Clenolanib for the treatment of relapsed / refractory FLT3 mutation-induced proliferative disorder to previous treatments.

Abstract

To provide methods for treating a proliferative disorder in a subject with mutated or constitutively active FLT3 who has relapsed after or is refractory to one or more prior tyrosine kinase inhibitors.SOLUTION: A method for treatment comprises: obtaining a tumor sample from the subject who has relapsed after or is refractory to one or more prior tyrosine kinase inhibitors; measuring expression of a mutated or constitutively active FLT3 mutant in the tumor sample; and administering to the subject a therapeutically effective amount of crenolanib or a pharmaceutically acceptable salt thereof sufficient to treat the proliferative disorder.SELECTED DRAWING: None

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS None TECHNICAL FIELD

[0002] The present invention is directed to the use of krenolanib, or a salt thereof, as a single agent or in combination with another agent for the treatment of cancer, and to methods for treating an animal suffering from a relapsed / refractory FLT3 mutation proliferative disorder against prior cancer treatment.

[0003] STATEMENT REGARDING FEDERALLY SPONSORED RESEARCH None

[0004] INCORPORATION BY REFERENCE OF MATERIALS FILED ON COMPACT DISC None BACKGROUND ART

[0005] Without limiting the scope of the present invention, the background art is described in relation to its ability to inhibit FLT3 tyrosine kinase in the treatment of proliferative disorders involving mutated or otherwise activated FLT3.

[0006] Protein kinases are enzymes that chemically modify other proteins by catalyzing the transfer of phosphate groups to the amino acid residues serine, threonine, or tyrosine. Approximately 30% of all human proteins can be modified by kinase activity, and kinase signaling is involved in several cellular processes including growth, proliferation, and survival.

[0007] Abnormal expression or activation of protein kinases is often associated with proliferative disorders, including various cancers, due to their involvement in cell proliferation and survival, including through mutation, copy number increase, amplification, gene fusion, or other mechanisms. Therefore, investigating compounds that potently inhibit the activity and function of protein kinases can lead to a better understanding of their physiological roles.

[0008] The FMS-like tyrosine kinase 3 (FLT3) gene encodes a membrane-bound receptor tyrosine kinase that affects hematopoiesis, and abnormalities in FLT3 signaling can lead to hematological disorders and malignancies (Gilliland & Griffin, 2002; Stirewalt & Radich, 2003). Activation of FLT3 receptor tyrosine kinase is initiated via the binding of FLT3 ligand (FLT3L) to the FLT3 receptor, and activation initiates homodimerization, cross-phosphorylation, and recruitment of downstream signaling factors of the ligand-bound receptor.

[0009] FLT3 is one of the most frequently mutated genes in hematological malignancies, present in approximately 30% of adult acute myeloid leukemia (AML) cases (Papaemmanuil et al., 2016; Rucker et al., 2021; Tyner et al., 2018), and the presence or absence of FLT3 mutations is included in international guidelines for AML risk stratification (Dohner et al., 2017).

[0010] The most common FLT3 mutation is internal tandem duplication (ITD), resulting in an in-frame insertion within the near-membrane domain of the FLT3 receptor, and is reported in 20–30% of adult AML patients. FLT3-ITD mutations are an independent predictor of poor patient prognosis and are associated with an increased risk of relapse after standard therapy and reduced disease-free time and overall survival (DiNardo & Lachowiez, 2019; Papaemmanuil et al., 2016; Rucker et al., 2021; Sakaguchi et al., 2019; Tyner et al., 2018). Point mutations with ligand-binding or kinase domain mutations are less frequent than ITD mutations but are also prognostically significant. The most commonly affected amino acid residue is aspartate 835 (D835) in the activation loop. Missense mutations (nucleotide substitutions) at D835 occur in approximately 5–10% of adult AML patients (Stirewalt & Radich, 2003; Tyner et al., 2018). While ITD and D835 mutations are detectable using PCR-based techniques that enable relatively low-cost and relatively rapid results, the current commercial availability of next-generation sequencing panels, as well as the wider availability of reagents and equipment required for such panels performed outside commercial laboratories, has redefined the mutation landscape of common AML and, in particular, FLT3-mutant AML. Detailed genetic analysis of 500 AML patient samples found that up to 20% of the point mutations identified in FLT3 did not contain amino acid D835 (Tyner et al., 2018).

[0011] Due to the frequency of FLT3-activating mutations in AML, this kinase is an interesting target in drug discovery. Several FLT3 inhibitors with varying degrees of potential and selectivity for activated FLT3 have been and are currently being investigated in AML patients. To date, two FLT3 inhibitors have been approved by the United States Food and Drug Administration (FDA) or the European Medicines Agency (EMA) for use in FLT3-mutated AML: midostaurin is approved for use in combination with chemotherapy for the treatment of newly diagnosed FLT3-mutated AML; gilteritinib is approved for use as a monotherapy in relapsed or refractory FLT3-mutated AML (FDA, 2019, 2020). While midostaurin and gilteritinib are the only currently approved FLT3 inhibitors, several other compounds have been investigated in the past or are currently under investigation.

[0012] While FLT3 inhibition has proven to be a desirable treatment option, it has some drawbacks compared to monotherapy. The majority of FLT3 inhibitors tested are tyrosine kinase inhibitors (TKIs), including midostaurine, gilteritinib, and the present invention. Tyrosine kinase inhibitors can be susceptible to resistance mutations, which are mutations in target genes that confer resistance to specific TKIs. The presence of resistance-conferring mutations not only predicts an incomplete response before TKI administration, but patients may also acquire resistance-conferring mutations while receiving TKIs, leading to relapses. Often, these resistance-conferring mutations cause structural changes in kinases that block TKI binding.

[0013] For example, a criterion FLT3 kinase domain mutation at amino acid residue D835 confers resistance to the TKIs quizartinib and sorafenib. Both quizartinib and sorafenib are "type II" inhibitors and bind to a hydrophobic site near the ATP binding pocket. Mutations at D835 alter the structure of this hydrophobic site, preventing inhibitor binding (CCSmith, Lin, Stecula, Sali, & Shah, 2015). Mutations at amino acids F691 and N701, "gatekeeper residues," confer resistance to gilteritinib by altering the structure of the ATP binding pocket to which gilteritinib binds (Joshi et al., 2021). Up to 12% of patients who relapse after monotherapy with gilteritinib develop a relapse-contributing FLT3-F691 mutation (McMahon et al., 2019). Mutations at amino acid residues A637, N676, G697, Y842, and A848 have been associated with resistance to several FLT3 inhibitors (Wang et al., 2021). Acquisition of secondary FLT3 mutations can explain the relatively short event-free survival (EFS) observed with monotherapy of FLT3 inhibitors, with a reported mean EFS of 2.8 months for gilteritinib monotherapy (Perl et al., 2019).

[0014] The majority of patients treated with either midostaurin or gilteritinib retain mutated FLT3 at relapse, either retaining the original mutated clone or acquiring secondary mutations (Altman et al., 2021; McMahon et al., 2019; Schmalbrock et al., 2021). These patients benefit from FLT3 inhibitor therapy at relapse because salvage chemotherapy alone does not provide high cure rates for relapsed / refractory FLT3 mutant AML (see control arm of the gilteritinib phase III trial) (Perl et al., 2019). However, patients who are relapsed / refractory to one FLT3 TKI are less likely to benefit from monotherapy with a second, or even third, FLT3 TKI, partly due to the potential acquisition of resistance-constituting mutations (Perl et al., 2021). [Overview of the project] [Problems that the invention aims to solve]

[0015] To ensure patients receive the best possible benefit, it is necessary to use pan-FLT3 inhibitors that are active against several resistance-constituting FLT3 variants, either alone or in combination with other drugs. Therefore, the need to treat patients with FLT3 mutational proliferation disorders who have been treated with one or more FLT3 tyrosine kinase inhibitors remains unaddressed. [Means for solving the problem]

[0016] This invention overcomes the limitations of the prior art by using crenolanib (and its pharmaceutically acceptable salts), a potent pan-FLT3 inhibitor active against resistance-constituting mutations, in the treatment of relapsed or refractory FLT3 mutational disorder following prior treatment with FLT3 inhibitors.

[0017] In one embodiment, the present invention relates to a method for treating proliferative disorders in subjects having mutated or constitutively active FLT3 that are recurrent / refractory to one or more prior tyrosine kinase inhibitors, the method comprising the steps of: obtaining a tumor sample from a subject that is recurrent / refractory to one or more prior tyrosine kinase inhibitors; measuring the expression of mutated or constitutively active FLT3 mutants in the tumor sample; and administering to the subject a therapeutically effective dose of crenolanib or a pharmaceutically acceptable salt thereof sufficient to treat the proliferative disorder. In one embodiment, mutated or constitutively active FLT3 is at least one of FLT3-ITD; FLT3-TKD; activating mutation in FLT3; copy number increase or amplification of the FLT3 gene; or gene fusion including fusion of FLT3 with another gene. In another embodiment, subjects have been provided with prior tyrosine kinase inhibitors selected from midostaurin, sorafenib, gilteritinib, quizaltinib, pexidartinib, FF-10101, CG-806, restoltinib, AG1295, AG1296, CEP-5214, CEP-7055, HM43239, pacritinib, MAX-40279, FYSYN, NMS-03592088, or TG02 citrate; or subjects have an FLT3 mutation that confers resistance to prior tyrosine kinase inhibitors. In another embodiment, the resistance-constituting FLT3 mutation is selected from missense mutations occurring alone or in combination with an FLT3-ITD mutation at at least one of the following amino acid residues: K429, A627, N676, A680, F691, Y693, G697, D698, N701, D835, N841, Y842, and A848. In another embodiment, the resistance-constituting FLT3 mutation was present before administration of the previous tyrosine kinase inhibitor, or was acquired during or after administration of the previous tyrosine kinase inhibitor.In another embodiment, the proliferative disorder is selected from at least one of the following: gastrointestinal stromal tumors, leukemia, myeloma, myeloproliferative disorders, myelodysplastic syndromes, idiopathic eosinophilic syndrome (HES), bladder cancer, breast cancer, cervical cancer, CNS cancer, colon cancer, esophageal cancer, head and neck cancer, liver cancer, lung cancer, nasopharyngeal cancer, neuroendocrine cancer, ovarian cancer, pancreatic cancer, prostate cancer, kidney cancer, salivary gland cancer, small cell lung cancer, skin cancer, gastric cancer, testicular cancer, thyroid cancer, uterine cancer, and hematological malignancies. In another embodiment, the therapeutically effective dose of crenolanib or a pharmaceutically acceptable salt thereof is approximately 50–500 mg per day, 100–450 mg per day, 200–400 mg per day, 300–500 mg per day, 350–500 mg per day, or 400–500 mg per day. In another embodiment, a therapeutically effective dose of crenolanib or a pharmaceutically acceptable salt thereof is administered sequentially, intermittently, systemically, or topically in at least one of these methods. In another embodiment, a therapeutically effective dose of crenolanib or a pharmaceutically acceptable salt thereof is administered orally, intravenously, or intraperitoneally. In another embodiment, a therapeutically effective dose of crenolanib or a pharmaceutically acceptable salt thereof is administered up to three times a day or more, as long as the subject requires treatment for the proliferative disorder. In another embodiment, a therapeutically effective dose of crenolanib or a pharmaceutically acceptable salt thereof is provided: sequentially or concurrently with at least one other agent to maintain remission in an existing patient; provided as monotherapy or in combination with another agent to maintain remission in a patient with relapsing / refractory proliferative disorder; or provided as monotherapy or in combination with another agent to maintain remission in a pediatric patient with relapsing / refractory proliferative disorder. In another embodiment, clenolanib or a pharmaceutically acceptable salt thereof is clenolanib besylate, clenolanibrine, clenolanib lactate, clenolanib hydrochloride, clenolanib citrate, clenolanib acetate, clenolanib toluenesulfonate, or clenolanib succinate.

