Methods for treating acute leukemia

Abstract

Provided herein are methods of treating acute leukemia, e.g., acute myeloid leukemia (AML), in an individual, e.g., an individual having a mutation in the nucleophosmin 1 (NPM1) gene or a rearrangement in the lysine [K]-methyltransferase 2A (KMT2A) gene, the methods comprising administering the menin inhibitor diftomenib to the individual at a dose of 600 milligrams daily.

Description

[Technical Field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims the benefit of U.S. Provisional Application No. 63 / 382,084, filed November 2, 2022, U.S. Provisional Application No. 63 / 386,649, filed December 8, 2022, U.S. Provisional Application No. 63 / 497,125, filed April 19, 2023, and U.S. Provisional Application No. 63 / 504,995, filed May 30, 2023, each of which is incorporated by reference herein in its entirety. [Background technology]

[0002] Acute leukemia is a group of blood cancers characterized by a rapid increase in the number of immature blood cells, including acute myeloid leukemia (AML) and acute lymphocytic leukemia (ALL). AML is a diverse group of highly lethal blood cancers characterized by the proliferation of myeloid precursors (myeloid blasts or progranulocytes) that fail to differentiate normally. AML develops as a result of a series of genetic alterations in hematopoietic progenitor cells. These alterations alter normal hematopoietic growth and differentiation, leading to the abnormal accumulation of large numbers of these immature myeloid blasts in the bone marrow and peripheral blood, which eventually prevents the production of normal blood cells. As with other malignancies, genetic alterations in AML include mutations in oncogenes and loss of tumor suppressor genes. However, in contrast to most solid tumors, many hematologic malignancies are associated with a single, characteristic cytogenetic abnormality. This clinically heterogeneous disease is characterized by numerous chromosomal abnormalities and gene mutations, which lead to marked differences in response and survival after chemotherapy and significant challenges to successful and sustained AML treatment (Kumar, CC, Genes Cancer 2011, 2(2), 95-107.). Similarly, ALL involves genetic alterations that result in the generation of leukemic lymphoblasts in the bone marrow, which affect the production of new red blood cells, white blood cells, and platelets. ALL is the most common type of leukemia in young children and the most common cause of cancer death in children. While most cases of ALL occur in children, 80% of ALL deaths occur in adults. Summary of the Invention

[0003]

[0003] Described herein are methods for treating acute leukemia, such as acute myeloid leukemia (AML) or acute lymphocytic leukemia (ALL), in an individual, or for inhibiting proliferation and / or inducing apoptosis in leukemia cells in an individual, wherein the acute leukemia or leukemia cells contain a nucleophosmin 1 (NPM1) mutation, a lysine methyltransferase 2a (KMT2A) rearrangement, a SET domain-containing 2 (SETD2) mutation, or a runt-related transcription factor 1 (RUNX1) mutation, the method comprising administering 600 milligrams of diftomenib or an isotope thereof, or a pharmaceutically acceptable salt of such a compound or isotope thereof, or a solvate of any of the foregoing (collectively, "diftomenib or a pharmaceutically acceptable form thereof"), or a pharmaceutical composition comprising diftomenib or a pharmaceutically acceptable form thereof, to the individual daily. In some embodiments, the acute leukemia is menin-dependent. In some embodiments, the acute leukemia or leukemia cells contain an NPM1 mutation, optionally in combination with a FLT3 mutation (e.g., an FLT3-internal tandem duplication (ITD) mutation or a FLT3 mutation in the tyrosine kinase domain (FLT3-TKD mutation)) or an IDH mutation (e.g., an IDH1 mutation or an IDH2 mutation), or a combination thereof. As described herein, diftomenib administered at 600 milligrams daily exhibits significant clinical efficacy in acute leukemia, as demonstrated by clinical response rates, combined with a manageable safety and tolerability profile, particularly when administered to individuals with specific genetic abnormalities, including NPM1 mutations or KMT2A rearrangements.Also described herein are methods of treating acute leukemia, e.g., AML or ALL, in an individual, or methods of inhibiting proliferation and / or inducing apoptosis in leukemic cells in an individual, wherein the acute leukemia is menin-dependent leukemia, and optionally the leukemia comprises an NPM1 mutation, a KMT2A rearrangement, a KMT2A partial tandem duplication (KMT2A-PTD), a SETD2 mutation, a RUNX1 mutation, a FLT3 mutation (e.g., a FLT3-ITD mutation or a FLT3-TKD mutation), an IDH mutation (e.g., an IDH1 mutation or an IDH2 mutation), a TERT mutation, or a BRAF mutation, or any combination thereof, the method comprising administering 600 mg of diftomenib or a pharmaceutically acceptable form thereof, or a pharmaceutical composition comprising diftomenib or a pharmaceutically acceptable form thereof, to the individual daily. In some embodiments, the acute leukemia comprises an NPM1 mutation, optionally in combination with a FLT3 mutation (e.g., a FLT3-ITD mutation or a FLT3-TKD mutation) or an IDH mutation (e.g., an IDH1 mutation or an IDH2 mutation), or a combination thereof.

[0004] Provided herein is a method for treating extramedullary leukemia in an individual with acute leukemia, the method comprising daily administering to the individual a therapeutically effective amount of a menin inhibitor, particularly 600 milligrams of diftomenib or a pharmaceutically acceptable form thereof, wherein optionally the acute leukemia comprises an NPM1 mutation, a KMT2A rearrangement, a SETD2 mutation, or a RUNX1 mutation. The extramedullary leukemia is characterized by the presence of leukemia cell aggregates outside the bone marrow cavity, optionally in the form of a solid tumor of myeloblasts. In some embodiments, the extramedullary leukemia is menin-dependent. Provided and described herein are methods of treating extramedullary leukemia in an individual with acute leukemia, wherein the acute leukemia is menin-dependent leukemia, and optionally the leukemia comprises an NPM1 mutation, a KMT2A rearrangement, a KMT2A-PTD mutation, a SETD2 mutation, a RUNX1 mutation, a FLT3 mutation (e.g., a FLT3-ITD mutation or a FLT3-TKD mutation), an IDH mutation (e.g., an IDH1 mutation or an IDH2 mutation), a TERT mutation, or a BRAF mutation, or any combination thereof, the method comprising administering 600 milligrams of diftomenib or a pharmaceutically acceptable form thereof, or a pharmaceutical composition comprising diftomenib or a pharmaceutically acceptable form thereof, to the individual daily. In some embodiments, the extramedullary leukemia comprises an NPM1 mutation, optionally in combination with a FLT3 mutation (e.g., a FLT3-ITD mutation or a FLT3-TKD mutation) or an IDH mutation (e.g., an IDH1 mutation or an IDH2 mutation), or a combination thereof. In some embodiments, the acute leukemia is AML or ALL.

[0005] A method for increasing the level of myeloid blasts in the blood of an individual with AML, comprising daily administering a therapeutically effective amount of a menin inhibitor, particularly 600 mg of diftomenib or a pharmaceutically acceptable form thereof, to the individual, wherein the AML optionally comprises NPM1 mutation, KMT2A rearrangement, SETD2 mutation, or RUNX1 mutation.Diftomenib has been observed to cause a significant and stable increase in the level of blasts in the blood, a situation that usually indicates the progression of leukemia disease.However, with diftomenib, it has been found that the release of blasts from the bone marrow to the intramedullary space correlates with the sensitivity of leukemia to diftomenib therapy, and in such cases, this effect is not an indicator of disease progression.In some embodiments, AML is menin-dependent. In some embodiments, the AML comprises an NPM1 mutation, a KMT2A rearrangement, a KMT2A-PTD mutation, a SETD2 mutation, a RUNX1 mutation, an FLT3 mutation (e.g., an FLT3-ITD mutation or an FLT3-TKD mutation), an IDH mutation (e.g., an IDH1 mutation or an IDH2 mutation), a TERT mutation, or a BRAF mutation, or any combination thereof. In some aspects, the AML comprises an NPM1 mutation, optionally in combination with an FLT3 mutation (e.g., an FLT3-internal tandem duplication (ITD) mutation or an FLT3 mutation in the tyrosine kinase domain (FLT3-TKD mutation)) or an IDH mutation (e.g., an IDH1 mutation or an IDH2 mutation), or a combination thereof. The increase in the level of myeloid blasts may be transient, followed by a decrease in myeloid blasts in the blood.

[0006] Also provided herein is a method for identifying acute leukemia in an individual as being susceptible to administration of a menin inhibitor, particularly diftomenib, said method comprising: administering to the individual daily an effective amount of a menin inhibitor, particularly 600 mg of diftomenib or a pharmaceutically acceptable form thereof; receiving an identification of the level of myeloid blasts in a first blood sample taken from the individual at a first time point, either before the start of administration or during administration; receiving an identification of the level of myeloid blasts in a second blood sample taken from the individual at a second time point after the first time point and during administration; determining that the acute leukemia is susceptible to the administration if the level of myeloid blasts at the second time point is greater than the level at the first time point; Optionally, the acute leukemia is menin-dependent; alternatively, the acute leukemia comprises an NPM1 mutation, a KMT2A rearrangement, a KMT2A-PTD mutation, a SETD2 mutation, a RUNX1 mutation, a FLT3 mutation (e.g., a FLT3-ITD mutation or a FLT3-TKD mutation), an IDH mutation (e.g., an IDH1 mutation or an IDH2 mutation), a TERT mutation, or a BRAF mutation, or any combination thereof; alternatively, the acute leukemia comprises an NPM1 mutation, a KMT2A rearrangement, a SETD2 mutation, or a RUNX1 mutation; preferably, the acute leukemia comprises an NPM1 mutation or a KMT2A rearrangement. In some embodiments, the acute leukemia comprises an NPM1 mutation, optionally in combination with a FLT3 mutation (e.g., a FLT3-ITD mutation or a FLT3-TKD mutation) or an IDH mutation (e.g., an IDH1 mutation or an IDH2 mutation), or a combination thereof.

