelogenous leukemia cells with overexpression of breakpoint cluster region-abelson (Bcr-Abl) oncoprotein. Emodin greatly enhanced cell sensitivity to imatinib, suppressed resistant cell proliferation and increased potentiated apoptosis induced by imatinib in K562/G01 cells. After treatment of emodin and imatinib together, the levels of p-Bcr-Abl and Bcr-Abl were significantly downregulated. Moreover, Bcr-Abl important downstream target, STAT5 and its phosphorylation were affected. Furthermore, the expression of Bcr-Abl and signal transducers and activators of transcription 5 (STAT5) related molecules, including c-MYC, MCL-1, poly(ADP-ribose)polymerase (PARP), Bcl-2 and caspase-3, were changed. Emodin also decreased Src expression and its phosphorylation. More importantly, emodin simultaneously targeted both the ATP-binding and allosteric sites on Bcr-Abl by molecular docking, with higher affinity with the myristoyl-binding site for enhanced Bcr-Abl kinase inhibition. Overall, these data indicated emodin might be an effective therapeutic agent for inhibiting resistance to imatinib in CML treatment.

ZNF224 is a KRAB-zinc finger transcription factor that exerts a key tumor suppressive role in chronic myelogenous leukemia. In this study, we identify the receptor tyrosine kinase Axl as a novel target of ZNF224 transcriptional repression activity. Axl overexpression is found in many types of cancer and is frequently associated with drug resistance. Interestingly, we also found that sensitivity to imatinib can be partly restored in imatinib-resistant chronic myelogenous leukemia cells by ZNF224 overexpression and the resulting suppression of Axl expression. These results, in accordance with our previous findings, support the role of ZNF224 in imatinib responsiveness and shed new insights into potential therapeutic use of ZNF224 in imatinib-resistant chronic myelogenous leukemia., (Copyright © 2018 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.)

The Sonic hedgehog (Shh) signaling pathway may be interrelated with other signaling pathways, such as the phosphoinositide 3-kinase (PI3K) and mitogen-activated protein kinase (MAPK) pathways in gastrointestinal stromal tumor (GIST). The present study investigated the interaction among Shh, PI3K and MAPK signaling pathways in GIST cells. The expression of PI3K, MAPK and Shh signaling pathways in GIST-H1 cells were upregulated by endothelial growth factor (EGF) and recombinant Shh (N-shh) stimulation, and were downregulated by specific inhibitors of each signaling pathway. The proliferation rate of GIST-H1 cells were significantly increased under EGF or N-shh treatment (P<0.01). In addition, this effect was partially prevented by the pretreatment of the inhibitors of these signaling pathways. In summary, a cross regulation exists among the Shh, PI3K and MAPK signaling pathways in GIST-H-1 cells. The combined use of the inhibitors of these signaling pathways is a potentially novel option for GIST targeted therapy.

Imatinib (IMT), a specific tyrosine kinase inhibitor (TKI), has drastically changed the treatment strategy for Ph+ ALL (Philadelphia chromosome-positive acute lymphoblastic leukemia). However, TKI resistance remains a serious problem for patient prognosis. Here, a Ph+ ALL cell line NphA2 and the IMT-resistant subline NphA2/STIR were analyzed to identify a potential novel treatment strategy. We also examined other Ph+ ALL cells, MR87 and its IMT-resistant subline, MR87/STIR. IMT induced apoptosis of NphA2 and MR87 but had no effect on resistant sublines. Increased phosphorylated ERK and BCL2, but not BCL-XL, were observed in NphA2/STIR compared with NphA2. NphA2/STIR but not NphA2 was moderately sensitive to U0126, an ERK inhibitor. Interestingly, SP600125, a JNK inhibitor, was potent in cell growth inhibition and apoptosis induction of both parental and IMT-resistant NphA2 and MR87 cells. Moreover, NphA2 and MR87 and their IMT-resistant sublines were sensitive to ABT-199, a specific BCL2 inhibitor. The combination of SP600125 and ABT-199 synergistically suppressed both parental and IMT-resistant cells, including one with T315I mutation, suggesting that Ph+ ALL exhibits high sensitivity to ABT-199 and SP600125 regardless of TKI resistance. This combination might be a possible therapeutic strategy for Ph+ ALL in the future.