[0018] In another embodiment, the present invention provides a method for inhibiting or reducing mutant FLT3 tyrosine kinase activity or expression in subjects suffering from proliferative disorders, comprising the steps of: identifying subjects who have discontinued previous tyrosine kinase inhibitor therapy due to refractory or recurrent proliferative disorders; obtaining a tumor sample from the subjects; measuring the expression of mutated FLT3 or a constitutively active FLT3 mutant in the tumor sample; and, if the subjects have mutated FLT3 or a constitutively active FLT3 mutant, administering to the subjects a therapeutically effective dose of crenolanib or a pharmaceutically acceptable salt thereof, wherein crenolanib or a salt thereof reduces the proliferative disorder burden or prevents the progression of the proliferative disorder. In one embodiment, the mutated or constitutively active FLT3 is at least one of FLT3-ITD; FLT3-TKD; activating mutation in FLT3; copy number increase or amplification of the FLT3 gene; or gene fusion including fusion of FLT3 with another gene. In another embodiment, subjects have been provided with prior tyrosine kinase inhibitors selected from midostaurin, sorafenib, gilteritinib, quizaltinib, pexidartinib, FF-10101, CG-806, restoltinib, AG1295, AG1296, CEP-5214, CEP-7055, HM43239, pacritinib, MAX-40279, FYSYN, NMS-03592088, or TG02 citrate; or subjects have an FLT3 mutation that confers resistance to prior tyrosine kinase inhibitors. In another embodiment, the subject is recurrent or refractory to a prior tyrosine inhibitor and has a resistance-constitutive FLT3 mutation selected from missense mutations occurring alone or in combination with an FLT3-ITD mutation in at least one of the following amino acid residues: K429, A627, N676, A680, F691, Y693, G697, D698, N701, D835, N841, Y842, A848. In another embodiment, the resistance-constitutive FLT3 mutation was present before administration of the prior tyrosine kinase inhibitor or was acquired during or after administration of the prior tyrosine kinase inhibitor.In another embodiment, the proliferative disorder is selected from at least one of the following: gastrointestinal stromal tumors, leukemia, myeloma, myeloproliferative disorders, myelodysplastic syndromes, idiopathic eosinophilic syndrome (HES), bladder cancer, breast cancer, cervical cancer, CNS cancer, colon cancer, esophageal cancer, head and neck cancer, liver cancer, lung cancer, nasopharyngeal cancer, neuroendocrine cancer, ovarian cancer, pancreatic cancer, prostate cancer, kidney cancer, salivary gland cancer, small cell lung cancer, skin cancer, gastric cancer, testicular cancer, thyroid cancer, uterine cancer, and hematological malignancies. In another embodiment, the therapeutically effective dose of crenolanib or a pharmaceutically acceptable salt thereof is approximately 50–500 mg per day, 100–450 mg per day, 200–400 mg per day, 300–500 mg per day, 350–500 mg per day, or 400–500 mg per day. In another embodiment, a therapeutically effective dose of crenolanib or a pharmaceutically acceptable salt thereof is administered sequentially, intermittently, systemically, or topically in at least one of these methods. In another embodiment, a therapeutically effective dose of crenolanib or a pharmaceutically acceptable salt thereof is administered orally, intravenously, or intraperitoneally. In another embodiment, a therapeutically effective dose of crenolanib or a pharmaceutically acceptable salt thereof is administered up to three times a day or more, as long as the subject requires treatment for the proliferative disorder. In another embodiment, a therapeutically effective dose of crenolanib or a pharmaceutically acceptable salt thereof is provided sequentially or concurrently with at least one other agent to maintain remission in an existing patient; provided as monotherapy or in combination with another agent in a patient to maintain remission in a patient with relapsing / refractory proliferative disorder; or provided as monotherapy or in combination with another agent in a pediatric patient with relapsing / refractory proliferative disorder to maintain remission. In another embodiment, clenolanib or a pharmaceutically acceptable salt thereof is clenolanib besylate, clenolanibrine, clenolanib lactate, clenolanib hydrochloride, clenolanib citrate, clenolanib acetate, clenolanib toluenesulfonate, or clenolanib succinate.

[0019] In another embodiment, the present invention provides a method for treating a subject suffering from a proliferative disorder, comprising the steps of: obtaining or having obtained a biological sample from the patient to determine whether the patient has a gene mutation in the FLT3 gene, a change in the kinase activity of FLT3 tyrosine kinase, overexpression of FLT3 tyrosine kinase, or a change in the phenotype or genotype of FLT3 tyrosine kinase; and performing or having performed an assay on the biological sample to determine whether the subject has increased FLT3 tyrosine kinase activity; treating the patient with a first tyrosine kinase inhibitor; and, if the patient is refractory or relapses after the first tyrosine kinase inhibitor, and the patient has a gene mutation in FLT3; a change in the kinase activity of FLT3, overexpression of FLT3, or a change in the phenotype of FLT3 tyrosine kinase, administering an effective dose of crenolanib to the patient to reduce the proliferative disorder burden or prevent the progression of the proliferative disorder. In one embodiment, the mutated or constitutively active FLT3 is at least one of the following: FLT3-ITD; FLT3-TKD; activating mutation in FLT3; copy number increase or amplification of the FLT3 gene; or gene fusion including fusion of FLT3 with another gene. In another embodiment, the subject has been provided with a prior tyrosine kinase inhibitor selected from midostaurin, sorafenib, gilteritinib, quizaltinib, pexidartinib, FF-10101, CG-806, restoltinib, AG1295, AG1296, CEP-5214, CEP-7055, HM43239, pacritinib, MAX-40279, FYSYN, NMS-03592088, or TG02 citrate; or the subject has an FLT3 mutation that confers resistance to the prior tyrosine kinase inhibitor.In another embodiment, a mutation in the FLT3 gene or a change in the phenotype or genotype of FLT3 is a resistance-constituting mutation selected from missense mutations occurring alone or in conjunction with an FLT3-ITD mutation in at least one of the following amino acid residues: K429, A627, N676, A680, F691, Y693, G697, D698, N701, D835, N841, Y842, and A848. In another embodiment, the resistance-constituting FLT3 mutation was present before administration of the previous tyrosine kinase inhibitor, or the resistance-constituting FLT3 mutation was acquired during or after administration of the previous tyrosine kinase inhibitor. In another embodiment, the proliferative disorder is selected from at least one of the following: gastrointestinal stromal tumors, leukemia, myeloma, myeloproliferative disorders, myelodysplastic syndromes, idiopathic eosinophilic syndrome (HES), bladder cancer, breast cancer, cervical cancer, CNS cancer, colon cancer, esophageal cancer, head and neck cancer, liver cancer, lung cancer, nasopharyngeal cancer, neuroendocrine cancer, ovarian cancer, pancreatic cancer, prostate cancer, kidney cancer, salivary gland cancer, small cell lung cancer, skin cancer, gastric cancer, testicular cancer, thyroid cancer, uterine cancer, and hematological malignancies. In another embodiment, the therapeutically effective dose of crenolanib or a pharmaceutically acceptable salt thereof is approximately 50–500 mg per day, 100–450 mg per day, 200–400 mg per day, 300–500 mg per day, 350–500 mg per day, or 400–500 mg per day. In another embodiment, a therapeutically effective dose of crenolanib or a pharmaceutically acceptable salt thereof is administered in at least one of the following manners: continuous, intermittent, systemic, or topical. In another embodiment, a therapeutically effective dose of crenolanib or a pharmaceutically acceptable salt thereof is administered orally, intravenously, or intraperitoneally. In another embodiment, a therapeutically effective dose of crenolanib or a pharmaceutically acceptable salt thereof is administered up to three times a day or more, as long as the subject requires treatment for proliferative disorders.In another embodiment, a therapeutically effective dose of crenolanib or a pharmaceutically acceptable salt thereof is provided sequentially or concurrently with at least one other agent to maintain remission in an existing patient; provided monotherapy or in combination with another agent in a patient to maintain remission in a patient with a relapsing / refractory proliferative disorder; or provided monotherapy or in combination with another agent to maintain remission in a pediatric patient with a relapsing / refractory proliferative disorder. In another embodiment, crenolanib or a pharmaceutically acceptable salt thereof is crenolanib besylate, crenolanibuline, crenolanib lactate, crenolanib hydrochloride, crenolanib citrate, crenolanib acetate, crenolanib toluenesulfonate, or crenolanib succinate.