[0007] Also provided herein is a method of treating a differentiation disorder in an individual diagnosed with, or reducing the risk of developing a severe differentiation disorder in an individual with, acute leukemia, wherein optionally, the acute leukemia is menin-dependent; or the acute leukemia comprises an NPM1 mutation, a KMT2A rearrangement, a KMT2A-PTD mutation, a SETD2 mutation, a RUNX1 mutation, an FLT3 mutation (e.g., an FLT3-ITD mutation or an FLT3-TKD mutation), an IDH mutation (e.g., an IDH1 mutation or an IDH2 mutation), a TERT mutation, or a BRAF mutation, or any combination thereof; or the acute leukemia comprises an NPM1 mutation, a KMT2A rearrangement, a SETD2 mutation, or a RUNX1 mutation; preferably, the acute leukemia comprises an NPM1 mutation or a KMT2A rearrangement; and (a) administering to an individual a therapeutically effective amount of a menin inhibitor, particularly 600 mg of diftomenib or a pharmaceutically acceptable form thereof, daily; (b) administering IV hydration to the individual; and optionally, (c) administering to the individual an effective amount of a xanthine oxidase inhibitor, optionally wherein the xanthine oxidase inhibitor is allopurinol; and optionally administering the allopurinol at a dose of about 200-400 mg / m 2 and administering daily at a dose of 1 to 3 divided doses up to a maximum of about 800 mg / day. In some embodiments, the acute leukemia comprises an NPM1 mutation, optionally in combination with a FLT3 mutation (e.g., a FLT3-ITD mutation or a FLT3-TKD mutation) or an IDH mutation (e.g., an IDH1 mutation or an IDH2 mutation), or a combination thereof.

[0008] Also provided herein is a method of treating a differentiation disorder in an individual with acute leukemia, or a method of reducing the risk of developing a severe differentiation disorder in an individual with acute leukemia, optionally wherein the acute leukemia is menin-dependent; or wherein the acute leukemia comprises an NPM1 mutation, a KMT2A rearrangement, a KMT2A-PTD mutation, a SETD2 mutation, a RUNX1 mutation, an FLT3 mutation (e.g., an FLT3-ITD mutation or an FLT3-TKD mutation), an IDH mutation (e.g., an IDH1 mutation or an IDH2 mutation), a TERT mutation, or a BRAF mutation, or any combination thereof; or wherein the acute leukemia comprises an NPM1 mutation, a KMT2A rearrangement, a SETD2 mutation, or a RUNX1 mutation, preferably an NPM1 mutation or a KMT2A rearrangement; (a) administering to an individual daily an effective amount of a menin inhibitor, particularly 600 mg of diftomenib or a pharmaceutically acceptable form thereof; (b) administering IV hydration to the individual; (c) administering a therapeutically effective amount of rasburicase, optionally at a dose of about 0.2 mg / kg, optionally as an intravenous infusion over about 30 minutes, for up to about 5 days. In some embodiments, the acute leukemia comprises an NPM1 mutation, optionally in combination with a FLT3 mutation (e.g., a FLT3-ITD mutation or a FLT3-TKD mutation) or an IDH mutation (e.g., an IDH1 mutation or an IDH2 mutation), or a combination thereof.

[0009] Also provided herein is a method of treating a differentiation disorder or reducing the risk of developing a severe differentiation disorder in an individual with acute leukemia, optionally wherein the acute leukemia is menin-dependent; or wherein the acute leukemia comprises an NPM1 mutation, a KMT2A rearrangement, a KMT2A-PTD mutation, a SETD2 mutation, a RUNX1 mutation, an FLT3 mutation (e.g., an FLT3-ITD mutation or an FLT3-TKD mutation), an IDH mutation (e.g., an IDH1 mutation or an IDH2 mutation), a TERT mutation, or a BRAF mutation, or any combination thereof; or wherein the acute leukemia comprises an NPM1 mutation, a KMT2A rearrangement, a SETD2 mutation, or a RUNX1 mutation, preferably an NPM1 mutation or a KMT2A rearrangement; and (a) administering to the individual a prophylactic and / or effective amount of a corticosteroid, optionally the corticosteroid being prednisone (or an equivalent dose of an alternative corticosteroid) at a dose of about 0.5 mg / kg; (b) administering a therapeutically effective amount of a menin inhibitor to the individual. In some embodiments, the acute leukemia comprises an NPM1 mutation, optionally in combination with a FLT3 mutation (e.g., a FLT3-ITD mutation or a FLT3-TKD mutation) or an IDH mutation (e.g., an IDH1 mutation or an IDH2 mutation), or a combination thereof.

[0010] Also provided herein is a method of reducing transfusion dependency in an individual with acute leukemia, optionally wherein the acute leukemia is menin-dependent; or wherein the acute leukemia comprises an NPM1 mutation, a KMT2A rearrangement, a KMT2A-PTD mutation, a SETD2 mutation, a RUNX1 mutation, a FLT3 mutation (e.g., a FLT3-ITD mutation or a FLT3-TKD mutation), an IDH mutation (e.g., an IDH1 mutation or an IDH2 mutation), a TERT mutation, or a BRAF mutation, or any combination thereof; or wherein the acute leukemia comprises an NPM1 mutation, a KMT2A rearrangement, a SETD2 mutation, or a RUNX1 mutation, preferably an NPM1 mutation or a KMT2A rearrangement, the method comprising administering 600 milligrams of diftomenib or a pharmaceutically acceptable form thereof, or a pharmaceutical composition comprising diftomenib or a pharmaceutically acceptable form thereof, to the individual daily.

[0011] Also provided herein are pharmaceutical compositions comprising an optimal biological dose, a recommended Phase 2 dose, a safe and effective dose, or a submaximal tolerated dose of diftomenib or a pharmaceutically acceptable form thereof. In some embodiments, the optimal biological dose, the recommended Phase 2 dose, a safe and effective dose, or a submaximal tolerated dose is 600 mg. In some embodiments, diftomenib or a pharmaceutically acceptable form thereof has breakthrough therapy designation.

[0012] Incorporation by Reference All publications, patents, and patent applications mentioned in this specification are herein incorporated by reference to the same extent as if each individual publication, patent, or patent application was specifically and individually indicated to be incorporated by reference. [Brief explanation of the drawings]

[0013] [Figure 1] Trough concentrations of diftomenib in plasma (ng / mL) or bone marrow, heart, or spleen tissue (ng / g) after daily dosing. [Figure 2]Mean change from baseline in peripheral blasts (orange) and white blood cells (blue) at (a) cycle 1, day 8, (b) cycle 1, day 15, and (c) cycle 2, day 1. [Figure 3] Exposure levels determined by AUC0-24, Ctrough, and Cmax at steady state after daily diftomenib doses of 50, 200, 400, 600, and 800 mg. A: AUC0-24 at steady state. B: Ctrough levels at steady state. C: Cmax levels at steady state. [Figure 4A] MEIS1 expression by gene subtype as a function of dose. Figure 4A: Individual subject data. [Figure 4B] MEIS1 expression by gene subtype as a function of dose. Figure 4B: Mean expression levels. DETAILED DESCRIPTION OF THE INVENTION

[0014] Epigenetic modifications in the lysine methyltransferase 2A (KMT2A) gene, resulting in KMT2A fusions with over 60 partner genes, play a causative role in the onset, development, and progression of some acute leukemias (Borkin et al., Cancer Cell 2015, 27(4), 589-602). HOXA9 and MEIS1 are key oncogenes overexpressed in KMT2A-rearranged leukemias (Thiel et al., Bioessays 2012, 34(9), 771-780). HOXA9 and MEIS1 transcription factors drive AML by upregulating stem cell programs and blocking myeloid differentiation. KMT2A (MLL) rearrangements alter the normal histone methyltransferase function of KMT2A (MLL), deregulating these HOX genes, resulting in persistently high HOX levels and a blockade of hematopoietic (myeloid) differentiation, ultimately leading to acute leukemia (Kuehn et al., Cancer Discov. 2016, 6(10), 1166-1181; Klossowski et al., J. Clin. Invest. 2020, 130(2), 981-997; Issa et al., Blood Cancer J. 2021, 11(9), 162; Chan et al., Front. Cell Dev. Biol. 2019, 7, 81). Translocations (rearrangements) in the KMT2A gene (KMT2A-r) occur in 5-10% of AML patients, and the 5-year survival rate for KMT2A-r patients is less than 20% (Issa, 2021).

[0015] Nucleoplasmin 1 (NPPM1) encodes a protein involved in cellular protein transport to the nucleolus. NPM1 further depends on the interaction between menin and wild-type KMT2A (MLL) to drive leukemogenic gene expression (Kuehn, 2016). The NPM1 gene is upregulated, mutated, and undergoes chromosomal translocation in many tumor types.

[0016] In AML, a key common factor in the regulation of these leukemogenic genes by KMT2A(MLL) is the interaction between the N-terminal portion of KMT2A(MLL) and menin, which is essential for the gene activation effects of both wild-type KMT2A(MLL) and KMT2A(MLL) fusion proteins to the HOXA9 and MEIS1 promoter regions. Menin is a highly specific and direct binding partner of KMT2A(MLL) and KMT2A(MLL) fusion proteins required for the regulation of their target genes (Yokoyama et al., Cell 2005, 123(2), 207-218). Many studies have demonstrated the important role of menin as an oncogenic cofactor in leukemic transformation mediated by the KMT2A(MLL) fusion protein, and in recent years, menin has been shown to further drive overexpression of HOXA9 and MEIS1 in a subset of normal karyotype AML associated with NPM1 mutations (Yokoyama 2005; Caslini et al., Cancer Res. 2007, 67(15), 7275-7283; Yokoyama et al., Cancer Cell 2008, 14(1), 36-46; Kuehn, 2016). NPM1 mutations occur in 25-30% of AML patients with or without other mutations, and the 5-year survival rate for AML patients with NPM1 mutations (NPM1-m) is approximately 50% (Angenendt et al., J. Clin. Oncol. 2019, 37(29), 2632-2642; Thiede et al., Blood 2006, 107(10), 4011-4020).

