Mutations In Tyrosine Kinase Domain Research Articles

Mutation Patterns Identify Adult Patients with De Novo Acute Myeloid Leukemia (AML) Aged 60 Years or Older Who Respond Favorably to Standard Chemotherapy: an Analysis of Alliance Studies

Abstract Background: Despite changes in treatment regimens, the patient's age (and the corresponding treatment intensity) remains one of the most important prognostic factors in AML. Thus far, only 5-15% of AML patients aged ≥60 years (y) are cured with chemotherapy. Identification of patients who are more (or less) likely to respond to standard cytarabine/daunorubicin-based chemotherapy might help to enable early risk stratification toward alternative treatment regimens. Aims: To provide a comprehensive molecular profile of a large cohort of adults with de novo acute myeloid leukemia (AML) aged ≥60 y, and to determine if specific molecular and/or cytogenetic features could identify patients who are more likely to respond to standard chemotherapy. Methods: We used a next-generation sequencing panel of 80 cancer- and/or leukemia-associated genes to profile molecularly 423 patients aged ≥60 y with de novo AML and available cytogenetic data similarly treated on Cancer and Leukemia Group B/Alliance for Clinical Trials in Oncology. Results: Overall, we detected 1377 mutations, with a median of 3 mutations per patient (range: 0-8). The outcome of the whole patient cohort was poor, with complete remission (CR) rate of 55% and disease-free (DFS) and overall survival (OS) 3y-rates of 14%. In multivariable analyses (MVA), mutated NPM1 was the only factor positively associated with achieving CR (P Summary: We identified combinations of mutations whose presence is associated with favorable outcome in older AML patients treated with standard chemotherapy. Patients with intermediate and adverse molecular features should be considered for alternative treatment regimens. Download : Download high-res image (186KB) Download : Download full-size image Figure 1 . Disclosures Stone: Otsuka: Membership on an entity's Board of Directors or advisory committees; Astellas: Membership on an entity's Board of Directors or advisory committees; Janssen: Membership on an entity's Board of Directors or advisory committees; Ono: Membership on an entity's Board of Directors or advisory committees; Orsenix: Membership on an entity's Board of Directors or advisory committees; Cornerstone: Membership on an entity's Board of Directors or advisory committees; Agios: Membership on an entity's Board of Directors or advisory committees, Research Funding; Amgen: Membership on an entity's Board of Directors or advisory committees; Argenix: Other: DSMB; Arog: Membership on an entity's Board of Directors or advisory committees, Research Funding; Novartis: Membership on an entity's Board of Directors or advisory committees, Research Funding; Merck: Membership on an entity's Board of Directors or advisory committees; Pfizer: Membership on an entity's Board of Directors or advisory committees; Roche: Membership on an entity's Board of Directors or advisory committees; Actinium: Membership on an entity's Board of Directors or advisory committees; Karyopharm: Membership on an entity's Board of Directors or advisory committees; Fujifilm: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees, Other: DSMB; Jazz: Membership on an entity's Board of Directors or advisory committees; Abbvie: Membership on an entity's Board of Directors or advisory committees; Seattle genetics: Membership on an entity's Board of Directors or advisory committees; Sumitomo: Membership on an entity's Board of Directors or advisory committees. Kolitz: Gilead, Magellan, Novartis, Pharmacyclics, and Seattle Genetics: Consultancy; Gilead, Magellan, and Novartis: Honoraria; Celgene, Jazz: Equity Ownership; Boehringer Ingelheim, Cantex, Erytech, and Millennium: Research Funding; Gilead, Novartis, and Seattle Genetics: Other: Travel Support. Powell: Rafael Pharmaceuticals: Consultancy, Membership on an entity's Board of Directors or advisory committees. Byrd: Janssen: Research Funding; Genentech: Research Funding; Acerta Pharma: Research Funding; Pharmacyclics: Research Funding; The Ohio State University: Patents & Royalties: OSU-2S.

De Novo Mutation in Non-Tyrosine Kinase Domain of ROS1 as a Potential Predictor of Immune Checkpoint Inhibitors in Melanoma.