[0020] In another embodiment, the present invention provides a method for treating a subject suffering from leukemia, comprising the steps of: obtaining a sample from the subject; determining from the subject sample whether the patient has a dysregulated FLT3 receptor or a constitutively active FLT3 receptor; further determining whether the subject is refractory to administration or has relapsed after administration of a previous tyrosine kinase inhibitor; and administering to the subject in need of such treatment a therapeutically effective dose of crenolanib or a pharmaceutically acceptable salt thereof sufficient to treat leukemia. In one embodiment, the mutated or constitutively active FLT3 is at least one of the following: FLT3-ITD; FLT3-TKD; activating mutation in FLT3; copy number increase or amplification of the FLT3 gene; or gene fusion including fusion of FLT3 with another gene. In another embodiment, subjects have been provided with prior tyrosine kinase inhibitors selected from midostaurin, sorafenib, gilteritinib, quizaltinib, pexidartinib, FF-10101, CG-806, restoltinib, AG1295, AG1296, CEP-5214, CEP-7055, HM43239, pacritinib, MAX-40279, FYSYN, NMS-03592088, or TG02 citrate; or subjects have an FLT3 mutation that confers resistance to prior tyrosine kinase inhibitors. In another embodiment, the mutation in the FLT3 gene or the change in the phenotype or genotype of FLT3 is a resistance-constituting mutation selected from missense mutations occurring in at least one of the amino acid residues K429, A627, N676, A680, F691, Y693, G697, D698, N701, D835, N841, Y842, and A848, either alone or in conjunction with an FLT3-ITD mutation. In another embodiment, the resistance-constituting FLT3 mutation was present before administration of the previous tyrosine kinase inhibitor, or the resistance-constituting FLT3 mutation was acquired during or after administration of the previous tyrosine kinase inhibitor.In another aspect, the leukemia is selected from at least one of Hodgkin's disease, myeloma, acute promyelocytic leukemia (APL), chronic lymphocytic leukemia (CLL), chronic myeloid leukemia (CML), chronic neutrophilic leukemia (CNL); acute undifferentiated leukemia (AUL), anaplastic large cell lymphoma (ALCL), prolymphocytic leukemia (PML); juvenile myelomonocytic leukemia (JMML); adult T-cell ALL, acute myeloid leukemia (AML), AML with trilineage myelodysplasia, myelodysplastic syndrome (MDS), myeloproliferative neoplasm (MPN), or multiple myeloma (MM). Modes for Carrying Out the Invention

[0021] While the making and using of various embodiments of the present invention are discussed in detail below, it should be appreciated that the present invention provides many applicable inventive concepts that can be embodied in a wide variety of specific contexts. The specific embodiments discussed herein are merely illustrative of specific manners to make and use the invention, and do not delimit the scope of the present invention.

[0022] To facilitate understanding of the present invention, several terms are defined below. The terms defined herein have meanings as commonly understood by a person of ordinary skill in the areas relevant to the present invention. For example, terms such as "a", "an" and "the" are not intended to refer only to a singular entity, but are intended to include general classes for which specific examples may be used for illustration. Although these terms are used herein to describe particular embodiments of the present invention, their use is not intended to delimit the scope of the present invention, except as outlined in the claims.

[0023] The present invention is directed to administering clenoquinib, or a pharmaceutically acceptable salt thereof, to a subject suffering from cancer, for treating cancer, preventing cancer recurrence, and / or preventing cancer progression.

[0024] Crenolanib is an orally bioavailable TKI that targets FLT3. Crenolanib is significantly more selective for FLT3 than for other kinases including c-KIT, VEGFR2, TIE2, FGFR2, EGFR, erbB2, and SRC (Lewis et al., 2009). As a type I TKI, crenolanib binds to both the active and inactive conformations of the kinase. Importantly, crenolanib exhibits preclinical activity against quizartinib- and gilteritinib-resistant FLT3 mutations, including missense mutations at D835 or F691 (C.C. Smith et al., 2014). In a direct enzyme inhibition assay, crenolanib was found to inhibit >99% of the kinase activity of the FLT3-F691L mutant at a concentration of 10 nM. FLT3-F691L is overexpressed in cell lines. Crenolanib blocks FLT3 phosphorylation at nanomolar concentrations. Thus, crenolanib is ideally suited for the treatment of subjects suffering from constitutively active FLT3 proliferative disorders whose disease has progressed as a result of secondary mutations that confer resistance, leading to discontinuation of treatment with other FLT3 TKIs. As a pan-FLT3 inhibitor, crenolanib has shown activity in subjects with cancers associated with FLT3 copy number gain, amplification, fusion, or constitutively active variants.

[0025] The present invention includes methods of inhibiting mutant or constitutively active FLT3 in a cell or subject, or methods of treating a disorder associated with FLT3 activity or expression in a subject. In one embodiment, the present invention provides a method for reducing or inhibiting the kinase activity of mutant FLT3 in a subject, comprising the step of administering a compound of the present invention to the subject. In other embodiments, the present invention provides therapeutic methods for treating a subject having a proliferative disorder driven by aberrant kinase activity of mutant FLT3. The present invention also provides methods for treating patients suffering from a proliferative disorder that is relapsed / refractory to prior tyrosine kinase inhibitors.

[0026] As used herein, the term “subject” refers to an animal, such as a mammal or a human, which is the subject of treatment, observation or experimentation.

[0027] As used herein, the term “to bring into contact” means to add crenolanib or a pharmaceutically acceptable salt thereof to cells so that the compound may be absorbed by the cells.

[0028] As used herein, the term “therapeutic dose” means an amount of crenolanib or a pharmaceutically acceptable salt thereof, which is the amount that elicits a biological or pharmacokinetic response in a subject, as explored by researchers, veterinarians, doctors of medicine or other clinicians, and which includes an increase in progression-free survival or overall survival, including a reduction in the symptoms of the disease or disorder being treated, a reduction in the burden of proliferative disorders (e.g., a reduction in tumor size), and / or an increase in long-term stable disease. Methods for determining a therapeutic dose for a pharmaceutical composition comprising the compounds of the present invention are known in the art.

[0029] As used herein, the term “composition” is intended to encompass products containing the specified components in specified amounts, as well as any products directly or indirectly produced from combinations of the specified components in specified amounts.

[0030] As used herein, the terms “mutant FLT3-related disorders” or “mutant FLT3-driven cell proliferation disorders” include disorders associated with or related to mutations that result in mutant FLT3 activity, such as constitutive activation of FLT3.

[0031] As used herein, the term “proliferative disorder” refers to the excessive proliferation of one or more subsets of cells within a multicellular organism that causes harm to the organism (i.e., discomfort or reduced estimated lifespan). Proliferative disorders can occur in different types of animals and in humans. Examples of practicing proliferative disorders include gastrointestinal stromal tumors (GISTs), leukemia, myeloma, myeloproliferative disorders, myelodysplastic syndromes, idiopathic eosinophilic syndrome (HES), bladder cancer, breast cancer, cervical cancer, CNS cancer, colon cancer, esophageal cancer, head and neck cancer, liver cancer, lung cancer, nasopharyngeal cancer, neuroendocrine cancer, ovarian cancer, pancreatic cancer, prostate cancer, kidney cancer, salivary gland cancer, small cell lung cancer, skin cancer, gastric cancer, testicular cancer, thyroid cancer, uterine cancer, and hematological malignancies.

[0032] As used herein, the terms “recurrent / refractory” or “recurring” refer to subjects who have previously received medication to treat a proliferative disorder but either did not respond to treatment (refractory) or progressed after an initial response (recurrent).

[0033] The detection of mutant FLT3 can be carried out using any suitable means known in the art. For example, the detection of gene mutations can be achieved by detecting nucleic acid molecules (e.g., DNA) using nucleic acid amplification methods (e.g., RT-PCR) or high-throughput sequencing (i.e., "next-generation sequencing"). As an example, a next-generation sequencing platform, e.g., Illumina, can be used to determine the precise gene sequence of a specific gene or a portion of a target gene. Briefly, DNA from a tumor sample is fragmented, ligated with appropriate primers and adapters, and amplified using PCR during "library preparation." The prepared library is then sequenced using one of several commercially available systems that generate sequences of a selected target gene, all exomes, or the entire genome. The sequence is then analyzed using commercially available software to align the tumor sample sequence with a known sequence of the target gene, perform a variant calling step to identify differences at the DNA level in the tumor sample, and determine whether such mutations result in amino acid sequence modifications in the translated protein. Using these systems, a person skilled in the art can determine whether a subject has one of the FLT3-specific mutations. Further information about FLT3, including the whole gene and protein sequence, known clinically relevant variants and mutations, tissue expression, and signaling interaction partners, can be found in UniProt (accession number P36888-1), GenBank (accession number NM_04119.2), and GenPept (accession number rNP_004110.2).

[0034] As used herein, the term “missense mutation” refers to a modification of the FLT3 gene sequence in which, when the sequence is converted into a protein, one amino acid is consequently replaced by a different amino acid.

[0035] As used herein, the term “missense mutation” refers to a modification of the FLT3 gene sequence in which, when the sequence is converted into a protein, one amino acid is consequently replaced by a different amino acid.

[0036] As used herein, the term “ITD” or “internal tandem duplication” refers to a nucleotide insertion at the DNA level in which the number of nucleotides is a multiple of three, which results in the addition of amino acids at the protein level but does not shift the gene's readability frame.

[0037] As used herein, the terms “resistance mutation,” “mutation-constituting resistance,” or “secondary mutation” refer to non-ITD mutations in the FLT3 gene that make the patient ineligible to gilteritinib, midostaurin, quizaltinib, or other TKIs not of the present invention. In other words, these mutations, whether present alone or in combination with an ITD, retain kinase activity when treated with midostaurin, gilteritinib, or other TKIs, but are inhibited by the present invention. Non-limiting examples of resistance mutations are missense mutations at amino acid residues K429, A627, N676, A680, F691, Y693, G697, D698, N701, D835, N841, Y842, or A848. Additional mutations in the immunoglobulin-like domain, near-membrane domain, tyrosine kinase domain, and hinge region are also within the scope of the present invention.

[0038] As used herein, the terms “copy number increase” or “copy number variation” refer to the presence of more than two copies but less than five copies of the FLT3 gene. As used herein, “amplification” refers to an increase of more than five copies of the FLT3 gene or signal per cell. Number increases and / or amplifications can be detected by any means known in the art, for example, by fluorescence in situ hybridization (FISH), which involves incubating cells with a fluorescently labeled probe that binds to a specific region of DNA and counting the “signal” number (the number of regions of DNA to which the probe binds).