[0017] Additional genetic alterations that negatively impact disease progression and prognosis include alterations to another histone methyltransferase gene called SET domain-containing 2 (SETD2), which contains a truncating mutation seen in 1-2% of AML cases, and to the gene encoding runt-related transcription factor 1, also known as acute myeloid leukemia 1 protein (RUNX1), a transcription factor that regulates the differentiation of hematopoietic stem cells into mature blood cells. Chromosomal translocations involving RUNX1 are associated with several types of leukemia, including AML.

[0018] The standard treatment for AML and ALL is intensive chemotherapy (a combination of anthracyclines, e.g., daunorubicin or idarubicin, and cytarabine in a "7+3" regimen), which has been used for over 40 years. However, intensive chemotherapy has various difficult side effects, is not suitable for patients with poor health (e.g., elderly patients), and results with standard chemotherapy remain unsatisfactory. In addition, even when patients respond, more than half of adult patients and approximately 80% of elderly patients experience primary refractory disease, relapse, or treatment-related death. Although improvements in supportive care have improved overall survival (OS), up to 30–40% of patients have refractory disease (i.e., fail to achieve a morphological complete response (CR) after one to two cycles of induction therapy), and these patients have a median survival of less than one year (Horibata et al., Proc. Natl. Acad. Sci. 2019, 116(21), 10494–10503). Patients can be further treated with stem cell transplantation.

[0019] AML comprises approximately 30% of all adult leukemia cases and 80% of all acute leukemia cases in adults. The outcomes for patients with AML are often considered severe or poor, further characterized by high mortality rates. Notably, more than 50% of patients who achieve a complete response to induction therapy relapse within 1–3 years, whereas the presence of NPM1-m correlates with a risk of refractory disease to standard intensive induction therapy, with KMT2A-r representing an aggressive and poor-prognostic group of hematologic cancers (Horibata, 2019; Wang et al., Blood 2020, 136(Suppl. 1), 7.). Novel therapeutic treatments for NPM1-m and KMT2A-r AML remain needed.

[0020] Diftomenib (KO-539) is a potent and selective inhibitor of the menin-KMT2A (MLL) complex with downstream effects on HOXA9 / MEIS1 expression (Burrows et al., Proceedings of the AACR EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol. Cancer Ther. 2018;17(1 Suppl): Abstract nr LB-A27.). KOMET-001 (NCT04067336) is a phase 1 / 2 open-label trial evaluating diftomenib in adult patients with relapsed and / or refractory AML, including select NPM1 mutations or KMT2A rearrangements, both of which represent subtypes of this particularly aggressive form of hematologic cancer for which there is a high unmet need, especially once the disease has relapsed or become refractory to available therapies (see https: / / kuraoncology.com / clinical-trials / clinical-trials-komet-001 / , accessible October 2023). The study will investigate the safety and tolerability, pharmacokinetics, and antitumor activity of diftomenib in these AML populations. Additional study endpoints included determining the optimal biologic dose or recommended phase 2 dose.

[0021] Provided herein is a method for treating acute leukemia such as AML or ALL, particularly AML, and in some embodiments, the method comprises administering diftomenib to an individual at a daily dose of 600 mg.The method provided herein generally encompasses the finding that individuals with acute leukemia, for example, AML or ALL, particularly AML, demonstrate increased response rate and / or improved safety profile when treated with diftomenib, and the acute leukemia is menin-dependent, or the acute leukemia comprises a mutation in the NPM1 gene, a rearrangement in the KMT2A gene, a mutation in the SETD2 gene, or a mutation in the RUNX1 gene, or comprises an NPM1 mutation, a KMT2A rearrangement, a KMT2A-PTD mutation, a SETD2 mutation, a RUNX1 mutation, an FLT3 mutation (for example, an FLT3-ITD mutation or an FLT3-TKD mutation), an IDH mutation (for example, an IDH1 mutation or an IDH2 mutation), a TERT mutation, or a BRAF mutation, or any combination thereof, and particularly comprises a mutation in the NPMI gene. For example, treatment of NPM1-m AML comprising administering 600 mg of the menin inhibitor diftomenib was more effective and resulted in a more favorable safety profile than treatment of non-NPM1-m AML at the same dose. Thus, in some embodiments, provided herein is a method for treating AML in an individual, wherein the AML comprises an NPM1 mutation, the method comprising administering 600 mg of diftomenib, or a pharmaceutically acceptable form thereof, to the individual. In some aspects, the acute leukemia comprises an NPM1 mutation, optionally in combination with an FLT3 mutation (e.g., an FLT3-ITD mutation or an FLT3-TKD mutation) or an IDH mutation (e.g., an IDH1 mutation or an IDH2 mutation), or a combination thereof.

[0022] Furthermore, in some embodiments, the use of diftomenib or a pharmaceutically acceptable form thereof in a method for treating acute leukemia (e.g., AML or ALL, particularly AML) in an individual is provided, wherein the acute leukemia is menin-dependent, or the acute leukemia comprises an NPM1 mutation, a rearrangement in the KMT2A gene, a mutation in the SETD2 gene, or a mutation in the RUNX1 gene, or the acute leukemia comprises an NPM1 mutation, a KMT2A rearrangement, a KMT2A-PTD mutation, a SETD2 mutation, a RUNX1 mutation, an FLT3 mutation (e.g., an FLT3-ITD mutation or an FLT3-TKD mutation), an IDH mutation (e.g., an IDH1 mutation or an IDH2 mutation), a TERT mutation, or a BRAF mutation, or any combination thereof, particularly a mutation in the NPM1 gene. Similarly, the use of diftomenib or a pharmaceutically acceptable form thereof in a method for treating AML with an NPM1 mutation is also provided. Similarly, in some embodiments, diftomenib or a pharmaceutically acceptable form thereof is provided for use in treating acute leukemia (e.g., AML or ALL, particularly AML) in an individual, wherein the AML comprises an NPM1 mutation, a rearrangement in the KMT2A gene, a mutation in the SETD2 gene, or a mutation in the RUNX1 gene, particularly a mutation in the NPM1 gene. In some aspects, the acute leukemia comprises an NPM1 mutation, optionally in combination with an FLT3 mutation (e.g., an FLT3-ITD mutation or an FLT3-TKD mutation) or an IDH mutation (e.g., an IDH1 mutation or an IDH2 mutation), or a combination thereof.

[0023] Diftomenib and pharmaceutically acceptable forms Diftomenib is a potent inhibitor of menin-MLL (KMT2A) interaction (IC) in clinical development for the treatment of acute leukemias, including NPM1-mutant (NPM1-m) and KMT2A-rearranged (KMT2A-r) AML, as well as some other genetically defined acute leukemias with high unmet need. 50 22 nM) and is a selective inhibitor.

[0024] Diftomenib has the following structure:

[0025] [ka] is a compound having the formula Alternatively, it is named (S)-4-methyl-5-((4-((2-(methylamino)-6-(2,2,2-trifluoroethyl)thieno[2,3-d]pyrimidin-4-yl)amino)piperidin-1-yl)methyl)-1-(2-(4-(methylsulfonyl)piperazin-1-yl)propyl)-1H-indole-2-carbonitrile. In some embodiments, the methods described herein employ a pharmaceutically acceptable form of diftomenib. In some embodiments, the methods described herein employ diftomenib or a pharmaceutically acceptable salt thereof. In some embodiments, the methods described herein employ diftomenib or a solvate thereof. In certain embodiments, diftomenib comprises the free base form or a solvate thereof. In some embodiments, stereoisomers and / or metabolites of diftomenib are also included.

[0026] In some embodiments, the menin inhibitor described herein is diftomenib or a pharmaceutically acceptable form thereof.

[0027] Diftomenib Dosage and Administration Regimen In some embodiments, the methods provided herein comprise administering to an individual an effective amount, for example, 600 mg, of diftomenib or a pharmaceutically acceptable form thereof daily. In some embodiments, the methods comprise administering to an individual diftomenib or a pharmaceutically acceptable form thereof at an optimal biological dose, a recommended Phase 2 dose, a safe and effective dose, or a dose less than the maximum tolerated dose. In some embodiments, the optimal biological dose is 600 mg. In some embodiments, the recommended Phase 2 dose (RP2D) is 600 mg. In some embodiments, the safe and effective dose is 600 mg. In some embodiments, the dose less than the maximum tolerated dose is 600 mg. In some embodiments, a dose of 600 mg is a safe and effective amount. In some embodiments, diftomenib or a pharmaceutically acceptable form thereof has breakthrough therapy designation.

[0028] In some embodiments, administering diftomenib or a pharmaceutically acceptable form thereof comprises administering to the individual for at least 3 days, or at least 5 days, or at least 7 days, or at least 10 days, or at least 14 days, or at least 21 days, or at least 28 days, or over a cycle comprising at least 28 days, or over a cycle comprising 28 days.

[0029] In certain embodiments, diftomenib, or a pharmaceutically acceptable form thereof, is administered to an individual daily for N cycles, including at least 28 days, or for a cycle including 28 days, where N is at least 1. In certain embodiments, N is at least 2. In certain embodiments, N is at least 3. In certain embodiments, N is at least 4. In certain embodiments, N is 2. In certain embodiments, N is 3. In certain embodiments, N is 4. In certain embodiments, N is 5. In certain embodiments, N is 6. In certain embodiments, N is 7. In certain embodiments, the cycles are continuous (i.e., 0 days between cycles).

[0030] In certain embodiments, diftomenib or a pharmaceutically acceptable form thereof is administered orally.

[0031] In some embodiments, daily administration is administration once or twice daily. In some embodiments, daily administration is once daily.