PurposeDespite the success of targeted therapy in c-ros oncogene 1 (ROS1)-rearranged cancers, especially non-small cell lung cancer (NSCLC), the clinical significance of ROS1 de novo mutation has not yet been understood. We sought to elucidate the predictive effect of ROS1 mutation for immune checkpoint inhibitor (ICI) therapy in melanoma.MethodsThe Cancer Genome Atlas [TCGA (n = 10967)] and Memorial Sloan Kettering Cancer Center [MSK (n = 10,945)] datasets, as well as two clinical cohorts of melanoma received ICI [CA209-038 (n = 73) and MEL-IPI (n = 110)], were included to explore the prevalence, prognostic effect, and immunotherapeutic predictive effect of ROS1 mutation in melanoma. Overall survival (OS) was defined as the primary outcome.ResultsOverall, melanoma accounted for the highest proportion of ROS1 mutation (~20%) which made up the majority (~95%) of the ROS1-alterated cases. Remarkably, ROS1 mutation yielded longer OS from ICI than the wild-type counterpart in the MSK melanoma population [hazard ratio (HR) 0.47, 95% confidence interval (CI) 0.30–0.74], and two external melanoma cohorts (CA209-038: HR 0.42, 95% CI 0.20–0.89; MEL-IPI: HR 0.55, 95% CI 0.34–0.91), without affecting the prognosis of patients. Elevated tumor mutational burden and enrichment of DNA damage repair was observed in ROS1 mutated patients, providing an explanation for the favorable responses to ICI therapy. Precisely, ROS1 mutation in non-protein tyrosine kinase (PTK) domain but not PTK mutation was responsible for the immunotherapy-specific responses of the ROS1 mutated patients in melanoma.ConclusionsCollectively, ROS1 mutation, specifically the non-PTK mutation, is a potential predictor of ICI therapy in melanoma, which is distinct from the well-established role of ROS1 rearrangement for targeted therapy in NSCLC.

Inhibition of translation initiation factor eIF4a inactivates heat shock factor 1 (HSF1) and exerts anti-leukemia activity in AML.

Eukaryotic initiation factor 4A (eIF4A), the enzymatic core of the eIF4F complex essential for translation initiation, plays a key role in the oncogenic reprogramming of protein synthesis, and thus is a putative therapeutic target in cancer. As important component of its anticancer activity, inhibition of translation initiation can alleviate oncogenic activation of HSF1, a stress-inducible transcription factor that enables cancer cell growth and survival. Here, we show that primary acute myeloid leukemia (AML) cells exhibit the highest transcript levels of eIF4A1 compared to other cancer types. eIF4A inhibition by the potent and specific compound rohinitib (RHT) inactivated HSF1 in these cells, and exerted pronounced in vitro and in vivo anti-leukemia effects against progenitor and leukemia-initiating cells, especially those with FLT3-internal tandem duplication (ITD). In addition to its own anti-leukemic activity, genetic knockdown of HSF1 also sensitized FLT3-mutant AML cells to clinical FLT3 inhibitors, and this synergy was conserved in FLT3 double-mutant cells carrying both ITD and tyrosine kinase domain mutations. Consistently, the combination of RHT and FLT3 inhibitors was highly synergistic in primary FLT3-mutated AML cells. Our results provide a novel therapeutic rationale for co-targeting eIF4A and FLT3 to address the clinical challenge of treating FLT3-mutant AML.

Role of Antioxidant Gene Polymorphisms in Risk and Prognosis of Chronic Myeloid Leukemia

Introduction: We aimed to investigate the possible role of antioxidant enzyme polymorphisms CAT -21A/T (rs7943316), CAT -262C/T (rs1001179), GPX1 -198C/T (rs1050450), MPO -463G/A (rs2333227), GSTM1 (rs366631) & GSTT1 (rs17856199) with susceptibility to chronic myeloid leukemia (CML) and their association with tyrosine kinase inhibitor (TKI, imatinib) response.Methods: Six single nucleotide polymorphisms (SNPs) in antioxidant enzyme genes were genotyped in a total of 325 samples, of which 125 were from CML patients and 200 from healthy controls. The SNPs were correlated with various confounding variables lke BCR-ABL1 levels and tyrosine kinase domain mutation status in CML patients.Results: Genotyping results revealed statistically significant associations with CAT -21A/T (p=0.037) and GPX1 -198C/T (p=<0.0001) polymorphisms with risk of CML. No associations were observed between CAT -262C/T, MPO -463G/A, GSTM1 & GSTT1 polymorphisms and CML. The CAT -21A/T polymorphism conferred 2.95 folds increased risk of CML under co-dominant model (p=0.024) and 2.51 folds risk under dominant models (p=0.05). In addition, the haplotypes of CAT -21A/T and -262C/T polymorphisms, ATCC and ATCT conferred higher incidence of CML risk by 2.67 times (p=0.05) and 2.99 times (p=0.045). The GPX1 -198C/T polymorphism conferred significantly increased risk of CML under co-dominant model [CC vs CT (p=<0.0001), CC vs TT (p=<0.0001)] and dominant models [CC vs CT+TT (p=<0.0001)]. The heterozygous GPX1 CT genotype frequency significantly elevated in poor molecular responders (p=0.005) and TKD mutation carriers (p=0.114) as compared to respective groups.Conclusions: Our results suggest that the reduced activity of antioxidant enzymes caused by the CAT -21A/T and GPX1-198C/T polymorphisms might contribute to increased risk of CML. In addition, the GPX1-198C/T polymorphism was associated with poor molecular response and acquired TKD mutations. Hence, the present study indicates that defective antioxidant defense system might have a strong influence on CML susceptibility and TKI (imatinib) response through oxidative stress.