[0039] FLT3 kinase inhibitors known in the art include Restoltinib (also known as CEP-701, licensed to Kyowa Hakko and Cephalon); CHIR-258 (Chiron Corp.); EB10 and IMC-EB10 (ImcLone Systems Inc.); Midostaurin (also known as PKC412, Novartis AG); Tanzutinib (also known as MLN-518, licensed to COR Therapeutics Inc. and Millennium Pharmaceuticals Inc.); Sunitinib (also known as SU11248, Pfizer USA); Quizartinib (also known as AC220, Daiichi Sankyo); XL-999 (Exelixis USA); and GTP 14564 (Merck Biosciences). UK); AG1295 and AG1296; CEP-5214 and CEP-7055 (Cephalon); Gilteritinib (also known as ASP2215, Astellas Pharma Inc.); FF-10101-01 (Fujifilm Pharmaceuticals); HM43239 (Hanni Pharmaceuticals); Pacritinib (also known as SB1518, CTI Biopharma); MAX-40279 (Maxinovel Pty. Ltd.); FLYSYN (Synimmune GmBH); NMS-03592088 (also known as NMS-P088, Nerviano Medical Sciences); LT-171-861; and TG02 citrate (Tragara Pharmaceuticals). Also (Griswold et al., 2004; Levis et al., 2002; Levis & Small, 2004;Majothi et al., 2020; Murata et al., 2003; O'Farrell et al., 2003; BD Smith et al., 2004; Stone et al., 2005;Yee et al., See also (2002).

[0040] The aforementioned inhibitors have been or are currently being investigated in preclinical settings, in Phase I and II clinical trials as monotherapy for relapsed AML, or in Phase III combination studies for relapsed AML. While FLT3 inhibition by these compounds has been reported successfully in preclinical studies, very few patients with FLT3 mutant AML have achieved complete remission in clinical practice. Clinical responses are short-lived for the majority of patients. Response criteria for AML clinical trials are adopted from the International Working Group on AML (Cheson et al., 2003). Responders are patients who achieve complete response (CR), complete response with incomplete blood cell count recovery (CRi), or partial remission (PR). Briefly, the criteria are as follows: 1. Complete remission (CR): a. Peripheral blood count: i. No circulating blasts ii. Neutrophil count ≥ 1.0 × 10⁻⁶ 9 / L iii. Platelet count ≥ 100 × 10⁻¹⁰ 9 / L b. Bone marrow aspiration and biopsy: i. ≤ 5% blast cells ii. Absence of Auer's corpuscles iii. No extramedullary leukemia 2. Complete remission (CRi) with incomplete recovery of blood cell counts: a. Peripheral blood count: i. No circulating blasts ii. Neutrophil count < 1.0 × 10⁻⁶ 9 / L, or iii. Platelet count < 100 × 10⁻¹⁰ 9 / L b. Bone marrow aspiration and biopsy i. ≤ 5% blast cells ii. Absence of Auer's corpuscles iii. No extramedullary leukemia 3. Partial remission: a. If abnormalities are present before treatment, all CR criteria apply, except for the following: b. Myeloblasts decreased by ≥50%, but are still >5%.

[0041] To date, the clinical response to FLT3 inhibitors has been primarily limited to the clearance of peripheral blood (PB) blasts, which often returns within a few weeks, while bone marrow (BM) blasts remain largely unaffected. For example, treatment with sorafenib, a previously described multi-kinase inhibitor with activity against mutant FLT3, is effective in eliminating PB blasts, but yields only a moderate reduction in BM blasts (Borthakur et al., 2011). The BM blast percentage plays a central role in the diagnosis and classification of AML. The presence of an increasing percentage of blasts in BM is associated with significantly shorter overall survival (Amin et al., 2005; Small, 2006). To effectively treat FLT3-mutated AML patients and overcome the significant unaddressed needs in this patient population, there is a need for inhibitors that can significantly deplete both PB and BM blasts, bridge high-risk, previously well-treated patients to stem cell transplantation, reduce relapse rates in early-stage disease patients, and increase overall survival.

[0042] As used herein, the terms “proliferative impairment burden” or “proliferative disease burden” refer to the overall impact on the health of a subject or patient with cancer. This impact on the health of a subject or patient, compared to a subject without proliferative impairment or disease, may include, to name a few, a reduction in overall life expectancy, an increase in the number of years of disability due to the disease, and a decrease in wellness or overall health.

[0043] In one embodiment, the present invention relates to a compound having formula I:

[0044] [ka]

[0045] The therapeutically effective dose of the pharmaceutically acceptable salt or solvate thereof is the therapeutically effective dose for proliferative disorders selected from at least one of the following: gastrointestinal stromal tumors, leukemia, myeloma, myeloproliferative disorders, myelodysplastic syndrome, idiopathic eosinophilic syndrome (HES), bladder cancer, breast cancer, cervical cancer, CNS cancer, colon cancer, esophageal cancer, head and neck cancer, liver cancer, lung cancer, nasopharyngeal cancer, neuroendocrine cancer, ovarian cancer, pancreatic cancer, prostate cancer, kidney cancer, salivary gland cancer, small cell lung cancer, skin cancer, gastric cancer, testicular cancer, thyroid cancer, uterine cancer, and hematological malignancies. Pharmacologically acceptable salts, such as hydrochloride, phosphate, and lactate, are prepared in a manner similar to that of benzenesulfonate and are well known to those skilled in the art. The following representative compounds of the present invention, including crenolanib as crenolanib besylate, crenolanibrine, crenolanib lactate, crenolanib hydrochloride, crenolanib citrate, crenolanib acetate, crenolanib toluenesulfonate, and crenolanib succinate, are for illustrative purposes only and are not intended to limit the present invention.

[0046] The compounds of the present invention can be administered systemically to a subject, for example, orally, intravenously, subcutaneously, intramuscularly, intradermally, or parenterally. The compounds of the present invention can also be administered topically to a subject.

[0047] The compounds of the present invention can be formulated as sustained-release or immediate-release compounds for the purpose of maintaining contact with a target tissue for a desired time range.

[0048] Compositions suitable for oral administration include solid forms such as pills, tablets, caplets, capsules, granules, and powders, and liquid forms such as solutions, emulsions, and suspensions. Forms useful for parenteral administration include sterile solutions, emulsions, and suspensions.

[0049] The daily dose of the compound of the present invention may vary over a wide range of 50 to 500 mg per adult human per day. For oral administration, the composition is preferably provided in the form of tablets containing 20 and 100 milligrams. The compound of the present invention may be administered in regimens of up to three times per day or more. Preferably, it may be administered three times per day. The optimal dose to be administered may be determined by those skilled in the art and may vary with the compound of the present invention used, mode of administration, time of administration, strength of the preparation, and details of the disease state. Factors related to patient characteristics, such as age, weight, and diet, may require dose adjustment. In other examples, the daily dose of the compound of the present invention may vary over a wide range of 15 to 500, 25 to 450, 50 to 400, 100 to 350, 150 to 300, 200 to 250, 15, 25, 50, 75, 100, 150, 200, 250, 300, 400, 450, or 500 mg per day. The compounds of the present invention may be administered in a daily regimen, once, twice, three times, or more times per day. The optimal dose can be determined by those skilled in the art and may vary with the compound of the present invention used, the mode of administration, the timing of administration, the strength of the preparation, and the details of the disease state. One or more factors related to the characteristics of the subject, such as age, weight, and diet, may require dose adjustment.Techniques and compositions for developing useful dosage forms using clenolanib are described in one or more of the following references: Anderson, Philip O.; Knoben, James E.; Troutman, William G, eds., Handbook of Clinical Drug Data, Tenth Edition, McGraw-Hill, 2002; Pratt and Taylor, eds., Principles of Drug Action, Third Edition, Churchill Livingston, New York, 1990; Katzung, ed., Basic and Clinical Pharmacology, Ninth Edition, McGraw Hill, 20037ybg; Goodman and Gilman, eds., The Pharmacological Basis of Therapeutics, Tenth Edition, McGraw Hill, 2001; Remingtons Pharmaceutical Sciences, 20th Ed., Lippincott Williams & Wilkins., 2000; Martindale, The Extra Pharmacopoeia, Thirty-Second Edition (The Pharmaceutical Press, London, 1999); relevant portions are incorporated herein by reference.

[0050] The dose units for the use of crenolanib may be the single compound or a mixture thereof with other compounds, such as enhancers. The compounds may also be mixed together to form ionic or covalent bonds. All of the compounds of the present invention can be administered orally, intravenously (bolus or infusion), intraperitoneally, subcutaneously, or intramuscularly using dosage forms well known to those skilled in the pharmaceutical art. Depending on the specific site or method of delivery, different dosage forms, such as tablets, capsules, pills, powders, granules, elixirs, tinctures, suspensions, syrups, and emulsions, can be used to provide crenolanib of the present invention to patients in need of therapy.

[0051] Clenolanib is typically administered in mixture with appropriate pharmaceutically acceptable salts, buffers, diluents, expanders, excipients, and / or carriers (collectively referred to herein as pharmaceutically acceptable carriers or carrier materials), selected to be consistent with conventional pharmacovigilance based on the intended dosage form. Depending on the best site for administration, clenolanib can be formulated to provide maximum and / or consistent dosing for specific forms, for example, oral, rectal, topical, intravenous injection, or parenteral administration. Clenolanib can be administered alone, but is commonly supplied in stable salt forms mixed with pharmaceutically acceptable carriers. The carrier may be solid or liquid, depending on the type and / or site of administration.

[0052] Preparation of the compounds of the present invention. General synthetic methods that can be referenced for the preparation of the compounds of Formula I are provided in U.S. Patent No. 5,990,146 (issued November 23, 1999) (Warner-Lambert Co.), PCT Publication No. WO99 / 16755 (published April 8, 1999) (Merck & Co.), WO01 / 40217 (published July 7, 2001) (Pfizer, Inc.), U.S. Patent Application No. US2005 / 0124599 (Pfizer, Inc.), and U.S. Patent No. 7,183,414 (Pfizer, Inc.), the relevant portions of which are incorporated herein by reference.

[0053] pharmaceutically acceptable salts, such as hydrochlorides, phosphates, and lactates, are prepared in a manner similar to that of benzenesulfonates and are well known to those skilled in the art. The following representative compounds of the present invention are for illustrative purposes only and are not intended to limit the invention. [Examples]

[0054] Summary of Examples

[0055] Example A: A patient with FLT3-ITD and FLT3F691 mutation. Following disease progression during treatment with the following two prior FLT3 tyrosine kinase inhibitors, midostaurin and gilteritinib, and cytotoxic chemotherapy, the patient achieved complete clearance of leukemia blasts in the blood, bone marrow, and central nervous system (CNS) after combination therapy with clenolanib besylate.