[0032] The dose of diftomenib presented herein refers to the amount of free base (when using the free form) or the amount of free base equivalent (when using salts and / or solvates).Thus, for example, when using salt forms, the total mass of a daily dose will be more than 600 mg, but the total mass will be selected to provide 600 mg of diftomenib free base equivalent.

[0033] In some embodiments, diftomenib or a pharmaceutically acceptable form thereof is administered in combination with a P-gp inhibitor or a breast cancer resistance protein (BCRP) inhibitor.

[0034] Pharmaceutical Composition Also provided herein are pharmaceutical compositions comprising an optimal biological dose, a recommended Phase 2 dose, a safe and effective dose, or a submaximal tolerated dose of diftomenib or a pharmaceutically acceptable form thereof. In some embodiments, the optimal biological dose, the recommended Phase 2 dose, the safe and effective dose, or the submaximal tolerated dose is 600 mg. In some embodiments, diftomenib or a pharmaceutically acceptable form thereof has breakthrough therapy designation. In some embodiments, the pharmaceutical composition comprises one or more dosage forms, such as one or more oral dosage forms, optionally each containing 50-600 mg of diftomenib, or containing 50 mg, 100 mg, 200 mg, 300 mg, 400 mg, 500 mg, or 600 mg of diftomenib. In some embodiments, the total amount of diftomenib in the one or more oral dosage forms is 600 mg. In some embodiments, each oral dosage form contains 100 mg, 200 mg, or 300 mg of diftomenib. In some embodiments, each oral dosage form contains 200 mg or 300 mg of diftomenib. In some embodiments, the diftomenib administered in the methods provided herein is administered using such a pharmaceutical composition or oral dosage form.

[0035] AML and ALL In some embodiments, the methods provided herein are directed to the treatment of acute myeloid leukemia (AML). AML is a bone marrow disease and hematopoietic stem cell disorder characterized by genetic alterations in blood cell precursors that lead to the overproduction of tumor clonal myeloid stem cells. While extramedullary manifestations (e.g., myeloid sarcoma, leukemia cutis) can occur, the underlying disease generally results from abnormalities in blood cell production. In certain embodiments, acute myeloid leukemia (AML) includes relapsed AML, refractory AML, or both relapsed and refractory AML. In certain embodiments, the AML is refractory AML. In certain embodiments, the refractory disease is refractory to intensive chemotherapy (first-line standard of care) including cytarabine and an anthracycline (idarubicin or daunorubicin), typically as a 7+3 combination of continuous infusion of cytarabine and intermittent administration of the anthracycline over 7 days and 3 days, respectively. In certain embodiments, the first-line standard of care for newly diagnosed AML in ineligible patients or as part of a 7+3 regimen is venetoclax plus azacitadine (ven / aza). In some embodiments, the AML is resistant to ven or ven / aza. In some embodiments, the AML has progressed during or after treatment with ven or ven / aza. In some embodiments, the AML is relapsed AML. In some embodiments, the AML is both refractory and relapsed AML. In some embodiments, the AML is acute promyelocytic leukemia, acute myeloblastic leukemia, or acute megakaryoblastic leukemia.

[0036] In some embodiments, the methods provided herein are directed to the treatment of acute lymphocytic leukemia (ALL). ALL is a type of cancer of the blood and bone marrow involving uncontrolled proliferation of abnormal immature lymphocytes, resulting in bone marrow replacement and blood infiltration. In certain embodiments, ALL includes relapsed ALL, refractory ALL, or both relapsed and refractory ALL. In certain embodiments, ALL is refractory ALL. In some embodiments, ALL is relapsed ALL. In some embodiments, ALL is both refractory and relapsed ALL. In some embodiments, ALL is precursor B-cell acute lymphoblastic leukemia, precursor T-cell acute lymphoblastic leukemia, Burkitt's leukemia, or acute mixed lineage leukemia.

[0037] Genetic alterations in acute leukemia In some embodiments, the acute leukemia or leukemia cells are characterized by a genetic alteration (e.g., one or more) that can be selected from an NPM1 mutation, a KMT2A rearrangement, a SETD2 mutation, and a RUNX1 mutation. In some embodiments, the acute leukemia or leukemia cells are menin dependent. In some embodiments, the acute leukemia or leukemia cells comprise an NPM1 mutation. In some embodiments, the acute leukemia or leukemia cells comprise a KMT2A rearrangement. In some embodiments, the acute leukemia or leukemia cells comprise a KMT2A-PTD. In some embodiments, the acute leukemia or leukemia cells comprise a SETD2 mutation. In some embodiments, the acute leukemia or leukemia cells comprise a RUNX1 mutation. In some embodiments, the acute leukemia or leukemia cells comprise a SETD2 mutation and a RUNX1 mutation. In some embodiments, the acute leukemia or leukemia cells comprise one or more mutations selected from FLT3, FLT3-ITD, FLT3-TKD, IDH, IDH1, IDH2, TERT, and BRAF. In some embodiments, the acute leukemia or leukemia cells comprise an NPM1 mutation, a KMT2A rearrangement, a KMT2A-PTD mutation, a SETD2 mutation, a RUNX1 mutation, a FLT3-ITD mutation, a FLT3-TKD mutation, an IDH mutation (e.g., an IDH1 mutation or an IDH2 mutation), a TERT mutation, or a BRAF mutation, or any combination thereof. In some embodiments, the acute leukemia or leukemia cells comprise an NPM1 mutation, and optionally one or more mutations selected from FLT3 (such as FLT3-ITD or FLT3-TKD), an IDH (such as an IDH1 mutation or an IDH2 mutation), TERT, or BRAF. In some embodiments, the acute leukemia comprises an NPM1 mutation, optionally in combination with a FLT3 mutation (e.g., a FLT3-internal tandem duplication (ITD) mutation or a FLT3 mutation in the tyrosine kinase domain (FLT3-TKD mutation)) or an IDH mutation (e.g., an IDH1 mutation or an IDH2 mutation), or a combination thereof.

[0038] In certain embodiments, the mutant NPM1 gene contains one or more mutations relative to the wild-type NPM1 gene sequence. NPM1 generally refers to and encompasses the gene encoding the NPM1 protein (see, e.g., UniProt ID P06748). In certain instances, the NPM1 gene encompasses NCBI gene ID 4869 and / or NCBI reference sequence: NG_016018.1 (5001..28181). NPM1 mutations are found in approximately 25-30% of AML cases and, in certain instances, are generally characterized by the presence of a primary four-base pair insertion that creates a new N-terminal nuclear export signal, resulting in abnormal cytoplasmic accumulation of the mutant NPM1c protein. NPM1 mutations include type A, type B, and type D mutations, each characterized by a four-nucleotide insertion in exon 12 that results in cytoplasmic localization of NPM1 (NPM1-c). In certain instances, NPM1-c, which normally promotes myeloid lineage differentiation, is re-transported into the nucleus by XPO1 and binds to transcription factors that can directly affect gene expression, resulting in mislocalization of the transcription factors.

[0039] In certain embodiments, the NPM1 mutation comprises a type A, type B, type C, or type D mutation. In certain embodiments, the NPM1 mutation comprises a type A mutation. In certain embodiments, the NPM1 mutation comprises a type B mutation. In certain embodiments, the NPM1 mutation comprises a type C mutation. In certain embodiments, the NPM1 mutation comprises a type D mutation. In certain embodiments, the NPM1 mutation results in cytoplasmic localization of NPM1. In certain embodiments, the NPM1 mutation comprises an insertion (e.g., a 4-nucleotide insertion) in exon 12 of the NPM1 gene.

[0040] Multiple translocations involving the KMT2A gene have been reported in both AML and ALL. The KMT2A-MLLT3 fusion, caused by t(9;11)(p21.3;q23.3), is the most common KMT2A rearrangement in adults with AML, although over 80 different fusion partners have been described. Translocations such as t(9;11)(p22;q23)(MLLT3;KMT2A) or t(10;11)(p12;q23)(AF10;KMT2A) have been reported in 4% of adult myeloid leukemias. Similarly, multiple SETD2 and RUNX1 mutations have been reported to be involved in acute leukemia.

[0041] In some embodiments, the acute leukemia or leukemia cells comprise more than one mutation or rearrangement. In some embodiments, the acute leukemia or leukemia cells comprise (a) an NPM1 mutation or a KMT2A rearrangement, and (b) at least one mutation selected from FLT3 (such as FLT3-ITD or FLT3-TKD), IDH (such as IDH1 or IDH2), TERT, and BRAF. In some embodiments, the acute leukemia or leukemia cells comprise (a) an NPM1 mutation, and (b) at least one mutation selected from FLT3 mutation (such as FLT3-ITD or FLT3-TKD) or IDH (such as IDH1 or IDH2).

[0042] In certain instances, characterization of gene mutations or rearrangements can be achieved by collecting bone marrow (BM aspirates), whole blood samples, and / or tumor samples followed by known assays for nucleic acid analysis. In certain embodiments, mutations or rearrangements are detected by sequencing (e.g., genomic sequencing), such as MyAML®, a CLIA-certified, next-generation sequencing assay for mutations in 194 genes associated with AML. In certain embodiments, specific mutations or rearrangements are detected by molecular testing, such as polymerase chain reaction (e.g., followed by fragment analysis and / or capillary gel electrophoresis). In certain embodiments, mutations or rearrangements are detected by RT-PCR or quantitative PCR. In some embodiments, the methods provided herein include detecting mutations or rearrangements or receiving identification of mutations or rearrangements before administering diftomenib, particularly NPM1 mutations, and optionally, NPM1 mutations are detected by next-generation sequencing or PCR assays.