Non‐small‐cell lung cancer with ERBB2 mutation in non‐tyrosine kinase domain benefits from pyrotinib: A case report

Tyrosine kinase domain (TKD) mutation and particularly exon 20 insertion mutations of erb‐b2 receptor tyrosine kinase 2 (ERBB2/HER2) have been extensively reported in non‐small cell lung cancer (NSCLC). Nevertheless, the clinical significance of non‐TKD mutations remains unknown. To date, no clinical trials have revealed that tyrosine kinase inhibitors are effective in NSCLC patients with non‐TKD ERBB2 mutations. Here we report a patient with advanced lung adenocarcinoma harboring non‐TKD mutation of ERBB2, S335C, without other actionable alterations benefited from pyrotinib. After first‐line treatment of pyrotinib monotherapy, a pan‐HER inhibitor, the patient achieved a durable partial response with good tolerance. This case powerfully illustrates that pyrotinib might be a promising first‐line treatment strategy for NSCLC patients with non‐TKD mutation of ERBB2.

81030 Utilizing a synergistic drug combination to target relapsed/refractory FLT3 mutant AML

ABSTRACT IMPACT: This work will help to understand a novel therapeutic approach to a common type of acute myeloid leukemia. OBJECTIVES/GOALS: FMS-like tyrosine kinase 3 (or FLT3) mutations occur in ˜30% of acute myeloid leukemia (AML) cases. FLT3 tyrosine kinase domain (TKD) mutations are particularly important in relapsed/refractory FLT3 mutant AML, which portends poor prognosis. This study describes a therapeutic approach to overcoming resistance conferred by FLT3-TKD mutations. METHODS/STUDY POPULATION: To understand the efficacy of a novel type 1 FLT3 inhibitor (NCGC1481), as a monotherapy and combination therapy, several assays were utilized to interrogate functionality of these therapies. Cell lines and patient samples containing aspartate 835 to tyrosine mutations (D835Y, the most common TKD alteration) and phenylalanine 691 to leucine (F691L) were utilized to examine the effects of NCGC1481 with and without other targeted therapies like MEK inhibitors. Specifically, assays measuring viability, cell death using flow cytometry, in vitro clonogenicity, cellular signaling, and xenograft survival were examined in these FLT3-TKD AML models. Synergy was also measured using well described methods, which also allowed for appropriate dose finding for drug combination experiments. RESULTS/ANTICIPATED RESULTS: Our novel type 1 FLT3 inhibitor (NCGC1481) was particularly effective in the most common FLT3 TKD mutant, D835Y. NCGC1481 reduced viability and cell signaling, while also inducing cell death and prolonging xenograft survival in the FLT3-D835Y model system. In contrast, clinically approved FLT3 inhibitors were less effective at suppressing AML cells expressing FLT3-D835Y. In the case of FLT3-F691L, most of the FLT3 inhibitors tested, including NCGC1481, suppressed canonical FLT3 signaling, but did not significantly reduce viability or leukemic clonogenicity. However, when NCGC1481 was combined with other targeted agents like MEK inhibitors, at synergistic doses, eradication of the FLT3-F691L AML clone was substantially increased. DISCUSSION/SIGNIFICANCE OF FINDINGS: In AML, response to FLT3 inhibitor therapy is often short-lived, with resistance sometimes occurring via FLT3-TKD mutations. Given the dismal prognosis of relapsed FLT3 mutant AML, novel therapies are necessary. This study describes efficacy of a novel FLT3 inhibitor, along with its synergistic activity when combined with other targeted agents.

Evaluation of cytogenetic response in CML patients with variant Philadelphia translocation.