[0056] Example B: A patient with an FLT3-ITD mutation that persisted after disease progression during previous treatment with an FLT3 tyrosine kinase inhibitor, gilteritinib, and cytotoxic chemotherapy. The patient achieved complete remission with recovery of all counts after combination therapy with clenolanib besylate.

[0057] Example C: A patient with an FLT3-ITD mutation that persisted after disease progression during treatment with two previous FLT3 tyrosine kinase inhibitors, midostaurin and gilteritinib, as well as cytotoxic chemotherapy. The patient achieved complete remission and was bridged to hematopoietic stem cell transplantation after combination therapy with clenolanib besylate.

[0058] Example D: A patient with an FLT3-ITD mutation. Following disease progression during treatment with three previous FLT3 tyrosine kinase inhibitors, sorafenib, gilteritinib, and midostaurin, as well as cytotoxic chemotherapy, the patient achieved complete remission with incomplete count recovery after combination therapy with clenolanib besylate.

[0059] Example E: A patient with FLT3-ITD, FLT3D835, and FLT3Y842 mutations. Following disease progression during previous treatment with FLT3 tyrosine kinase inhibitors, sorafenib, and cytotoxic chemotherapy, the patient achieved partial remission after treatment with clenolanib besylate monotherapy.

[0060] Example F: A patient with FLT3-ITD, FLT3D835, and FLT3N841 mutations. Following disease progression during previous treatment with FLT3 tyrosine kinase inhibitors, sorafenib, and cytotoxic chemotherapy, the patient achieved complete remission with incomplete hematological recovery after treatment with clenolanib besylate monotherapy.

[0061] Example A: Efficacy of clenolanib besylate therapy after prior administration of midostaurin and gilteritinib in relapsed / refractory patients with acquired resistance-constituting FLT3 mutations: Achievement of clearance of blood, bone marrow, and CNS leukemia blasts.

[0062] A 54-year-old woman was diagnosed with recurrent AML-positive for FLT3-ITD and FLT3-F691 missense mutation, specifically F691L. This mutation, sometimes called a "gatekeeper" mutation, is known to confuse resistance to the FLT3 tyrosine kinase inhibitor gilteritinib (McMahon et al., 2019).

[0063] In March 2019, the patient was initially diagnosed with FLT3-ITD-positive AML and was treated with a standard chemotherapy regimen plus the FLT3 inhibitor midostaurin, achieving complete remission after two cycles. Approximately six months later, in October 2019, the patient relapsed. At this point, the patient remained FLT3-positive and received a salvage combination therapy regimen including the FLT3 inhibitor gilteritinib. The patient achieved partial remission and remained stable with this regimen, but experienced disease progression in May 2020, at which point the patient enrolled in a phase 1 clinical trial of a menin inhibitor. One month into the trial, the patient experienced significant disease progression, including CNS involvement, and was withdrawn from the trial.

[0064] As of June 2020, the patient had received three previous therapies, including two FLT3 inhibitors. Molecular testing revealed that after gilteritinib treatment, the patient acquired the FLT3 mutation F691L, which conferred secondary resistance. The persistence of the FLT3-ITD mutation, the acquisition of the F691L mutation, and the fact that the patient was relapsed / refractory to multiple FLT3 inhibitors placed this patient in a particularly high-risk group with a reduced likelihood of treatment response and shortened overall survival. (Perl et al., 2021)

[0065] As there were no other available approved standard treatment options, the treating physician applied for special use authorization for clenolanib besilate, which was granted in June 2020. The patient was treated with salvage chemotherapy consisting of high doses of cytarabine and clenolanib besilate, 80 mg three times daily. At the start of treatment, the patient had 72% myeloblasts, 23% peripheral blasts, and 88% blasts (of all nucleated cells) in the cerebrospinal fluid (CSF), indicating significant CNS involvement in these leukemias.

[0066] Bone marrow, peripheral blood, and CSF samples taken on day 21 of treatment revealed complete clearance of leukemic blasts from all three compartments. Molecular testing revealed clearance of all FLT3 clones in the bone marrow. A second bone marrow biopsy taken on day 43 of treatment confirmed that the patient remained morphologically leukemia-free (complete remission without recovery of counts) and CNS leukemia-free. The patient received treatment for 90 days and died in remission from sepsis 3.5 months after initiating crenolanib therapy.

[0067] Table A below illustrates the ability of clenolanib to eliminate malignant leukemic blasts from the bone marrow, peripheral blood, and CSF of patients with FLT3 mutations that confer resistance after treatment with previous tyrosine kinase inhibitors. [Table 1]

[0068] Example B: Efficacy of clenolanib besylate therapy in relapsed / refractory patients who were refractory to gilteritinib treatment: Achievement of complete remission with recovery of all counts.

[0069] A 35-year-old woman was diagnosed with relapsed AML-positive due to an FLT3-ITD mutation. In March 2018, the patient was initially diagnosed with FLT3-ITD-positive AML and achieved complete remission with a standard chemotherapy regimen. Approximately three months later, in June 2018, the patient relapsed and received the FLT3 inhibitor gilteritinib as salvage therapy. The patient was refractory to this treatment, and after two cycles of gilteritinib therapy, 20% myeloblasts persisted. The patient also experienced pericarditis as a side effect of gilteritinib treatment. After discontinuation of gilteritinib, the patient received standard salvage chemotherapy, achieved a second remission, and underwent a hematopoietic stem cell transplant in November 2018. Approximately 17 months later, the patient's FLT3-ITD-positive disease relapsed.

[0070] In April 2020, the patient was receiving three previous lines of therapy, including the FLT3 inhibitor gilteritinib. Molecular testing revealed that the patient's initial FLT3-ITD mutation persisted. The persistence of this mutation and the fact that the patient experienced disease relapse after HSCT placed this patient in a particularly high-risk group, associated with a reduced likelihood of treatment response and shortened overall survival (Bejanyan et al., 2015).

[0071] Due to limited treatment options, the treating physician applied for special use authorization for clenolanib besilate, which was granted in April 2020. The patient was treated with salvage chemotherapy consisting of fludarabine, cytarabine, idarubicin, and granulocyte colony-stimulating factor, followed by clenolanib besilate, 100 mg three times daily. At the start of treatment, the patient had 75% myeloblasts and 85% peripheral blasts, and diagnostic imaging showed extramedullary disease (leukemic blasts outside the bone marrow or blood) in the spleen and lymph nodes.

[0072] A bone marrow biopsy performed on day 36 of treatment revealed complete clearance of peripheral blasts, clearance of less than 5% of myeloblasts, and recovery of neutrophils and platelets, which was classified as complete remission with recovery of the total count. Molecular testing revealed clearance of all FLT3 clones. A second bone marrow biopsy performed on day 81 of treatment confirmed that the patient remained in complete remission and had complete clearance of all extramedullary disease. The patient remained in remission for four months during treatment with crenolanib besylate.

[0073] Table B below illustrates the ability of clenolanib to eliminate malignant leukemic blasts from the bone marrow and peripheral blood of patients with relapsed / refractory disease following prior tyrosine kinase inhibitor treatment. [Table 2]

[0074] Example C: Efficacy of clenolanib besylate therapy in relapsed / refractory patients with FLT3-ITD mutations following prior gilteritinib administration: achievement of complete remission with recovery of all counts and bridging to transplantation.

[0075] A 22-year-old woman was diagnosed with relapsed AML-positive for FLT3-ITD mutation. In November 2019, the patient was initially diagnosed with AML with FLT3-ITD mutation and was treated with a standard chemotherapy regimen including cytarabine and daunorubicin plus the FLT3 inhibitor midostaurin, achieving complete remission after two cycles. However, the patient remained measurable residual disease (MRD) and FLT3-positive. Subsequently, the patient received intensive high-dose cytarabine therapy in combination with midostaurin, but MRD and FLT3-ITD mutation persisted. In May 2020, because the FLT3-ITD mutation was still detectable in a bone marrow sample obtained from the patient, and because residual FLT3-ITD-positive blasts could potentially cause relapse, the patient was administered the FLT3 inhibitor gilteritinib with the aim of eliminating them. A bone marrow biopsy performed four weeks after monotherapy with gilteritinib revealed that the patient had relapsed, with 40% myeloblasts, and gilteritinib was discontinued.

[0076] In June 2020, the patient was receiving two previous therapies, including two FLT3 inhibitors. Molecular testing confirmed that the FLT3-ITD mutation present at diagnosis persisted through all therapies (more detailed sequencing, including tests to identify the presence of resistance-constituting point mutations, was not performed). The persistence of the FLT3-ITD mutation and the patient's relapse / refractory nature to multiple FLT3 inhibitors placed this patient in a particularly high-risk group, associated with reduced likelihood of treatment response and shortened overall survival (Perl et al., 2021).

[0077] As there were no other available approved standard treatment options, the treating physician applied for special use authorization for clenolanib besilate, which was granted in June 2020. The patient was treated with salvage chemotherapy consisting of fludarabine, cytarabine, idarubicin, and granulocyte colony-stimulating factor, followed by treatment with clenolanib besilate, 100 mg three times daily. At the start of treatment, the patient had 43% myeloblasts.

[0078] A bone marrow biopsy taken on day 56 of treatment revealed that the percentage of myeloblasts had dropped to less than 5%, and the patient underwent a hematopoietic stem cell transplant. Subsequently, the patient received monotherapy with clenolanib besylate as post-transplant maintenance, starting on day 49 post-transplant, with the aim of preventing further relapses. A bone marrow biopsy performed on day 30 post-transplant, prior to the initiation of clenolanib maintenance therapy, showed that the patient remained in remission, but the FLT3-ITD mutation was still detectable. A second bone marrow biopsy performed on day 68 post-transplant, 19 days after the initiation of clenolanib maintenance, revealed that the FLT3-ITD mutation had been eliminated.

[0079] Table C below illustrates the ability of clenolanib to eliminate malignant leukemic blasts from the bone marrow of patients who were relapsed / refractory to two previous tyrosine kinase inhibitors, and to eliminate FLT3-ITD MRD after hematopoietic stem cell transplantation. [Table 3]

[0080] Example D: Efficacy of clenolanib besylate therapy in relapsed / refractory patients with FLT3-ITD mutations following prior administration of sorafenib, gilteritinib, and midostaurin: Achievement of complete remission with incomplete count recovery.