[0043] Effectiveness As described in Example 3, efficacy outcomes of patients treated with diftomenib were assessed according to the rates of CR, CR / CRh, and CRc, as well as by ORR. In some embodiments, the CR or CR / CRh rate is at least about 18%, or at least about 19%, or at least about 20%, or at least about 21%, or at least about 22%, or at least about 23%, or at least about 24%, or at least about 25%, or at least about 26%, or at least about 27%, or at least about 28%, or at least about 29%, or at least about 30%, or at least about 31%, or at least about 32%, or at least about 33%, or at least about 34%, or at least about 35%, or is about 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, or 35%, or is about 35%. In some embodiments, the CR rate is at least about 35%, or is about 35%. In some embodiments, the CRc rate is at least about 20%, or at least about 21%, or at least about 22%, or at least about 23%, or at least about 24%, or at least about 25%, or at least about 26%, or at least about 27%, or at least about 28%, or at least about 29%, or at least about 30%, or at least about 31%, or at least about 32%, or at least about 33%, or at least about 34%, or at least about 35%, or at least about 36%, or at least about 37%, or at least about 38%, or at least about 39%, or at least about 40%, or is about 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, or 40%.In some embodiments, the ORR is at least about 20%, or at least about 21%, or at least about 22%, or at least about 23%, or at least about 24%, or at least about 25%, or at least about 26%, or at least about 27%, or at least about 28%, or at least about 29%, or at least about 30%, or at least about 31%, or at least about 32%, or at least about 33%, or at least about 34%, or at least about 35%, or at least is about 36%, or at least about 37%, or at least about 38%, or at least about 39%, or at least about 40%, or at least about 41%, or at least about 42%, or at least about 43%, or at least about 44%, or at least about 45%, or is about 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, 40%, 41%, 42%, 43%, 44%, or 45%.

[0044] In some embodiments, the CR, CR / CRh, CRc, and / or ORR rates are higher for patients with particular genetic characteristics, hi some embodiments, the CR, CR / CRh, CRc, and / or ORR rates are higher in patients with acute leukemia (such as AML) that have an NPM1 mutation than in patients that do not have an NPM1 mutation.

[0045] In some embodiments, the duration of remission (DoR) of patients, such as NPM1-m AML patients, who achieve CRc is at least 2 months, or at least 3 months, or at least 4 months, or at least 5 months, or at least 6 months, or at least 7 months, or at least 8 months, or at least 9 months, or at least 10 months, or at least 11 months, or at least 12 months, or at least about 18 months, or at least about 24 months. In some embodiments, the DoR of NPM1-m patients who achieve CRc is at least about 8 months, or at least about 8.2 months, or about 8.2 months. In some embodiments, the DoR of NPM1-m patients who achieve CRc is at least about 5 months, or at least about 6 months, or at least about 7 months, or about 5.6 months, or about 6.6 months, or about 7.7 months. In some embodiments, the DoR of NPM1-m patients who achieve CRc from administration of 600 mg of diftomenib is about 5-7 months, or about 5.6 months, or about 6.6 months. In some embodiments, the DoR for NPM1-m patients who achieve CR or CRh is about 5-7 months, or about 5.6 months, or about 6.6 months. In some embodiments, the DoR for NPM1-m patients who achieve CR or CRh from administration of 600 mg of diftomenib is 5-6 months, or about 5.6 months. In some embodiments, the DoR for NPM1-m patients treated according to the methods described herein is longer than the DoR for KMT2A-r patients.

[0046] In some embodiments, the median overall survival (OS) is at least about 5 months, or about 5.1 months, or about 5.6 months, or at least 5 months, or at least 5.5 months. In some embodiments, such OS is achieved by NPM1-m patients treated with 600 mg of diftomenib. In some embodiments, the OS of NPM1-m patients treated with 600 mg of diftomenib is longer than that of NPM1-m patients treated with 200 mg of diftomenib.

[0047] In some embodiments, administration results in a transient increase in white blood cell count or peripheral blast count, followed by a decrease in such count. In some embodiments, the transient increase occurs within 14 days of initiating treatment. In some embodiments, the decrease occurs within 14 days of the increase. In some embodiments, provided herein are methods for transiently increasing white blood cell count or peripheral blast count, comprising administering an effective amount of diftomenib or a pharmaceutically acceptable form thereof, for example, 600 mg of diftomenib or a pharmaceutically acceptable form thereof. In some embodiments, the increase from baseline occurs within 1 week, 2 weeks, 3 weeks, or 4 weeks of initiating administration. In some embodiments, the increase from baseline lasts for about 1 week, about 2 weeks, or about 3 weeks.

[0048] safety In some embodiments, the methods provided herein include administering diftomenib or a pharmaceutically acceptable form thereof to an individual, wherein the risk of an individual of one genetic subtype experiencing or developing a particular side effect (or a serious side effect) is reduced compared to another genetic subtype or the entire treated patient population. As used herein, the "risk" of an individual experiencing or developing a particular side effect is determined based on the incidence of the side effect across a modified intent-to-treat (mITT) patient population regardless of dose, or across different genetic cohorts (e.g., NPM1-m compared to KMT2A-r groups), since results have demonstrated that safety outcomes do not perfectly correlate with dose (see Example 3).

[0049] In some embodiments, the methods disclosed herein include the feature that an individual's risk of developing a particular adverse event is lower when the individual has acute leukemia with an NPM1 mutation (e.g., AML or ALL), particularly AML, than when the individual does not have an NPM1 mutation. In some embodiments, the risk of developing a Grade 3 or higher (i.e., Grade 3, 4, or 5) treatment-emergent adverse event (TEAE) regardless of causality, a serious adverse event regardless of causality, an adverse event suspected of differentiation syndrome, differentiation syndrome, or severe differentiation syndrome is lower in individuals with acute leukemia containing an NPM1 mutation than in individuals with acute leukemia not containing an NPM1 mutation.

[0050] In some embodiments, the method comprises administering diftomenib or a pharmaceutically acceptable form thereof, wherein the risk of an individual receiving diftomenib experiencing any Grade 3 or higher treatment-emergent adverse event (TEAE), regardless of causality, after administration is less than about 80%, or less than about 75%, or about 71%. In some embodiments, the method comprises administering diftomenib or a pharmaceutically acceptable form thereof, wherein the risk of an individual receiving diftomenib experiencing any serious adverse event, regardless of causality, after administration is less than about 65%, or less than about 60%, or less than about 55%, or about 53%.

[0051] In some embodiments, the method includes administering diftomenib or a pharmaceutically acceptable form thereof, wherein the risk of an individual experiencing any adverse event suspected of differentiation syndrome after administration is less than about 80%, or less than about 75%, or less than about 70%, or less than about 65%, or less than about 60%, or less than about 55%, or less than about 50%, or about 47%. As used herein, a "suspected adverse event of differentiation syndrome" is a differentiation syndrome, a treatment-emergent adverse event (TEAE) that meets the Norsworthy criteria for a potential differentiation syndrome, or a TEAE that meets the Norsworthy criteria for a differentiation syndrome that cannot be excluded (Norsworthy et al., Clin. Cancer Res. 2020, 26(16), 4280-4288).

[0052] In some embodiments, the method includes administering diftomenib or a pharmaceutically acceptable form thereof, and the risk of the individual developing differentiation syndrome after administration is less than about 25%, or less than about 20%, or less than about 19%, or less than about 18%, or about 18%. In some embodiments, the probability that the individual develops differentiation syndrome is severe differentiation syndrome of grade 3, 4, or 5 is less than about 50%, or less than about 45%, or less than about 40%, or less than about 35%, or about 33%.

[0053] In some embodiments, the method comprises administering diftomenib or a pharmaceutically acceptable form thereof, wherein the risk of the individual developing severe differentiation syndrome is less than about 20%, or less than about 15%, or less than about 10%, or less than about 7%, or about 6%. As used herein, "severe differentiation syndrome" is defined as differentiation syndrome at Grade 3, Grade 4, or Grade 5 according to the National Cancer Institute's standardized Common Toxicity Criteria for Adverse Events (available at http: / / ctep.cancer.gov / protocolDevelopment / electronic_applications / ctc.htm).

[0054] In some embodiments, the method includes administering diftomenib or a pharmaceutically acceptable form thereof, wherein the individual develops differentiation syndrome after administration, and the probability that the differentiation syndrome is not a severe differentiation syndrome (e.g., is Grade 1 or 2) is greater than about 50%, or greater than about 55%, or greater than about 60%, or greater than about 65%, or about 67%.

[0055] In some embodiments, the methods provided herein comprise administering diftomenib or a pharmaceutically acceptable form thereof without inducing QTc prolongation.

[0056] Differentiation disorders In preclinical studies, diftomenib was found to promote terminal differentiation and scheduled apoptosis. One potentially severe sequela is differentiation syndrome (DS), which can be life-threatening or fatal if untreated. Differentiation syndrome is prominent in patients treated with isocitrate dehydrogenase (IDH) inhibitors (Norsworthy, 2020), and some fatal outcomes have been reported in patients after diftomenib administration. Increased recognition of the signs and symptoms of DS via the Montesinos framework may lead to earlier diagnosis and treatment, and reduced severe morbidity and mortality. Montesinos et al. (Blood 2009, 113(4), 775–783) proposed diagnostic criteria for DS based on at least two of the following signs and symptoms: dyspnea, unexplained fever, weight gain, unexplained hypotension, acute kidney injury, and pulmonary infiltrates or pleuropericardial effusion. Patients with two or three criteria were classified as having moderate DS, and patients with at least four criteria were classified as having severe DS.

[0057] As used herein, "differentiation disorder" refers to differentiation syndrome (with or without leukocytosis), leukocytosis, and tumor lysis syndrome.Leukocytosis can be detected based on an increase in white blood cell count in the absence of infection.Tumor lysis syndrome can occur in the context of rapidly progressive leukocytosis and can be detected by the presence of two or more blood chemistry markers selected from hyperuricemia, hyperkalemia, hyperphosphatemia, and hypocalcemia, or by an increase in serum creatinine level.