Molecular mechanism of translocation and outcome in variant chronic myeloid leukaemia (vCML) has been a topic of debate. While several cytogenetic studies suggest a low response to Imatinib Mesylate, others demonstrate a similar disease course in both classical and vCML. Besides, many studies comprehensively also link tyrosine kinase domain (TKD) mutations with aggressive clinical outcome. Thus, we aim to study the molecular mechanism of translocation, identify the third partner chromosomes and comment on the disease course and clinical outcome. We cytogenetically characterised 25 vCML cases to determine the third partner chromosome, mechanism of translocation and prognostic outcome. We also compared vCML cases with and without TKD mutation to most appropriately outline the clinical consequence and ascertain the potent cause of unresponsiveness to treatment. Third partner chromosome in variant translocation was defined by conventional and molecular cytogenetics. Although in our study most cases showed inadequate clinical response attributable to TKD mutation rather than variant translocation, we observed an inferior outcome in cases involving chromosome 5 as the third partner. Thus, we conclude that characterising and reporting new cases of variant translocations, involving various different chromosomes as third partner (with different breakpoints) by cytogenetics, will lead to a better understanding of the disease. To the best of our knowledge, this kind of delineate study has not been applied to precisely comment on the prospects of cytogenetically characterised vCML.

Genomic Analysis of FLT3 Mutations in a Comprehensive NGS Multicenter Study of AML: HM-Screen-Japan 01

Genomic Analysis of FLT3 Mutations in a Comprehensive NGS Multicenter Study of AML: HM-Screen-Japan 01

Clinical Benefit and Tolerability of Crenolanib in Children with Relapsed Acute Myeloid Leukemia Harboring Treatment Resistant FLT3 ITD and Variant FLT3 TKD Mutations Treated on Compassionate Access

Background: FLT3-mutant pediatric AML represents a biological and clinically diverse disease and is associated with a poor outcome. Sorafenib has limited activity against tyrosine kinase domain mutations. Crenolanib is a pan-FLT3 inhibitor, which, at the time of this compassionate use program, had already been safely administered to over 55 children (age 2-18) with diffuse intrinsic pontine glioma as well as high grade gliomas. Crenolanib at a dose of 66mg/m2 TID has been well tolerated in pediatric patients. Methods: We here report our experience with five consecutive children (ages 4-12y) who received crenolanib for FLT3 mutant AML on a compassionate basis between April 2017 to October 2019. All patients treated had IRB/local ethics approval prior to treatment and all patients' guardians signed informed consent forms. Results: Patient Characteristics: Of the five patients, three were refractory to induction therapy and required salvage therapy to get into remission. All five patients were then able to undergo HSCT, but all subsequently relapsed. Two patients were successfully salvaged and underwent a second HSCT. Only two pts received sorafenib. By the time of the compassionate use request, all children had exhausted all standard and experimental therapies (3-9 prior therapies, Table 1). Two patients had FLT3-ITDs and three had FLT3 kinase domain mutations (A848P in one, D835H in another and both D835H and D835E in trans in third). Co-occurring events included KMT2A fusion/mutations in three pts as follows: i) one pt with KMT2A-rearranged infant AML, ii) one with treatment-related AML with KMT2A fusion and t(9;11) and iii) a third patient with KMT2A fusion along with p53 mutation. Another patient with biphenotypic AML had a co-occurring NOTCH1 mutation with 9;14 translocation. Three patients had complex karyotypes and a number of translocations were identified including t(9;11), t(3;5), t(1;16), and t(9;14) (Table 1). In addition to bone-marrow disease and circulating blasts, all patients had extramedullary AML. Three patients had CNS leukemia. Three patients had non-CNS extramedullary AML (submandibular, testicular, liver and spleen). Treatment: Crenolanib was given with curative intent to three patients, one in combination with Vyxeos (liposomal cytarabine/daunorubicin), one with high-dose cytarabine, and one as maintenance therapy after her second HSCT. Two patients received crenolanib as palliation for rapidly progressing AML. Tolerability: All children tolerated crenolanib well. Reasons for crenolanib discontinuation were bridge to HSCT in two patients and completion of 12 months of maintenance in another. One patient stopped crenolanib early as he developed fungal pneumonia and one patient stopped due to lack of benefit. Only one patient required dose reduction due to grade 2 transaminitis. There were no cardiac toxicities, pericardial effusion, fluid retention or weight gain. Response: Four of five patients reported clinical benefit with crenolanib. Interestingly, the child with KMT2A-rearranged infant AML achieved a molecular CR after salvage therapy with Vyxeos plus crenolanib. This child was successfully bridged to a second HSCT and remains in remission one year after the start of his compassionate use crenolanib. Another patient with KMT2A fusion, p53 mutation and CNS AML achieved a CR with crenolanib + sorafenib, was successfully bridged to 2nd HSCT and received one year of post-transplant crenolanib maintenance. She remains in remission 3.5 years after initiation of crenolanib. The third patient with KMT2A fusion was successfully bridged to allo HSCT but relapsed 4 months after HSCT (she did not receive maintenance). The patient with bi-phenotypic AML (and D835H mutation) had a quick reduction in circulating blasts (within 24 h) but died of fungal sepsis. The fifth patient received reduced doses of crenolanib due to transaminitis and did not have a clinical benefit. Conclusion: This series of five children with multiply relapsed FLT3-mutant AML shows that treatment with full doses of crenolanib can be safely combined with salvage chemotherapy. Rapid remissions could be obtained even in patients with co-occurring KMT2A, 3q, and p53 mutations. Crenolanib, which is novel type-I pan FLT3 inhibitor, was able to inhibit variant FLT3 mutations (D835H, D835E and D848P). Crenolanib can be safely combined with ITT (2 of these children remain alive 1-3.5 years). Figure Disclosures Rubnitz: AbbVie Inc.: Research Funding. Karol:AbbVie Inc.: Other: Unrelated to this study, St. Jude has received a charitable contribution from AbbVie, Inc. The charitable contribution is not being used for clinical or research activities, including any activities related to this study. . Pathan:Arog Pharmaceuticals: Current Employment. Messahel:AROG Pharmaceuticals: Current Employment.