[0081] A 76-year-old male was diagnosed with relapsed AML-positive due to an FLT3-ITD mutation. In 2008, the patient was initially diagnosed with myelodysplastic syndrome, which was converted to FLT3-ITD-mutated AML in August 2015. At the time of the conversion to AML, the patient was treated with a standard chemotherapy regimen, achieved complete remission, and progressed to hematopoietic stem cell transplantation. Approximately nine months after transplantation, in August 2016, the patient's disease relapsed due to CNS involvement in his leukemia. He was treated with cytarabine, methotrexate, and the FLT3 inhibitor sorafenib, and achieved remission again. Two and a half years later, in April 2019, the patient's disease relapsed again due to CNS involvement. The patient was treated with venetoclax, donor lymphocyte infusion (a common rescue method for patients who have relapsed after hematopoietic stem cell transplantation, in which leukocytes from the original transplant donor are infused into the recipient), and the FLT3 inhibitor gilteritinib, and achieved complete remission again. Seven months later, in November 2019, the patient relapsed again due to CNS involvement and received cytarabine, cladribine, and a third FLT3 inhibitor, midostaurin, achieving complete remission again. Eight months later, in July 2020, the patient experienced a fourth relapse with 16% myeloblasts, and the original FLT3-ITD mutation persisted.

[0082] As of July 2020, the patient had received four previous therapies, including three FLT3 inhibitors. Molecular testing confirmed that the FLT3-ITD mutation present at diagnosis persisted through all therapies (more detailed sequencing, including tests to identify the presence of resistance-constituting point mutations, was not performed). The persistence of the FLT3-ITD mutation and the patient's relapse / refractory nature to multiple FLT3 inhibitors placed this patient in a particularly high-risk group, associated with reduced likelihood of treatment response and shortened overall survival (Perl et al., 2021).

[0083] As there were no other available approved standard treatment options, the treating physician applied for special use authorization for clenolanib besilate, which was granted in July 2020. The patient was treated with salvage chemotherapy consisting of fludarabine, cytarabine, idarubicin, and granulocyte colony-stimulating factor, followed by treatment with clenolanib besilate, 100 mg three times daily. At the start of treatment, the patient had 16% myeloblasts.

[0084] A bone marrow biopsy performed on day 21 of treatment revealed a recovery in neutrophil counts and clearance of less than 5% of myeloblasts, which was classified as complete remission with incomplete hematological recovery. At this point, the FLT3-ITD mutation was also ruled out. Due to the patient's advanced age, regular bone marrow biopsies were not performed, and the patient continued crenolanib treatment for approximately six months.

[0085] Patient Example D illustrates the ability of clenolanib to eliminate leukemic blasts from the bone marrow of a patient who was relapsed / refractory to three previous FLT3 tyrosine kinase inhibitors.

[0086] Example E: Efficacy of clenolanib besylate monotherapy in relapsed / refractory patients with FLT3 mutations that confer acquired resistance after previous sorafenib administration: Achievement of partial remission.

[0087] An 87-year-old woman was diagnosed with recurrent AML-positive for FLT3-ITD and FLT3-D835 and Y842 missense mutations, specifically D835Y and Y842C. These mutations are known to confuse resistance to the FLT3 tyrosine kinase inhibitors sorafenib and quizartinib (Wang et al., 2021).

[0088] In August 2013, the patient was initially diagnosed with FLT3-ITD AML and treated with a standard chemotherapy regimen, achieving complete remission after two cycles. To prevent relapse, the patient was given sorafenib as maintenance therapy. Five months later, in March 2014, the patient experienced a relapse.

[0089] At this point, molecular studies revealed the persistence of the original FLT3-ITD mutation, as well as the acquisition of FLT3 mutations D835Y and Y842C that confer secondary resistance, following treatment with sorafenib. The persistence of the FLT3-ITD mutation, the acquisition of D835Y and Y842C mutations, and the fact that patients experienced disease relapse in response to previous FLT3 inhibitors placed them in a particularly high-risk group with reduced likelihood of treatment response and shortened overall survival (Perl et al., 2021).

[0090] Because there were no other available approved standard treatment options, the patient enrolled in a clinical trial of clenolanib besylate monotherapy administered at a dose of 100 mg three times daily (NCT01657682). At trial enrollment, the patient had 68% myeloblasts and 30% peripheral blasts.

[0091] A bone marrow biopsy performed on day 27 of treatment revealed that the patient's myeloblast count had decreased to 7% and peripheral blasts had been eliminated, which was classified as partial remission. Sadly, the patient died on day 61 of treatment from leukemia-related complications before further bone marrow biopsies could be performed.

[0092] Patient Example E illustrates the ability of clenolanib to significantly reduce malignant leukemic blasts from 30% to 7% in the bone marrow of a patient with FLT3 mutations conferring two types of resistance after prior tyrosine kinase inhibitor treatment, thereby completely eliminating malignant leukemic blasts in the peripheral blood.

[0093] Example F: Efficacy of clenolanib besylate monotherapy in relapsed / refractory patients with acquired resistance-contributing FLT3 mutations following previous sorafenib administration: Achievement of complete remission with incomplete hematological recovery.

[0094] A 31-year-old male was diagnosed with AML-positive, recurrent / refractory to FLT3-ITD and FLT3-D835 and N841 missense mutations, specifically D835V, D835Y, D835H, and N841K. These mutations are known to confuse resistance to the FLT3 tyrosine kinase inhibitors sorafenib and quizartinib (Wang et al., 2021).

[0095] In November 2012, the patient was initially diagnosed with FLT3-ITD AML and was treated with a standard chemotherapy regimen, achieving complete remission and proceeding to hematopoietic stem cell transplantation. Six months after transplantation, in November 2013, the patient relapsed and was treated with sorafenib and decitabine as salvage therapy, but after several cycles, did not respond to treatment.

[0096] In May 2014, molecular studies revealed the persistence of the original FLT3-ITD mutation, as well as the acquisition of multiple secondary resistance-constituting FLT3 mutations: D835V, D835Y, D835H, and N841K, after sorafenib treatment. The persistence of the FLT3-ITD mutation, the acquisition of the D835 and N841 mutations, and the fact that the patient experienced disease relapse in response to previous FLT3 inhibitors placed this patient in a particularly high-risk group, associated with reduced likelihood of treatment response and shortened overall survival. (Perl et al., 2021)

[0097] Because there were no other available, approved, standard treatment options, the patient enrolled in a clinical trial of clenolanib besylate monotherapy administered at a dose of 100 mg three times daily (NCT01657682). At trial enrollment, the patient had 84% myeloblasts and 96% peripheral blasts.

[0098] A bone marrow biopsy performed on day 29 of treatment revealed that the patient's myeloblast count had dropped to 23%, along with peripheral blast clearance, which was classified as partial remission. A second bone marrow biopsy performed on day 57 of treatment revealed that the patient's myeloblast count had dropped to 7%, which was still classified as partial remission. A third bone marrow biopsy performed on day 84 of treatment revealed that the patient's myeloblast count had dropped to less than 5%, along with neutrophil recovery, which was classified as complete remission with incomplete hematological recovery.

[0099] Table F below illustrates the ability of clenolanib to eliminate malignant leukemia blasts from the bone marrow of patients with FLT3 mutations that confer resistance after previous tyrosine kinase inhibitor treatment. [Table 4]

[0100] Any embodiment discussed herein may be introduced in connection with any method, kit, reagent, or composition of the present invention, and vice versa. Furthermore, the methods of the present invention can be achieved using the compositions of the present invention.

[0101] It should be understood that the specific embodiments described herein are illustrative and not intended to be limitations of the invention. The main features of the invention can be utilized in various embodiments without departing from the scope of the invention. Those skilled in the art will recognize or determine many equivalents to the specific procedures described herein simply by using the given experiments. Such equivalents are considered to be within the scope of the invention and are covered by the claims.

[0102] All publications and patent applications described herein represent the skill level of a person skilled in the art to which the invention relates. All publications and patent applications are incorporated herein by reference to the same extent as each individual publication or patent application is specifically and individually indicated as being incorporated by reference.

[0103] In the claims and / or specification, the use of the words “a” or “an” in conjunction with the term “including” may mean “one,” which also coincides with the meanings of “one or more,” “at least one,” and “one or more.” In the claims, the use of the term “or” is used to mean “and / or,” unless it is expressly indicated that it refers only to substitutes or that the substitutes are mutually exclusive, however this disclosure refers only to substitutes and supports the definition of “and / or.” Throughout this application, the term “about” is used to suggest that the value includes the specific error variability of the device, the method used to determine the value, or the variability present in the subject of study.

[0104] Where used herein and in the claims, the word “comprising” (and any form of including, e.g., “comprise” and “comprises”), “having” (and any form of having, e.g., “have” and “has”), “including” (and any form of including, e.g., “includes” and “include”), or “containing” (and any form of containing, e.g., “contains” and “contain”) is comprehensive or open-ended and does not exclude additional, unlisted features, elements, components, groups, integers, and / or processes, nor does it exclude the presence of other undescribed features, elements, components, groups, integers, and / or processes. In any embodiment of the compositions and methods provided herein, “comprising” may be replaced with “consisting essentially of” or “consisting of”. As used herein, the term “consisting” is used only to indicate the presence of an enumerated integer (e.g., feature, element, characteristic, method / process step or limitation) or a group of integers (e.g., feature(pl), element(pl), feature(pl), characteristic(pl), method / process step or limitation(pl))). As used herein, the phrase “consisting essentially of” requires the identified feature, element, component, group, integer, and / or step, but does not exclude the presence of other undescribed features, elements, components, base, integer, and / or step, as well as any fundamental and novel features(pl) and / or functions of the claimed invention that do not materially affect them.

[0105] The term “or combination thereof,” as used herein, refers to all permutations and combinations of the items listed prior to the term. For example, “A, B, C, or combination thereof” is intended to include at least one of A, B, C, AB, AC, BC, or ABC, and, where the order is important in a particular context, BA, CA, CB, CBA, BCA, ACB, BAC, or CAB. Continuing with this example, explicitly included are combinations containing repetitions of one or more items or terms, such as BB, AAA, AB, BBC, AAABCCCC, CBBAAA, CABABB, etc. Those skilled in the art will understand that, unless evident from the context, there is usually no limit to the number of items or terms in any combination.