Quiescent leukemia stem cells (LSCs) that are insensitive to BCR-ABL tyrosine kinase inhibitors confer resistance to imatinib in chronic myelogenous leukemia (CML). Identifying proteins to regulate survival and stemness of LSCs is urgently needed. Although histone deacetylase inhibitors (HDACis) can eliminate quiescent LSCs in CML, little is known about the underlying mechanism that HDACis kill LSCs. By fishing with a biotin-labeled probe, we identified that HDACi JSL-1 bound to the protein γ-catenin. γ-Catenin expression was higher in LSCs from CML patients than normal hematopoietic stem cells. Silencing γ-catenin in human CML CD34(+) bone-marrow (BM) cells sufficiently eliminated LSCs, which suggests that γ-catenin is required for survival of CML LSCs. Pharmacological inhibition of γ-catenin thwarted survival and self-renewal of human CML CD34(+) cells in vitro, and of murine LSCs in BCR-ABL-driven CML mice. γ-Catenin inhibition reduced long-term engraftment of human CML CD34(+) cells in NOD.Cg-Prkdc (scid) II2rg (tm1Sug)/JicCrl (NOG) mice. Silencing γ-catenin by shRNA in human primary CD34(+) cells did not alter β-catenin, implying a β-catenin-independent role of γ-catenin in survival and self-renewal of CML LSCs. Taken together, our findings validate that γ-catenin may be a novel therapeutic target of LSCs, and suppression of γ-catenin by HDACi may explain elimination of CML LSCs., Competing Interests: Authors have no conflicts of interest.

Chronic myeloid leukaemia (CML) is a myeloproliferative disorder promoted by the constitutive tyrosine kinase activity of Bcr-Abl oncoprotein. Although treatment with the Bcr-Abl-inhibitor imatinib represents the first-line therapy against CML, almost 20-30% of patients develop chemotherapeutic resistance and require alternative therapy. Here we show that a strong hyper-phosphorylation/activation of ERK1/2, Akt Ser473, and 40S ribosomal protein S6 (rpS6) is detectable in imatinib-resistant KCL22 and K562 CML cells as compared to the -sensitive cell variants. In imatinib-resistant CML cells, high concentration of imatinib is required to strongly inhibit Bcr-Abl, ERK1/2 and Akt Ser473 phosphorylation, but under these conditions the phosphorylation of rpS6, a common downstream effector of MEK/ERK1/2 and PI3K/Akt/mTOR pathways is only slightly reduced. By contrast, down-regulation of the protein kinase CK2 by the inhibitor CX-5011 or by silencing the CK2 subunits does not affect the activation state of MEK/ERK1/2 or PI3K/Akt/mTOR signalling, but causes a drop in rpS6 phosphorylation in parallel with reduced protein synthesis. CK2-inhibition by CX-5011 induces cell death by apoptosis and acts synergistically with imatinib or the MEK-inhibitor U0126 in reducing the viability of imatinib-resistant CML cells. The ternary mixture containing CX-5011, imatinib and U0126 represents the most effective synergistic combination to counteract CML cell viability. These results disclose a novel CK2-mediated mechanism of acquired imatinib-resistance resulting in hyper-phosphorylation of rpS6. We suggest that co-targeting CK2 and MEK protein kinases is a promising strategy to restore responsiveness of resistant CML cells to imatinib.

After an initial benefit from tyrosine kinase inhibitors (TKI), most gastrointestinal stromal tumors (GISTs) eventually develop disease progression or secondary resistance. An altered tumor (immune)phenotype with anaplasia and morphological changes secondary to therapy have occasionally been described in the literature. We present a 52-year old patient, diagnosed with high risk, spindle-cell, GIST (CD117 positive, Pankeratin negative) in 2003, showing a c-Kit exon 11 mutation. After TKI therapy, he developed drug resistance and disease progression. Pathological assessment of the last surgical specimen showed a pure epithelioid/clear cell histology, without evidence of cellular anaplasia. Tumor cells were CD117 positive, DOG1 positive but also E-cadherin positive and Pankeratin positive, whereas molecular analysis confirmed the presence of the c-Kit exon 11 mutation, with no additional mutations. We describe an unusual case of GIST showing peculiar (immuno)phenotypic changes under therapy, different from the vast majority of therapy-driven changes, which include marked cellular pleomorphisms and KIT immunonegativity. Possible molecular explanations to understand these phenomena and a brief review of the literature are also addressed., (Copyright © 2015 Elsevier GmbH. All rights reserved.)