FLT3 inhibitors in the treatment of acute myeloid leukemia: current status and future perspectives.

Mutations in the FMS-like tyrosine kinase 3 (FLT3) gene arise in 25-30% of all acute myeloid leukemia (AML) patients. These mutations lead to constitutive activation of the protein product and are divided in two broad types: internal tandem duplication (ITD) of the juxtamembrane domain (25% of cases) and point mutations in the tyrosine kinase domain (TKD). Patients with FLT3 ITD mutations have a high relapse risk and inferior cure rates, whereas the role of FLT3 TKD mutations still remains to be clarified. Additionally, growing research indicates that FLT3 status evolves through a disease continuum (clonal evolution), where AML cases can acquire FLT3 mutations at relapse - not present in the moment of diagnosis. Several FLT3 inhibitors have been tested in patients with FLT3-mutated AML. These drugs exhibit different kinase inhibitory profiles, pharmacokinetics and adverse events. First-generation multi-kinase inhibitors (sorafenib, midostaurin, lestaurtinib) are characterized by a broad-spectrum of drug targets, whereas second-generation inhibitors (quizartinib, crenolanib, gilteritinib) show more potent and specific FLT3 inhibition, and are thereby accompanied by less toxic effects. Notwithstanding, all FLT3 inhibitors face primary and acquired mechanisms of resistance, and therefore the combinations with other drugs (standard chemotherapy, hypomethylating agents, checkpoint inhibitors) and its application in different clinical settings (upfront therapy, maintenance, relapsed or refractory disease) are under study in a myriad of clinical trials. This review focuses on the role of FLT3 mutations in AML, pharmacological features of FLT3 inhibitors, known mechanisms of drug resistance and accumulated evidence for the use of FLT3 inhibitors in different clinical settings.

Abstract 3766: Development of new FLT3/AXL dual inhibitor JRF104 with potent FLT3-TKD inhibition