[0058] The following adverse events were monitored and evaluated for treated patients using an algorithm based on the Montesinos criteria as described by Norsworthy (2020), and diagnostic criteria were applied as described below in patients who received at least one dose of diftomenib:

[0059] A differentiation disorder such as DS, with or without leukocytosis, may be suspected based on one or more of the following symptoms: New or worsening progressive dyspnea or hypoxia with an increased need for supplemental oxygen and no clear alternative etiology Radiological evidence of new or worsening pulmonary infiltrates not attributable to another cause, Radiological evidence of new or worsening pleural or pericardial effusion without definitive etiology or refractory to treatment of the first suspected cause, New or worsening peripheral edema without a definitive etiology and accompanied by rapid weight gain (e.g., more than 5 kg over 7 days), Acute renal failure not attributable to other causes or medication (e.g., serum creatinine increase more than twofold from baseline), · Unexplained fever of 38°C (100.4°F) or higher, -Unexplained low blood pressure, Significant changes in inflammatory markers, Evidence of multiple organ dysfunction, or Rash, joint pain, bone pain, or swelling of extramedullary lesions.

[0060] The presence of two or more of the above signs and symptoms can be considered potential DS. Patients with two or three criteria are classified as having moderate DS, and patients with at least four criteria are classified as having severe DS.

[0061] In particular, the development of differentiation syndrome in patients with AML with NPM1 mutations has been found to correlate with a favorable outcome. In some embodiments, treated individuals develop differentiation syndrome, optionally a severe differentiation syndrome (e.g., grade 3, 4, or 5), but may achieve an ORR of at least about 60%, or at least about 65%, or at least about 70%, or at least about 75%, or about 65%, 66%, 67%, 68%, 69%, 70%, 71%, 72%, 73%, 74%, or 75%.

[0062] It has also been found that treating patients with diftomenib or its pharmaceutically acceptable form increases the level of myeloid blasts in the blood of treated individuals.Therefore, provided herein is a method for increasing the level of myeloid blasts in the blood of individuals with AML, comprising administering an effective amount of a menin inhibitor, particularly 600 mg of diftomenib or its pharmaceutically acceptable form, to individuals daily, wherein optionally, the AML is menin-dependent, or optionally, the AML comprises NPM1 mutation, KMT2A rearrangement, KMT2A-PTD mutation, SETD2 mutation, RUNX1 mutation, FLT3 mutation (for example, FLT3-ITD mutation or FLT3-TKD mutation), IDH mutation (for example, IDH1 mutation or IDH2 mutation), TERT mutation, or BRAF mutation, or any combination thereof, or optionally, the AML comprises NPM1 mutation, KMT2A rearrangement, SETD2 mutation, or RUNX1 mutation. In some embodiments, the AML comprises an NPM1 mutation or a KMT2A rearrangement. In some embodiments, the AML comprises an NPM1 mutation. In some embodiments, the relative level of myeloid blasts in the individual's blood is assessed by analysis of two time-separated blood samples from the individual, optionally taken at, for example, (a) a first time point and a second time point (both time points during administration), or (b) a first time point before administration begins and a second time point during administration. In some embodiments, the first time point is before administration, e.g., before cycle 1, day 1, and the second time point is at least 7, 14, 21, or 28 days later, or from cycle 1, day 2 through cycle 1, day 28. In some embodiments, each analysis is a complete blood count (CBC), optionally with differential. In some embodiments, the first and second time points are separated by 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, 1 week, 2 weeks, 3 weeks, 4 weeks, at least 1 week, at least 2 weeks, at least 3 weeks, or at least 4 weeks, or the duration of one cycle (such as cycle 1), or the second time point is the time at which the individual achieves complete remission (CR).In some embodiments, the first time point is screening or cycle 1, day 1. In some embodiments, an increase in the level of myeloid blasts in an individual's blood does not correlate with the progression of AML. In some embodiments, an increase in the level of myeloid blasts occurs in the extramedullary space. In some embodiments, an increase in the level of myeloid blasts indicates that the individual is sensitive to a menin inhibitor, particularly diftomenib or a pharmaceutical form thereof.

[0063] Monitoring and treatment of differentiation disorders It has been found that monitoring and early intervention can reduce the incidence and severity of differentiation disorders, leading to fewer adverse events and allowing patients to continue receiving treatment.

[0064] In some embodiments, the methods provided herein include administering an effective amount of a steroid (e.g., dexamethasone) if a differentiation disorder is detected in an individual during administration of diftomenib or a pharmaceutically acceptable form thereof.

[0065] Provided herein is a method for identifying acute leukemia (particularly AML) in an individual as being susceptible to administration of a menin inhibitor, particularly diftomenib, said method comprising: administering to the individual daily an effective amount of a menin inhibitor, particularly 600 mg of diftomenib or a pharmaceutically acceptable form thereof; receiving an identification of the level of myeloid blasts in a first blood sample taken from the individual at a first time point, either before the start of administration or during administration; receiving an identification of the level of myeloid blasts in a second blood sample taken from the individual at a second time point after the first time point and during administration; determining that the acute leukemia is susceptible to the administration if the level of myeloid blasts at the second time point is greater than the level at the first time point; Optionally, the acute leukemia is menin-dependent; or optionally, the acute leukemia comprises an NPM1 mutation, a KMT2A rearrangement, a KMT2A-PTD mutation, a SETD2 mutation, a RUNX1 mutation, a FLT3 mutation (e.g., a FLT3-ITD mutation or a FLT3-TKD mutation), an IDH mutation (e.g., an IDH1 mutation or an IDH2 mutation), a TERT mutation, or a BRAF mutation, or any combination thereof; or optionally, the acute leukemia comprises an NPM1 mutation, a KMT2A rearrangement, a SETD2 mutation, or a RUNX1 mutation, preferably an NPM1 mutation or a KMT2A rearrangement. In some embodiments, the acute leukemia comprises an NPM1 mutation, optionally in combination with a FLT3 mutation (e.g., a FLT3-ITD mutation or a FLT3-TKD mutation) or an IDH mutation (e.g., an IDH1 mutation or an IDH2 mutation), or a combination thereof.

[0066] In some embodiments, methods are provided for treating a differentiation disorder, particularly tumor lysis syndrome, in an individual diagnosed with a differentiation disorder in an individual with acute leukemia, or for reducing the risk of developing a severe differentiation disorder in an individual with acute leukemia, wherein optionally the acute leukemia is menin-dependent; or optionally the acute leukemia comprises an NPM1 mutation, a KMT2A rearrangement, a KMT2A-PTD mutation, a SETD2 mutation, a RUNX1 mutation, a FLT3 mutation (e.g., a FLT3-ITD mutation or a FLT3-TKD mutation), an IDH mutation (e.g., an IDH1 mutation or an IDH2 mutation), a TERT mutation, or a BRAF mutation, or any combination thereof; or optionally the acute leukemia comprises an NPM1 mutation, a KMT2A rearrangement, a SETD2 mutation, or a RUNX1 mutation, preferably an NPM1 mutation or a KMT2A rearrangement; and (a) administering to an individual daily an effective amount of a menin inhibitor, particularly 600 mg of diftomenib or a pharmaceutically acceptable form thereof; (b) administering IV hydration to the individual; and optionally, (c) administering to the individual an effective amount of a xanthine oxidase inhibitor, optionally wherein the xanthine oxidase inhibitor is allopurinol; and optionally administering the allopurinol at a dose of about 200-400 mg / m 2 and administering daily at a dose of 1 to 3 divided doses up to a maximum of about 800 mg / day. In some embodiments, the acute leukemia comprises an NPM1 mutation, optionally in combination with a FLT3 mutation (e.g., a FLT3-internal tandem duplication (ITD) mutation or a FLT3 mutation in the tyrosine kinase domain (FLT3-TKD mutation)) or an IDH mutation (e.g., an IDH1 mutation or an IDH2 mutation), or a combination thereof.

[0067] Some embodiments include administering a xanthine oxidase inhibitor, wherein prior to administering the xanthine oxidase inhibitor, the individual: (1) Approximately 25×10 9 a white blood cell count of less than 100 U / L and a lactate dehydrogenase level less than twice the upper limit of normal (e.g., if the normal level is about 280 U / L), or (2) Approx. 25 to approx. 100 x 10 9 / L white blood cell count, or (3) Approximately 25×10 9 White blood cell count less than / L and lactate dehydrogenase level more than twice the upper limit of normal The method includes diagnosing the individual as having or at low or moderate risk of developing a differentiation disorder, based on receiving an identification that the individual has the disorder.

[0068] Provided herein is a method of treating a differentiation disorder in an individual with acute leukemia, or a method of reducing the risk of developing a severe differentiation disorder in an individual with acute leukemia, wherein optionally the acute leukemia is menin-dependent; or optionally the acute leukemia comprises a mutation selected from an NPM1 mutation, a KMT2A rearrangement, a KMT2A-PTD mutation, a SETD2 mutation, a RUNX1 mutation, a FLT3 mutation (e.g., a FLT3-ITD mutation or a FLT3-TKD mutation), an IDH mutation (e.g., an IDH1 mutation or an IDH2 mutation), a TERT mutation, or a BRAF mutation, or any combination thereof; or optionally the acute leukemia comprises an NPM1 mutation, a KMT2A rearrangement, a SETD2 mutation, or a RUNX1 mutation, preferably an NPM1 mutation or a KMT2A rearrangement; and the method comprises: (a) administering to an individual daily an effective amount of a menin inhibitor, particularly 600 mg of diftomenib or a pharmaceutically acceptable form thereof; (b) administering IV hydration to the individual; (c) administering a therapeutically effective amount of rasburicase, optionally at a dose of about 0.2 mg / kg, optionally as an intravenous infusion over about 30 minutes, for up to about 5 days. In some embodiments, the acute leukemia comprises an NPM1 mutation, optionally in combination with a FLT3 mutation (e.g., a FLT3-internal tandem duplication (ITD) mutation or a FLT3 mutation in the tyrosine kinase domain (FLT3-TKD mutation)) or an IDH mutation (e.g., an IDH1 mutation or an IDH2 mutation), or a combination thereof.