Gain-of-function mutations of receptor tyrosine kinase KIT play a critical role in the pathogenesis of systemic mastocytosis (SM) and gastrointestinal stromal tumors. D816V KIT mutation, found in ∼80% of SM, is resistant to the currently available tyrosine kinase inhibitors (TKIs) (e.g. imatinib mesylate). Therefore, development of promising TKIs for the treatment of D816V KIT mutation is still urgently needed. We synthesized thiazole amine compounds and chose one representative designated 126332 to investigate its effect on human mast cells expressing KIT mutations. We found 126332 inhibited the phosphorylation of KIT and its downstream signaling molecules Stat3 and Stat5. 126332 inhibited the proliferation of D816V KIT expressing cells. 126332 induced apoptosis and downregulated levels of Mcl-1 and survivin. Furthermore, 126332 inhibited the tyrosine phosphorylation of β-catenin, inhibited β-catenin-mediated transcription and DNA binding of TCF. Moreover, 126332 also exhibited in vivo antineoplastic activity against cells harboring D816V mutation. Our findings suggest thiazole amine compounds may be promising agents for the treatment of diseases caused by KIT mutation., (Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.)

Chronic myeloid leukaemia (CML) is driven by the fusion protein Bcr-Abl, a constitutively active tyrosine kinase playing a crucial role in initiation and maintenance of CML phenotype. Despite the great efficacy of the Bcr-Abl-specific inhibitor imatinib, resistance to this drug is recognized as a major problem in CML treatment. We found that in LAMA84 cells, characterized by imatinib-resistance caused by BCR-ABL1 gene amplification, the pro-survival protein kinase CK2 is up-regulated as compared to the sensitive cells. CK2 exhibits a higher protein-level and a parallel enhancement of catalytic activity. Consistently, CK2-catalysed phosphorylation of Akt-Ser129 is increased. CK2 co-localizes with Bcr-Abl in the cytoplasmic fraction as judged by subcellular fractionation and fluorescence immunolocalization. CK2 and Bcr-Abl are members of the same multi-protein complex(es) in imatinib-resistant cells as demonstrated by co-immunoprecipitation and co-sedimentation in glycerol gradients. Cell treatment with CX-4945, a CK2 inhibitor currently in clinical trials, counteracts CK2/Bcr-Abl interaction and causes cell death by apoptosis. Interestingly, combination of CX-4945 with imatinib displays a synergistic effect in reducing cell viability. Consistently, knockdown of CK2α expression by siRNA restores the sensitivity of resistant LAMA84 cells to low imatinib concentrations. Remarkably, the CK2/Bcr-Abl interaction and the sensitization towards imatinib obtained by CK2-inhibition in LAMA84 is observable also in other imatinib-resistant CML cell lines. These results demonstrate that CK2 contributes to strengthen the imatinib-resistance phenotype of CML cells conferring survival advantage against imatinib. We suggest that CK2 inhibition might be a promising tool for combined strategies in CML therapy., (Copyright © 2013 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.)

Introduction: Chronic myelogenous leukemia (CML) is a progressive and often fatal hematopoietic neoplasm. The Bcr-Abl tyrosine kinase inhibitor imatinib mesylate represented a major therapeutic advance over conventional CML therapy, with more than 90% of patients obtaining complete hematologic response, and 70%-80% of patients achieving a complete cytogenetic response. Despite the high efficacy of imatinib, a minority of patients in chronic phase CML and more patients in advanced phases are resistant to imatinib, or develop resistance during treatment. This is attributed, in 40% to 50% of cases, to the development of mutations in the Bcr-Abl tyrosine kinase domain that impair imatinib binding. Attempts to circumvent resistance led to the discovery of nilotinib (Tasigna), a novel, potent and selective oral Bcr-Abl kinase inhibitor., Aims: To review the evidence for the use of nilotinib in the management of CML., Evidence Review: Preclinical and clinical investigations demonstrate that nilotinib effectively overcomes imatinib resistance, and has further improved the treatment of CML., Place in Therapy: Nilotinib is currently indicated for patients with CML in chronic and accelerated phases following imatinib failure. Randomized studies are ongoing to assess the efficacy of nilotinib in patients with newly diagnosed CML.

The development of imatinib has resulted in sustained hematologic and cytogenetic remissions in all phases of chronic myeloid leukemia (CML). Despite the high efficacy, relapses have been observed and are much more prevalent in patients with advanced disease. The most common mechanism of acquired resistance has been traced to Bcr-Abl kinase domain mutations. Several strategies have been developed to overcome the problem of imatinib resistance, including imatinib dose escalation, novel targeted agents and combination treatments. A second generation of tyrosine kinase inhibitors was developed, which displays increased potency towards Bcr-Abl and is able to target the majority of CML mutant clones. Nilotinib (Tasigna, AMN107, Novartis) is a close analog of imatinib with approximately 20-fold higher potency for BCR-ABL kinase inhibition. Preclinical and clinical investigations demonstrate that nilotinib effectively overcomes imatinib resistance, and has induced high rates of hematologic and cytogenetic responses in CML post imatinib failure, with a good tolerance. Nilotinib has been approved for CML patients in chronic and accelerated phases, post imatinib failure.