Abstract Background: More than 30% of de novo AML patients were identified with FLT3 mutations. Two kinds of mutations, internal tandem duplication (ITD) and tyrosine kinase domain (TKD) mutations, account for 23% and 7% of all AML cases, respectively. FLT3 inhibitors, which have been recommended by both NCCN and ELN guidelines, are often used as single or combo agent to treat FLT3 m+ AML. However, primary and secondary acquired resistance to FLT3 inhibitors is a remarkable challenge. It has been revealed that AXL can activate FLT3, resulting in reduction of response to FLT3 inhibitors. Furthermore, TKD mutations can also lead to development of drug resistance. Therefore, FLT3/AXL dual inhibitor with potent FLT3-TKD inhibition may provide better efficacy as novel AML treatment option. Results: JRF104 is a potent FLT3/AXL dual inhibitor with a kinase inhibition IC50 value of 0.42 nM and 1.22 nM, respectively. In vitro studies suggested that JRF104 could potently inhibit the proliferation of MV4-11 and Molm-13 cells, which endogenously express FLT3-ITD mutation. JRF104 shows wide spectrum in vitro antiproliferation activity against all FLT3 mutation types, including the “gate-keeper” F691L mutation. JRF104 can effectively reduce MDA-MB-231 cell migration in vitro, accomplished by blocking tumor migration pathway regulated by AXL. In MV4-11 xenograft model, JRF104 inhibited tumor growth at the dose of 1 mg/kg, QD (TGI: 65%); significantly reduce tumor volume at 5 mg/kg, QD (TGI: 118%). In Molm-13 xenograft model, tumor volume was well controlled at 15 mg/kg, QD (TGI: 94%), and all tumor volume reduced to zero at 50 mg/kg, QD (TGI: 104%). In CT-26 syngeneic model, JRF104 enhanced anti-tumor growth effect when combined with anti-PD-1 antibody and suppressed pERK and pAKT in tumor tissues. Additionally, JRF104 demonstrated high oral bioavailability in mouse, rat, and dog. In 28 days pre-clinical toxicology studies, JRF104 was well tolerated in all tested rats. HNSTD and NOEAL were determined to be 20 mg/kg QD and 3 mg/kg QD, respectively. All results support that JRF104 with high efficacy and safety. Conclusion: JRF104 demonstrated consistently potent inhibition of FLT3 and strong antitumor efficacy against FLT3-driven tumors in preclinical studies. Based on these datas, JRF104 may overcome resistance caused by AXL and FLT3-TKD mutation, representing a novel promising clinical antitumor agent for treating AML patients with FLT3 mutations. Citation Format: yuquan wei, Xiawei Wei, Yanzhong Zhang, Wanru Qin, Xiangyu Fu, Lihong Hu, Yuanfeng Xia, Jian Zuo, Zhong Xiong, Shanshan Yin, Shaohua Qin, Ge Dong, Ying Chen, Charles Ding, Chichung Chan, Shuhui Chen. Development of new FLT3/AXL dual inhibitor JRF104 with potent FLT3-TKD inhibition [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3766.

Molecular testing of metastatic non-small cell lung cancer in the Asia-Pacific region.

Molecular testing of metastatic non-small cell lung cancer in the Asia-Pacific region.

Gilteritinib: An FMS-like tyrosine kinase 3/AXL tyrosine kinase inhibitor for the treatment of relapsed or refractory acute myeloid leukemia patients.

Acute myeloid leukemia has recently undergone a significant transition into identifying and successfully inhibiting driver mutations leading to disease. One of the most common mutations in acute myeloid leukemia involves the protein FMS-like tyrosine kinase 3 (FLT3), which leads to ligand-independent activation of intracellular signaling cascades leading to the survival and proliferation of the acute leukemia blast cell. Preclinical studies have demonstrated the presence of two dominant types of mutations of this protein: internal tandem duplication and tyrosine kinase domain mutations. Successful inhibition of this protein has proven to be challenging. While FLT3 has been shown to be successfully inhibited and shown to improve overall survival in the frontline therapy of acute myeloid leukemia in combination with cytarabine and anthracycline, relapsed and refractory (R/R) patients have not been shown to be a successful population until recently. A phase III trial (ADMIRAL trial) demonstrated significant overall survival benefit in patients receiving gilteritinib compared to patients receiving salvage chemotherapy. This review will provide an overview of the preclinical, clinical, and practical use of gilteritinib in the treatment of patients with relapsed and refractory acute myeloid leukemia with FLT3 mutation.

Quizartinib-resistant FLT3-ITD acute myeloid leukemia cells are sensitive to the FLT3-Aurora kinase inhibitor CCT241736

Internal tandem duplication of FLT3 (FLT3-ITD) is one of the most common somatic mutations in acute myeloid leukemia (AML); it causes constitutive activation of FLT3 kinase and is associated with high relapse rates and poor survival. Small-molecule inhibition of FLT3 represents an attractive therapeutic strategy for this subtype of AML, although resistance from secondary FLT3 tyrosine kinase domain (FLT3-TKD) mutations is an emerging clinical problem. CCT241736 is an orally bioavailable, selective, and potent dual inhibitor of FLT3 and Aurora kinases. FLT3-ITD+ cells with secondary FLT3-TKD mutations have high in vitro relative resistance to the FLT3 inhibitors quizartinib and sorafenib, but not to CCT241736. The mechanism of action of CCT241736 results in significant in vivo efficacy, with inhibition of tumor growth observed in efficacy studies in FLT3-ITD and FLT3-ITD-TKD human tumor xenograft models. The efficacy of CCT241736 was also confirmed in primary samples from AML patients, including those with quizartinib-resistant disease, which induces apoptosis through inhibition of both FLT3 and Aurora kinases. The unique combination of CCT241736 properties based on robust potency, dual selectivity, and significant in vivo activity indicate that CCT241736 is a bona fide clinical drug candidate for FLT3-ITD and TKD AML patients with resistance to current drugs.