[0069] In some embodiments, prior to administering rasburicase, the individual (i) Approximately 100 × 10 9 have a white blood cell count level of 1 / L or higher, or (ii)(i) about 25 to about 100 × 10 9 / L white blood cell count level, or (ii) approximately 25 × 10 9have a white blood cell count level of less than 1 / L and a lactate dehydrogenase level of more than twice the upper limit of normal (e.g., where normal levels are about 280 units / L); For each of (i) and (ii), the individual has impaired renal function or uric acid, potassium, and / or phosphate levels above the applicable upper normal limit; and diagnosing the individual as having or at risk of developing a differentiation disorder, based on receiving an identification of the individual.

[0070] Provided herein are methods of treating differentiation disorders or reducing the risk of developing severe differentiation disorders in an individual with acute leukemia, optionally wherein the acute leukemia is menin-dependent; or optionally wherein the acute leukemia comprises a mutation selected from an NPM1 mutation, a KMT2A rearrangement, a KMT2A-PTD mutation, a SETD2 mutation, a RUNX1 mutation, a FLT3 mutation (e.g., a FLT3-ITD mutation or a FLT3-TKD mutation), an IDH mutation (e.g., an IDH1 mutation or an IDH2 mutation), a TERT mutation, or a BRAF mutation, or any combination thereof; or optionally wherein the acute leukemia comprises an NPM1 mutation, a KMT2A rearrangement, a SETD2 mutation, or a RUNX1 mutation, preferably an NPM1 mutation or a KMT2A rearrangement; and the method comprises: (a) administering to the individual a prophylactic and / or effective amount of a corticosteroid, optionally the corticosteroid being prednisone (or an equivalent dose of an alternative corticosteroid) at a dose of about 0.5 mg / kg; (b) administering to the individual a therapeutically effective amount of a menin inhibitor. In some embodiments, the acute leukemia comprises an NPM1 mutation, optionally in combination with a FLT3 mutation (e.g., a FLT3-internal tandem duplication (ITD) mutation or a FLT3 mutation in the tyrosine kinase domain (FLT3-TKD mutation)) or an IDH mutation (e.g., an IDH1 mutation or an IDH2 mutation), or a combination thereof.

[0071] In some embodiments in which the corticosteroid and menin inhibitor are administered daily, sequentially, or simultaneously, optionally including administering the first dose of each of the corticosteroid and menin inhibitor on the same or about the same day, or administering the first dose of the corticosteroid on a day before or after the first administration of the menin inhibitor.

[0072] In some embodiments, prior to administration of a corticosteroid or menin inhibitor, the individual receives 5×10 9 have one or more of the following: white blood cell count greater than / L, increased serum creatinine level, significant extramedullary disease, and proliferative acute leukemia.

[0073] In some embodiments, the method comprises administering a corticosteroid daily starting on day 1, administering a menin inhibitor daily starting on the same or a subsequent day, and, if the individual has not been diagnosed with a differentiation disorder (in methods of reducing risk), or if the individual develops a differentiation disorder (in methods of reducing risk or methods of treating), the differentiation disorder is ameliorated and bone marrow blasts are at a level of less than about 5%, decreasing the dose of the corticosteroid after about 28 days of administration of the menin inhibitor.

[0074] Some embodiments involve administering a dose of about 5 mg, 10 mg, or 15 mg, preferably 10 mg, or about 5-10 mg, of dexamethasone intravenously every 12 hours (or an equivalent dose of an alternative oral or IV corticosteroid) to an individual for 1, 2, or 3 days.

[0075] Some embodiments provide for a white blood cell count or leukocyte count of an individual of about 10×10 9 / L or doubles within about 24 to 48 hours, administering a therapeutically effective amount of hydroxyurea to the individual, and optionally administering a therapeutically effective amount of cytarabine, idarubicin, or gemtuzumab to the individual.

[0076] Some embodiments include tapering the dose and / or discontinuing administration of the corticosteroid, hydroxyurea, cytarabine, idarubicin, or gemtuzumab upon improvement of the differentiation disorder.

[0077] Some embodiments include discontinuing administration of a menin inhibitor during all or part of the administration of one or more of IV hydration, allopurinol, rasburicase, prednisone, dexamethasone, hydroxyurea, cytarabine, idarubicin, and gemtuzumab, and administering the menin inhibitor to a patient after improvement of the differentiation defect (e.g., white blood cell count is less than about 20×10) with a therapeutically effective dose or a reduced dose of the menin inhibitor. 9 and resuming administration when the blood glucose level drops below 1 / L.

[0078] In some embodiments, the differentiation disorder is differentiation syndrome with or without leukocytosis, hi some embodiments, the differentiation disorder is tumor lysis syndrome.

[0079] In some embodiments, the acute leukemia comprises AML. In some embodiments, the acute leukemia comprises ALL. In some embodiments, the acute leukemia comprises an NPM1 mutation. In some embodiments, the acute leukemia is AML comprising an NPM1 mutation.

[0080] definition The compounds of the present disclosure include crystalline and amorphous forms of these compounds, pharmaceutically acceptable salts of these compounds and active metabolites thereof having the same type of activity, including, for example, polymorphs, pseudopolymorphs, solvates, hydrates, nonsolvated polymorphs (including anhydrates), stereopolymorphs, and amorphous forms of the compounds, as well as mixtures thereof.

[0081] The compounds described herein may exhibit their natural isotopic abundance, or one or more atoms may be artificially enriched with a particular isotope having the same atomic number but a different atomic mass or mass number than that predominantly found in nature. All isotopic variations of the compounds of the present disclosure, whether radioactive or not, are encompassed within the scope of the present disclosure. For example, hydrogen has three naturally occurring isotopes, designated 1H (protium), 2H (deuterium), and 3H (tritium). Protium is the most abundant isotope of hydrogen in nature. Deuterium enrichment may provide certain therapeutic advantages, such as increased in vivo half-life and / or exposure, or provide compounds useful for investigating in vivo pathways of drug excretion and metabolism. Isotopically enriched compounds may be prepared by conventional techniques well known to those skilled in the art.

[0082] The term "isotopically enriched" refers to an isotopically enriched compound. The term "isotopically enriched" refers to an atom having an isotopic composition other than the natural isotopic composition of that atom. An "isotopically enriched" can refer to a compound containing at least one atom having an isotopic composition other than the natural isotopic composition of that atom. The term "isotopic composition" refers to the amount of each isotope present for a given atom. Radiolabeled and isotopically enriched compounds are useful as therapeutic agents, e.g., multiple myeloma therapeutic agents, research reagents, e.g., binding assay reagents, and diagnostic agents, e.g., in vivo imaging agents. Isotopic variations of the compounds described herein, whether radioactive or not, are intended to be encompassed within the scope of the embodiments provided herein.

[0083] "Isomers" are different compounds with the same molecular formula. "Stereoisomers" are isomers that differ only in the way their atoms are arranged in space. "Enantiomers" are a pair of stereoisomers that are non-superimposable mirror images of each other. A 1:1 mixture of a pair of enantiomers is a "racemic" mixture. The term "(±)" is used to designate a racemic mixture when appropriate. "Diastereoisomers" or "diastereomers" include stereoisomers that have at least two asymmetric atoms but are not mirror images of each other. Absolute stereochemistry is designated according to the Cahn-Ingold-Prelog RS system. When a compound is a pure enantiomer, the stereochemistry at each chiral carbon can be designated as either R or S. Resolved compounds whose absolute configuration is unknown can be designated as (+) or (-) depending on the direction (dextrorotatory or levorotatory) they rotate plane-polarized light at the wavelength of the sodium D line. Certain compounds described herein contain one or more asymmetric centers and can thus give rise to enantiomers, diastereomers, and other stereoisomeric forms, which can be defined, in terms of absolute stereochemistry, as (R)- or (S)-. The subject chemical entities, pharmaceutical compositions, and methods are meant to include all such possible stereoisomers, including racemic mixtures, optically pure forms, mixtures of diastereomers, and intermediate mixtures. Optically active (R)- and (S)-isomers can be prepared using chiral synthons or chiral reagents or resolved using conventional techniques. The optical activity of a compound can be analyzed via appropriate methods, including, but not limited to, chiral chromatography and polarimetry, to determine the degree to which one stereoisomer predominates over another.

[0084] Chemical entities having carbon-carbon or carbon-nitrogen double bonds can exist in Z or E forms (or cis or trans forms). Additionally, some chemical entities can exist in various tautomeric forms. Unless otherwise specified, chemical entities described herein are intended to include all Z, E, and tautomeric forms as well.

[0085] The term "solvate" generally refers to a compound (e.g., a free base) or a salt thereof that further contains a stoichiometric or non-stoichiometric amount of solvent bound by non-covalent intermolecular forces. When the solvent is water, the solvate is a hydrate.

[0086] The term "salt" or "pharmaceutically acceptable salt" refers to salts derived from various organic and inorganic counterions well known in the art. Pharmaceutically acceptable acid addition salts can be formed with inorganic and organic acids. Inorganic acids from which salts can be derived include, for example, hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, etc. Organic acids from which salts can be derived include, for example, acetic acid, propionic acid, glycolic acid, pyruvic acid, oxalic acid, maleic acid, malonic acid, succinic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, salicylic acid, etc. Pharmaceutically acceptable acid addition salts can be formed with inorganic and organic bases. Inorganic bases from which salts can be derived include, for example, sodium, potassium, lithium, ammonium, calcium, magnesium, iron, zinc, copper, manganese, aluminum, etc. Organic bases from which salts may be derived include, for example, primary, secondary, and tertiary amines, naturally occurring substituted amines, cyclic amines, substituted amines including basic ion exchange resins, and the like, specifically isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, and ethanolamine. In some embodiments, the pharmaceutically acceptable base addition salts are selected from ammonium, potassium, sodium, calcium, and magnesium salts.