[0106] Where used herein, approximation words, such as “about,” “substantial,” or “substantially,” without limitation, refer to a state that, when thus modified, is not necessarily absolute or perfect, but is considered by a person skilled in the art to be nearly sufficiently justifiable to describe as existing. The range by which the description may vary depends on how significant the change may be while still causing a person skilled in the art to perceive that the modified feature still possesses the required features and capabilities of the unmodified feature. Generally, but subject to the preceding considerations, numerical values ​​herein modified with approximation words, such as “about,” may vary from the stated value by at least ±0.1, 0.5, 1, 2, 3, 4, 5, 6, 7, 10, 12, or 15%, or are considered to be within the normal tolerances of the art, e.g., within two standard deviations of the mean. Unless otherwise evident from the context, all numerical values ​​provided herein are modified with the term “about.”

[0107] Furthermore, the section headings in this specification are provided to maintain consistency with the proposals of 37 CFR 1.77, or otherwise to provide organizational clues. These headings are not intended to limit or characterize the invention(s) presented in any claims that may arise from this disclosure. Specifically and as an example, although the headings refer to “Field of Invention,” such claims should not be limited by the language under this heading that describes the so-called technical field. Furthermore, the description of the technology in the “Background of Invention” section should not be construed as an acknowledgment that the technology is prior art to any invention(s) in this disclosure. The “Summary of Invention” should also not be considered a characterization of the invention(s) described in the granted claims. Furthermore, any reference to “Invention” in this disclosure, even singular, should not be used to assert that there is only one point of novelty in this disclosure. Multiple inventions may be described in accordance with the limits of multiple claims arising from this disclosure, and such claims should therefore define the invention(s) and their equivalents that are thereby protected. In all cases, such claims should be considered in light of the present disclosure for their own merits and should not be limited by the headings set forth herein.

[0108] All compositions and / or methods disclosed and claimed herein can be prepared and performed in light of this disclosure without excessive experimentation. While the compositions and methods of the present invention are described as preferred embodiments, it will be apparent to those skilled in the art that variations may be applied to the compositions and / or methods and the steps or sequences of steps described herein without departing from the concept, spirit and scope of the invention. All such similar substitutions and variations, which will be apparent to those skilled in the art, are considered to fall within the spirit, scope and concept of the invention as defined by the appended claims.

[0109] To help the Patent Office and any reader of any patent arising from this application interpret the claims accompanying this specification, the applicants would like to note that, unless the words “means for” or “process for” are expressly used in a particular claim, none of the attached claims are intended to exercise equivalents as existing in 35 U.S. SC § 112, paragraph 6, U.S.C § 112, paragraph (f), or as of the filing date of that application.

[0110] For each of the claims, each dependent claim may be dependent on both the independent claim and each of the prior dependent claims, insofar as the prior claims provide a proper precedent base for the terms or elements of the claims.

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[0122] Joshi, S. K., Sharzehi, S., Pittsenbarger, J., Bottomly, D., Tognon, C. E., McWeeney, S. K., . . . Traer, E. (2021). A noncanonical FLT3 gatekeeper mutation disrupts gilteritinib binding and confers resistance. Am J Hematol, 96(7), E226-E229. doi:10.1002 / ajh.26174

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[0126] Majothi, S., Adams, D., Loke, J., Stevens, S. P., Wheatley, K., & Wilson, J. S. (2020). FLT3 inhibitors in acute myeloid leukaemia: assessment of clinical effectiveness, adverse events and future research-a systematic review and meta-analysis. Syst Rev, 9(1), 285. doi:10.1186 / s13643-020-01540-1

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[0128] Murata, K., Kumagai, H., Kawashima, T., Tamitsu, K., Irie, M., Nakajima, H., . . . Kitamura, T. (2003). Selective cytotoxic mechanism of GTP-14564, a novel tyrosine kinase inhibitor in leukemia cells expressing a constitutively active Fms-like tyrosine kinase 3 (FLT3). J Biol Chem, 278(35), 32892-32898. doi:10.1074 / jbc.M210405200

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[0130] Papaemmanuil, E., Gerstung, M., Bullinger, L., Gaidzik, V. I., Paschka, P., Roberts, N. D., . . . Campbell, P. J. (2016). Genomic Classification and Prognosis in Acute Myeloid Leukemia. N Engl J Med, 374(23), 2209-2221. doi:10.1056 / NEJMoa1516192

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[0132] Perl, A. E., Martinelli, G., Cortes, J. E., Neubauer, A., Berman, E., Paolini, S., . . . Levis, M. J. (2019). Gilteritinib or Chemotherapy for Relapsed or Refractory FLT3-Mutated AML. New England Journal of Medicine, 381(18), 1728-1740. doi:10.1056 / NEJMoa1902688

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[0134] Sakaguchi, M., Yamaguchi, H., Kuboyama, M., Najima, Y., Usuki, K., Ueki, T., . . . Inokuchi, K. (2019). Significance of FLT3-tyrosine kinase domain mutation as a prognostic factor for acute myeloid leukemia. Int J Hematol. doi:10.1007 / s12185-019-02720-z

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[0138] Smith, C. C., Lasater, E. A., Lin, K. C., Wang, Q., McCreery, M. Q., Stewart, W. K., . . . Shah, N. P. (2014). Crenolanib is a selective type I pan-FLT3 inhibitor. Proc Natl Acad Sci U S A, 111(14), 5319-5324. doi:10.1073 / pnas.1320661111

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Claims

1. A pharmaceutical composition for treating proliferative disorders in subjects having mutated or constitutively active FLT3, which are recurrent or refractory to one or more prior tyrosine kinase inhibitors, wherein the treatment comprises the following steps: A step of obtaining a tumor sample from a subject that is recurrent or refractory to one or more prior tyrosine kinase inhibitors selected from midostaurin, gilteritinib, and quizartinib; A step of measuring the expression of mutated or constitutively active FLT3 mutants in the tumor sample; and The process of administering to the subject a therapeutically effective dose of crenolanib or a pharmaceutically acceptable salt thereof sufficient to treat the proliferative disorder. A pharmaceutical composition containing the above.

2. The pharmaceutical composition according to claim 1, wherein the mutated or constitutively active FLT3 is at least one of the following: FLT3-ITD; FLT3-TKD; activating mutation in FLT3; copy number increase or amplification of the FLT3 gene; or gene fusion including fusion of FLT3 with another gene.

3. The pharmaceutical composition according to claim 1, wherein the subject has an FLT3 mutation that confers resistance to the aforementioned prior tyrosine kinase inhibitor.

4. The pharmaceutical composition according to claim 3, wherein the resistance-constituting FLT3 mutation is selected from missense mutations occurring alone or in combination with the FLT3-ITD mutation in at least one of the amino acid residues K429, A627, N676, A680, F691, Y693, G697, D698, N701, D835, N841, Y842, and A848.

5. The pharmaceutical composition according to claim 3, wherein the resistance-constituting FLT3 mutation was present before administration of the previous tyrosine kinase inhibitor, or the resistance-constituting FLT3 mutation was acquired during or after administration of the previous tyrosine kinase inhibitor.

6. The pharmaceutical composition according to claim 1, wherein the proliferative disorder is selected from at least one of the following: gastrointestinal stromal tumor, leukemia, myeloma, myeloproliferative disorder, myelodysplastic syndrome, idiopathic eosinophilic syndrome (HES), bladder cancer, breast cancer, cervical cancer, CNS cancer, colon cancer, esophageal cancer, head and neck cancer, liver cancer, lung cancer, nasopharyngeal cancer, neuroendocrine cancer, ovarian cancer, pancreatic cancer, prostate cancer, kidney cancer, salivary gland cancer, small cell lung cancer, skin cancer, gastric cancer, testicular cancer, thyroid cancer, uterine cancer, and hematological malignancies.

7. The pharmaceutical composition according to claim 1, wherein the therapeutically effective dose of crenolanib or a pharmaceutically acceptable salt thereof is approximately 50 to 500 mg per day, 100 to 450 mg per day, 200 to 400 mg per day, 300 to 500 mg per day, 350 to 500 mg per day, or 400 to 500 mg per day.

8. The pharmaceutical composition according to claim 1, characterized in that a therapeutically effective dose of crenolanib or a pharmaceutically acceptable salt thereof is administered in at least one of the following ways: continuous, intermittent, systemic, or topical.

9. The pharmaceutical composition according to claim 1, for administration orally, intravenously, or intraperitoneally.

10. The pharmaceutical composition according to claim 1, characterized in that a therapeutically effective amount of crenolanib or a pharmaceutically acceptable salt thereof is administered up to three times a day or more, insofar as the subject requires treatment for a proliferative disorder.

11. A therapeutically effective dose of crenolanib or a pharmaceutically acceptable salt thereof is To maintain remission in existing patients, it is provided sequentially or concurrently with at least one other drug; To maintain remission in patients with relapsing or refractory proliferative disorders, it is provided to the patient as monotherapy or in combination with another drug; or Provided as monotherapy or in combination with other drugs to maintain remission in pediatric patients with relapsing or refractory proliferative disorders. The pharmaceutical composition according to claim 1, characterized in that...

12. The pharmaceutical composition according to claim 1, wherein the clenolanib or a pharmaceutically acceptable salt thereof is clenolanib besylate, clenolanibrinate, clenolanib lactate, clenolanib hydrochloride, clenolanib citrate, clenolanib acetate, clenolanib toluenesulfonate, or clenolanib succinate.

13. A pharmaceutical composition comprising crenolanib or a pharmaceutically acceptable salt thereof as the active ingredient for inhibiting or reducing mutant FLT3 tyrosine kinase activity or expression in subjects suffering from proliferative disorders, wherein the inhibition or reduction is achieved through the following steps: A step of identifying subjects who have discontinued previous tyrosine kinase inhibitor therapy due to a refractory or recurrent proliferative disorder, wherein the previous tyrosine kinase inhibitor is selected from midostaurin, gilteritinib, and quizartinib; A step of obtaining a tumor sample from the aforementioned target; A step of measuring the expression of mutated FLT3 or constitutively active FLT3 mutants in the tumor sample; and If the subject has the mutated FLT3 or a constitutively active FLT3 mutant, the procedure includes administering to the subject a therapeutically effective dose of crenolanib or a pharmaceutically acceptable salt thereof. A pharmaceutical composition comprising the crenolanib or a salt thereof, which reduces the proliferative impairment burden or prevents the progression of a proliferative disorder.