Activity of the multikinase inhibitor TP‐0903 in RAS mutant acute myeloid leukemia

Clonal heterogeneity and co‐occurring mutations contribute to drug resistance in AML. Leukemia samples from patients progressing on gilteritinib, a FLT3 inhibitor, were selected for RAS mutant clones, which are known to contribute preclinically to resistance (McMahon CM, Cancer Discov 2019). Previously, we demonstrated that TP‐0930, a multi‐kinase inhibitor, targets FLT3‐ITD and resistance‐conferring tyrosine kinase domain mutations in preclinical models. Here, we present the activity of TP‐0903 in RAS pathway mutant models of AML.KiNativ assay was performed in OCI‐AML3 cells (NRAS Q61L) treated with TP‐0903 (100 nM, 2h). Inhibition of cell viability was evaluated in AML cell lines (MTT) and primary samples (CellTiter Glo). Inhibition of AURKA/B, AKT, ERK, MCL‐1, and BCL‐2 were determined by western blot. Apoptosis, cell cycle, and differentiation assays were performed in OCI‐AML3 using annexin V, DAPI and CD11b‐APC staining. In vivo activity of TP‐0903 was evaluated in a systemic OCI‐AML3‐Luc+ xenograft model; 5e5 luciferase expressing cells were tail vein injected to NSG mice and TP‐0903 50 mg/kg or vehicle was given orally once dailyx5/week. Progression of leukemia was monitored via bioluminescence imaging and Kaplan Meier was used for survival analysis.In Kinativ assay, inhibition of native kinases were similar to those observed in a previous binding assay. We observed that TP‐0903 inhibited ACK1 (activated CDC42 kinase 1 or TNK2) and GCK (germinal center kinase or MAP4K2) by 87 and 46%, respectively. Inhibition of ACK1 (Kd = 7.3 nM, IC50 = 30.7 nM) and GCK (Kd = 1.8 nM, IC50 = 2.5 nM) was confirmed in binding and kinase assays. In RAS mutant AML cell lines (OCI‐AML3, THP1, HL60, and U937), TP‐0903 inhibited viability with IC50 values of 26 – 99 nM. In OCI‐AML3, TP‐0903 at 100 nM inhibited pAURKA/B, pAKT and MCL‐1; caused a G2‐M arrest and polyploidy (20 nM, 24h); and induced apoptosis (20–50 nM, 72h). TP‐0903, compared to gilteritinib, was more potent in inhibiting 4 ex vivo human primary AML samples with N/KRAS mutations co‐occurring with wt FLT3 or FLT3‐ITD, along with other mutations (IC50 range, 21 – 60 vs 260 – 1370 nM). In a 14‐day CFU assay, TP‐0903 inhibited colony formation of a FLT3‐ITD and NRAS mutant primary sample compared to DMSO (mean, 65 vs 188 CFUs). In an OCI‐AML3‐Luc+ xenograft study, TP‐0903 suppressed the outgrowth of leukemia at day 21 (P<0.0001) and prolonged median survival by 9 days compared to veh‐treated mice (P<0.0001). At study endpoint, spleen weight was higher in veh‐treated mice than in TP‐0903‐treated mice (mean ± SD, 333 ± 23 vs 181 ± 21 mg, P=0.0029).TP‐0903, a novel multi‐kinase inhibitor, shows promising in vitro and in vivo activity in RAS mutant AML. In a previous report (Nonami A, Blood 2015), an integrated approach involving cell‐based pharmacologic screening combined with KiNativ, gene expression profiling and mechanism studies, identified 2 kinase signaling pathways (ACK1/AKT and GCK) that synergistically contributed to the growth of NRAS mutant leukemia cells, including OCI‐AML3. Based on our data, TP‐0903 may exert its activity in RAS mutant AML via inhibiting ACK1 and GCK.