14. The pharmaceutical composition according to claim 13, wherein the mutated or constitutively active FLT3 is at least one of the following: FLT3-ITD; FLT3-TKD; activating mutation in FLT3; copy number increase or amplification of the FLT3 gene; or gene fusion including fusion of FLT3 with another gene.

15. The pharmaceutical composition according to claim 13, wherein the subject has an FLT3 mutation that confers resistance to the aforementioned prior tyrosine kinase inhibitor.

16. The pharmaceutical composition according to claim 13, wherein the subject is recurrent or unresponsive to a previous tyrosine inhibitor, and the subject has a resistance-constituting FLT3 mutation selected from a missense mutation occurring alone or in combination with an FLT3-ITD mutation in at least one of the amino acid residues K429, A627, N676, A680, F691, Y693, G697, D698, N701, D835, N841, Y842, A848.

17. The pharmaceutical composition according to claim 16, wherein the resistance-constituting FLT3 mutation was present before administration of the previous tyrosine kinase inhibitor, or the resistance-constituting FLT3 mutation was acquired during or after administration of the previous tyrosine kinase inhibitor.

18. The pharmaceutical composition according to claim 13, wherein the proliferative disorder is selected from at least one of the following: gastrointestinal stromal tumors, leukemia, myeloma, myeloproliferative disorders, myelodysplastic syndrome, idiopathic eosinophilic syndrome (HES), bladder cancer, breast cancer, cervical cancer, CNS cancer, colon cancer, esophageal cancer, head and neck cancer, liver cancer, lung cancer, nasopharyngeal cancer, neuroendocrine cancer, ovarian cancer, pancreatic cancer, prostate cancer, kidney cancer, salivary gland cancer, small cell lung cancer, skin cancer, gastric cancer, testicular cancer, thyroid cancer, uterine cancer, and hematological malignancies.

19. The pharmaceutical composition according to claim 13, wherein the therapeutically effective dose of crenolanib or a pharmaceutically acceptable salt thereof is approximately 50 to 500 mg per day, 100 to 450 mg per day, 200 to 400 mg per day, 300 to 500 mg per day, 350 to 500 mg per day, or 400 to 500 mg per day.

20. The pharmaceutical composition according to claim 13, characterized in that a therapeutically effective dose of crenolanib or a pharmaceutically acceptable salt thereof is administered in at least one of the following ways: continuous, intermittent, systemic, or topical.

21. The pharmaceutical composition according to claim 13, for administration orally, intravenously, or intraperitoneally.

22. The pharmaceutical composition according to claim 13, characterized in that a therapeutically effective amount of crenolanib or a pharmaceutically acceptable salt thereof is administered up to three times a day or more, insofar as the subject requires treatment for a proliferative disorder.

23. A therapeutically effective dose of crenolanib or a pharmaceutically acceptable salt thereof is To maintain remission in existing patients, it is provided sequentially or concurrently with at least one other drug; To maintain remission in patients with relapsing or refractory proliferative disorders, it is provided to the patient as monotherapy or in combination with another drug; or Provided as monotherapy or in combination with other drugs to maintain remission in pediatric patients with relapsing or refractory proliferative disorders. The pharmaceutical composition according to claim 13, characterized in that...

24. The pharmaceutical composition according to claim 13, wherein the clenolanib or a pharmaceutically acceptable salt thereof is clenolanib besylate, clenolanibrinate, clenolanib lactate, clenolanib hydrochloride, clenolanib citrate, clenolanib acetate, clenolanib toluenesulfonate, or clenolanib succinate.

25. A pharmaceutical composition for treating a subject suffering from a proliferative disorder, comprising crenolanib or a pharmaceutically acceptable salt thereof as the active ingredient, wherein the treatment comprises the following steps: To determine whether a patient has a gene mutation in the FLT3 gene, alterations in the kinase activity of FLT3 tyrosine kinase, overexpression of FLT3 tyrosine kinase, or alterations in the phenotype or genotype of FLT3 tyrosine kinase, By obtaining or having obtained a biological sample from the aforementioned patient; and By performing or having performed an assay on the aforementioned biological sample, A step to determine whether the subject increased FLT3 tyrosine kinase activity; The step of treating the patient with a first tyrosine kinase inhibitor selected from midostaurin, gilteritinib, and quizartinib; and A pharmaceutical composition comprising the step of discontinuing the administration of the first tyrosine kinase inhibitor and administering an effective dose of crenolanib to the patient if the patient is refractory or relapses after administration of the first tyrosine kinase inhibitor, and if the patient has a gene mutation in FLT3; a change in the kinase activity of FLT3, overexpression of FLT3, or a change in the phenotypic characteristics of the FLT3 tyrosine kinase genotype.

26. The pharmaceutical composition according to claim 25, wherein the mutated or constitutively active FLT3 is at least one of the following: FLT3-ITD; FLT3-TKD; activating mutation in FLT3; copy number increase or amplification of the FLT3 gene; or gene fusion including fusion of FLT3 with another gene.

27. The pharmaceutical composition according to claim 25, wherein the subject has an FLT3 mutation that confers resistance to a previous tyrosine kinase inhibitor.

28. The pharmaceutical composition according to claim 25, wherein the mutation in the FLT3 gene or the change in the phenotype or genotype of FLT3 is a mutation that confers resistance, selected from missense mutations occurring alone or in combination with the FLT3-ITD mutation in at least one of the amino acid residues K429, A627, N676, A680, F691, Y693, G697, D698, N701, D835, N841, Y842, and A848.

29. The pharmaceutical composition according to claim 28, wherein the resistance-constituting FLT3 mutation was present before administration of the previous tyrosine kinase inhibitor, or the resistance-constituting FLT3 mutation was acquired during or after administration of the previous tyrosine kinase inhibitor.

30. The pharmaceutical composition according to claim 25, wherein the proliferative disorder is selected from at least one of the following: gastrointestinal stromal tumor, leukemia, myeloma, myeloproliferative disorder, myelodysplastic syndrome, idiopathic eosinophilic syndrome (HES), bladder cancer, breast cancer, cervical cancer, CNS cancer, colon cancer, esophageal cancer, head and neck cancer, liver cancer, lung cancer, nasopharyngeal cancer, neuroendocrine cancer, ovarian cancer, pancreatic cancer, prostate cancer, kidney cancer, salivary gland cancer, small cell lung cancer, skin cancer, gastric cancer, testicular cancer, thyroid cancer, uterine cancer, and hematological malignancies.

31. The pharmaceutical composition according to claim 25, wherein the therapeutically effective dose of crenolanib or a pharmaceutically acceptable salt thereof is approximately 50 to 500 mg per day, 100 to 450 mg per day, 200 to 400 mg per day, 300 to 500 mg per day, 350 to 500 mg per day, or 400 to 500 mg per day.

32. The pharmaceutical composition according to claim 25, characterized in that a therapeutically effective amount of crenolanib or a pharmaceutically acceptable salt thereof is administered in at least one of the following ways: continuous, intermittent, systemic, or topical.

33. The pharmaceutical composition according to claim 25 for oral, intravenous, or intraperitoneal administration.

34. The pharmaceutical composition according to claim 25, characterized in that a therapeutically effective amount of crenolanib or a pharmaceutically acceptable salt thereof is administered up to three times a day or more, insofar as the subject requires treatment for a proliferative disorder.

35. A therapeutically effective dose of crenolanib or a pharmaceutically acceptable salt thereof is To maintain remission in existing patients, it is provided sequentially or concurrently with at least one other drug; To maintain remission in patients with relapsing or refractory proliferative disorders, it is provided to the patient as monotherapy or in combination with another drug; or Provided as monotherapy or in combination with other drugs to maintain remission in pediatric patients with relapsing or refractory proliferative disorders. The pharmaceutical composition according to claim 25, characterized in that...

36. The pharmaceutical composition according to claim 25, wherein the clenolanib or a pharmaceutically acceptable salt thereof is clenolanib besylate, clenolanibrinate, clenolanib lactate, clenolanib hydrochloride, clenolanib citrate, clenolanib acetate, clenolanib toluenesulfonate, or clenolanib succinate.

37. A pharmaceutical composition for treating a subject suffering from leukemia, comprising crenolanib or a pharmaceutically acceptable salt thereof as the active ingredient, wherein the treatment comprises the following steps: A step of obtaining a sample from the aforementioned target; A step of determining from the aforementioned sample whether the patient has a disregulated FLT3 receptor or a constitutively active FLT3 receptor; A further step of determining whether the subject is refractory to administration or has relapsed after administration of a prior tyrosine kinase inhibitor selected from midostaurin, gilteritinib, and quizaltinib; and The process of administering to the subject requiring such treatment a therapeutically effective dose of crenolanib or a pharmaceutically acceptable salt thereof sufficient to treat leukemia, A pharmaceutical composition containing the above.

38. The pharmaceutical composition according to claim 37, wherein the mutated or constitutively active FLT3 is at least one of the following: FLT3-ITD; FLT3-TKD; activating mutation in FLT3; copy number increase or amplification of the FLT3 gene; or gene fusion including fusion of FLT3 with another gene.

39. The pharmaceutical composition according to claim 37, wherein the subject has an FLT3 mutation that confers resistance to a previous tyrosine kinase inhibitor.

40. The pharmaceutical composition according to claim 37, wherein the mutation in the FLT3 gene or the change in the phenotype or genotype of FLT3 is a mutation that confers resistance, selected from missense mutations occurring alone or in combination with the FLT3-ITD mutation in at least one of the amino acid residues K429, A627, N676, A680, F691, Y693, G697, D698, N701, D835, N841, Y842, and A848.

41. The pharmaceutical composition according to claim 37, wherein the resistance-constituting FLT3 mutation was present before administration of the previous tyrosine kinase inhibitor, or the resistance-constituting FLT3 mutation was acquired during or after administration of the previous tyrosine kinase inhibitor.

42. The pharmaceutical composition according to claim 37, wherein the leukemia is selected from at least one of Hodgkin's disease, myeloma, acute promyelocytic leukemia (APL), chronic lymphocytic leukemia (CLL), chronic myeloid leukemia (CML), chronic neutrophilic leukemia (CNL); acute anaplastic leukemia (AUL), anaplastic large cell lymphoma (ALCL), prolymphoplastic leukemia (PML); juvenile myelomonocytic leukemia (JMML); adult T-cell ALL, acute myeloid leukemia (AML), AML with tricytic myelodysplasia, myelodysplastic syndrome (MDS), myeloproliferative neoplasm (MPN), or multiple myeloma (MM).

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