A common MET polymorphism harnesses HER2 signaling to drive aggressive squamous cell carcinoma

c-MET receptors are activated in cancers through genomic events like tyrosine kinase domain mutations, juxtamembrane splicing mutation and amplified copy numbers, which can be inhibited by c-MET small molecule inhibitors. Here, we discover that the most common polymorphism known to affect MET gene (N375S), involving the semaphorin domain, confers exquisite binding affinity for HER2 and enables METN375S to interact with HER2 in a ligand-independent fashion. The resultant METN375S/HER2 dimer transduces potent proliferative, pro-invasive and pro-metastatic cues through the HER2 signaling axis to drive aggressive squamous cell carcinomas of the head and neck (HNSCC) and lung (LUSC), and is associated with poor prognosis. Accordingly, HER2 blockers, but not c-MET inhibitors, are paradoxically effective at restraining in vivo and in vitro models expressing METN375S. These results establish METN375S as a biologically distinct and clinically actionable molecular subset of SCCs that are uniquely amenable to HER2 blocking therapies.

Targeting AML-associated FLT3 mutations with a type I kinase inhibitor.

Tyrosine kinase domain (TKD) mutations contribute to acquired resistance to FMS-like tyrosine kinase 3 (FLT3) inhibitors used to treat FLT3-mutant acute myeloid leukemia (AML). We report a cocrystal structure of FLT3 with a type I inhibitor, NCGC1481, that retained potent binding and activity against FLT3 TKD and gatekeeper mutations. Relative to the current generation of advanced FLT3 inhibitors, NCGC1481 exhibited superior antileukemic activity against the common, clinically relevant FLT3-mutant AML cells in vitro and in vivo.

Gilteritinib is a clinically active FLT3 inhibitor with broad activity against FLT3 kinase domain mutations

AbstractGilteritinib is the first FMS-like tyrosine kinase 3 (FLT3) tyrosine kinase inhibitor (TKI) approved as monotherapy in acute myeloid leukemia with FLT3 internal tandem duplication and D835/I836 tyrosine kinase domain (TKD) mutations. Sequencing studies in patients have uncovered less common, noncanonical (NC) mutations in FLT3 and have implicated secondary TKD mutations in FLT3 TKI resistance. We report that gilteritinib is active against FLT3 NC and TKI resistance-causing mutations in vitro. A mutagenesis screen identified FLT3 F691L, Y693C/N, and G697S as mutations that confer moderate resistance to gilteritinib in vitro. Analysis of patients treated with gilteritinib revealed that 2/9 patients with preexisting NC FLT3 mutations responded and that secondary TKD mutations are acquired in a minority (5/31) of patients treated with gilteritinib. Four of 5 patients developed F691L mutations (all treated at <200 mg). These studies suggest that gilteritinib has broad activity against FLT3 mutations and limited vulnerability to resistance-causing FLT3 TKD mutations, particularly when used at higher doses.

Outcomes of allogeneic stem cell transplantation for patients with t(6:9) AML- A strong case for allogeneic stem cell transplantation in first complete remission.

Outcomes of allogeneic stem cell transplantation for patients with t(6:9) AML- A strong case for allogeneic stem cell transplantation in first complete remission.

High incidence of FLT3 mutations in follicular thyroid cancer: potential therapeutic target in patients with advanced disease stage.

Background:Conventional treatments for follicular thyroid cancer (FTC) can be ineffective, leading to poor prognosis. The aim of this study was to identify mutations associated with FTC that would serve as novel molecular markers of the disease and its outcome and could potentially identify new therapeutic targets.Methods:FLT3 mutations were first detected in a 29-year-old White female diagnosed with metastasized, treatment-refractory FTC. Analyses of FLT3 mutational status through next-generation sequencing of formalin-fixed, paraffin-embedded FTC specimens were subsequently performed in 35 randomly selected patients diagnosed with FTC.Results:FLT3 mutations were found in 69% of patients. FLT3 mutation-positive patients were significantly older than those that were FLT3 mutation-negative [median age at diagnosis 54 (36–82) versus 45 (27–58) (p = 0.023)]. Patients over 60 years were 23 times more likely to be FLT3 mutation-positive (p = 0.006). However, the number of FLT3 mutations did not correlate with age (r-Pearson: –0.244, p-value: 0.25). A total of 26 mutations were identified in the FLT3 gene with 2–16 FLT3 mutations in each FLT3 mutation-positive patient (mean: 5.6 mutations/patient). Tyrosine kinase domain (TKD) mutations in the FLT3 gene were detected in 58% of FLT3 mutation-positive patients. All FLT3 mutation-positive patients with a disease stage of pT2N1 or worse harbored at least one mutation in the TKD of FLT3.Conclusions:There is a wide spectrum and high frequency of FLT3 mutations in FTC. The precise role of FLT3 mutations in the genesis of FTC, as well as its potential role as a therapeutic target, requires further investigation.