[0087] The term "pharmaceutical composition" generally refers to a composition comprising a menin inhibitor, particularly diftomenib, in combination with at least one additional pharmaceutically acceptable carrier. "Pharmaceutically acceptable carrier" refers to a vehicle generally accepted in the art for delivering biologically active agents to individuals, including, for example, adjuvants, excipients, or vehicles, such as diluents, preservatives, fillers, flow regulators, disintegrants, wetting agents, emulsifiers, suspending agents, sweeteners, flavoring agents, fragrances, antibacterial agents, antifungal agents, lubricants, and dispensing agents, depending on the mode of administration and the nature of the dosage form. Suitable carriers include, but are not limited to, any adjuvant, carrier, excipient, glidant, sweetener, diluent, preservative, dye, coloring agent, flavor enhancer, surfactant, wetting agent, dispersant, suspending agent, stabilizer, isotonic agent, solvent, or emulsifier approved by the US Food and Drug Administration for use in humans or livestock.

[0088] Pharmaceutical compounds are formulated according to several factors within the skill of those skilled in the art. These include, but are not limited to, the type and nature of the active agent to be formulated, the subject to which the drug-containing composition will be administered, the intended route of administration of the composition, and the targeted therapeutic indication. Pharmaceutically acceptable carriers include both aqueous and non-aqueous liquid media, as well as various solid and semi-solid dosage forms. Such carriers can contain several different components and additives in addition to the active agent, and such additional components are included in the formulation for various reasons, such as stabilizing the active agent, binders, etc., as known to those skilled in the art. Descriptions of suitable pharmaceutically acceptable carriers and the factors involved in their selection can be found in various readily available sources, such as Allen, LV, Jr. et al., Remington: The Science and Practice of Pharmacy (Volume 2), 22nd Edition, Pharmaceutical Press (2012).

[0089] As used herein, "treatment" or "treating" refers to an approach for obtaining a beneficial or desired result with respect to a disease, disorder, or medical condition (e.g., AML), including, but not limited to, in certain instances, therapeutic benefit and / or prophylactic benefit. Therapeutic benefit refers to the eradication or amelioration of the underlying disease being treated. Therapeutic benefit is further achieved by the eradication or amelioration of one or more physiological symptoms associated with the underlying disease, such that an improvement is observed in an individual, even though the individual may still be affected by the underlying disease. In certain embodiments, with regard to prophylactic benefit, a composition is administered to an individual at risk of developing a particular disease or who reports one or more physiological symptoms of the disease, even if a diagnosis of the disease has not been made.

[0090] A "therapeutic benefit," as that term is used herein, encompasses therapeutic benefits and / or prophylactic benefits, as described above. A prophylactic benefit includes delaying or eliminating the appearance of a disease or condition, delaying or eliminating the onset of symptoms of a disease or condition, slowing, halting, or reversing the progression of a disease or condition, or any combination thereof.

[0091] As used herein, the term "effective amount" in reference to a compound means an amount capable of treating, preventing, or managing a disorder, disease, or condition, or one or more symptoms thereof.

[0092] As used herein, the term "prophylactic amount" in reference to a compound means an amount that is capable of preventing a disorder, disease, or condition, or one or more symptoms thereof, or reducing the ultimate severity of a disorder, disease, or condition, or one or more symptoms thereof.

[0093] As used herein, the term "optimal biological dose" or "OBD" means the lowest dose of a drug, such as an investigational drug, that provides the highest rate of efficacy while being safely administered.

[0094] As used herein, the term "recommended Phase 2 dose" means a dose of an investigational drug that has been approved by a governmental authority (e.g., the U.S. Food and Drug Administration (FDA) or a similar agency in another country) for evaluation in a Phase 2 trial.

[0095] As used herein, the term "safe and effective dose" means a dose of a drug, such as an investigational drug, that produces a clinical effect without unacceptable side effects.

[0096] As used herein, the term "maximum tolerated dose" means the highest dose of a drug, such as an investigational drug, that does not cause unacceptable side effects, e.g., where the observed toxicity rate (e.g., dose-limiting toxicity rate) is less than 0.33.

[0097] "Breakthrough therapy designation" refers to a designation given by a government authority (e.g., the FDA or a similar agency in another country) to an active pharmaceutical ingredient that treats a serious or life-threatening condition and for which preliminary clinical evidence (e.g., Phase 1 clinical data) indicates that the active pharmaceutical ingredient may demonstrate substantial improvement in a clinically significant endpoint over available therapies. In the United States, the status is called Breakthrough Therapy Designation, and in Europe, the status is Priority Medicine (PRIME). As used herein, Breakthrough Therapy Designation may be given for a particular indication and / or genetic subtype, such as NPM1-m AML.

[0098] An "investigational drug" is a substance that has been tested in laboratory experiments and approved by a governmental authority (e.g., the FDA or similar agency in other countries) for testing in humans.

[0099] "Chemotherapy" refers to the administration of one or more chemotherapeutic and / or other agents to an individual by various methods, including intravenous, oral, intramuscular, intraperitoneal, intravesical, subcutaneous, transdermal, buccal, or inhaled, or in the form of a suppository. In the context of acute leukemia, chemotherapy is intensive chemotherapy involving a combination of an anthracycline, e.g., daunorubicin or idarubicin, with cytarabine in a "7+3" regimen (cytarabine is continued for 7 days with short infusions of the anthracycline each day for the first 3 days).

[0100] "Individual" refers to an animal, such as a mammal, e.g., a human. The methods described herein may be useful for both human therapy and veterinary applications. In some embodiments, the individual is a mammal, and in some embodiments, the individual is a human. "Mammal" includes both humans and domestic animals, such as laboratory animals and household pets (e.g., cats, dogs, pigs, cows, sheep, goats, horses, rabbits), and non-domestic animals, such as wildlife. In some embodiments, the human is 18 years of age or older. In some embodiments, the human is under 18 years of age, under 12 years of age, or under 6, 5, 4, 3, 2, or 1 year of age.

[0101] Clinical terms used herein include the following: "CR" means complete remission, with no visible evidence of leukemia cells in the blood or bone marrow, normal bone marrow function, and a normal number of healthy blood cells returning to circulation as confirmed by bone marrow biopsy and blood tests; CR rate is defined as the population of patients achieving a best overall response of CR; "CRh" means complete response with hematologic recovery; CR / CRh response rate is defined as the proportion of patients achieving a best overall response of CR, with or without MRD or CRh; "CRi" means complete response with incomplete hematologic recovery; "CRp" means complete remission with incomplete platelet recovery; CRc means combined complete remission; response rate is defined as the proportion of patients achieving a best overall response of CRi (including CRp), CRh, or CR (including MRD-); and "MRD" means measurable residual disease, referring to a level of leukemia that is not easily observed under a microscope but can be detected by laboratory methods. CR may or may not have measurable disease (CR MRD+ / MRD-), "MLFS" means morphological leukemia-free state, "PR" means partial response, "SD" means stable disease without progression, and "ORR" means overall response rate, determined by the formula ORR=CR (including CR MRD-) + CRh + MLFS (including CRp).

[0102] "BC" means blast count and refers to the percentage of blasts in the bone marrow or blood. In normal bone marrow, the blast count is 5% or less, while the blood usually contains no blasts. A level of at least 20% blasts in the bone marrow or blood usually indicates a diagnosis of AML.

[0103] Hydroxyurea, or hydrea, is an antimetabolite that prevents the overproduction of blood cells in proliferative diseases and is useful in AML to reduce elevated levels of leukemic white blood cells.

[0104] "DS" refers to differentiation syndrome, a potentially serious side effect that can occur in patients with acute leukemia, such as AML, treated with certain anticancer drugs. Differentiation syndrome usually occurs within one to two weeks of treatment. It is caused by the massive and rapid release of cytokines from leukemia cells affected by the anticancer drugs. Signs and symptoms of differentiation syndrome include fever, cough, difficulty breathing, weight gain, swelling of the arms, legs, and neck, excess fluid accumulation around the heart and lungs, low blood pressure, and kidney failure.

[0105] "SCT" or "HSCT" means hematopoietic stem cell transplantation. In some embodiments of the methods provided herein, the subject undergoes SCT after treatment or administration of diftomenib or a pharmaceutically acceptable form thereof. In some embodiments of the methods provided herein, the subject undergoes SCT before treatment or administration of diftomenib or a pharmaceutically acceptable form thereof. In some embodiments, diftomenib or a pharmaceutically acceptable form thereof is administered to the subject before and after SCT (e.g., as maintenance therapy).

[0106] Extramedullary hematopoiesis is the formation and activation of blood cells outside the bone marrow in response to hematopoietic stress caused by leukemia.

[0107] The terms "co-administration," "administered in combination with," and their grammatical equivalents, as used herein, encompass the administration of two or more agents to an animal, including a human, such that both agents and / or their metabolites are present in the individual at the same time. Co-administration includes simultaneous administration in separate compositions, administration at different times in separate compositions, or administration in a composition in which both agents are present.

[0108] The terms "antagonist" and "inhibitor" are used interchangeably and refer to compounds capable of inhibiting the biological function (e.g., activity, expression, binding, protein-protein interaction) of a target protein (e.g., menin, MLL1, MLL2, and / or MLL fusion protein). Thus, the terms "antagonist" and "inhibitor" are defined in the context of the biological role of the target protein. Preferred antagonists herein specifically interact (e.g., bind) with the target; however, specifically included within this definition are compounds that inhibit the biological activity of a target protein by interacting with other members of a signal transduction pathway in which the target protein is a member. A preferred biological activity inhibited by an antagonist is associated with tumor progression, growth, or spread.

[0109] The term "agonist," as used herein, refers to a compound capable of initiating or enhancing the biological function of a target protein, whether by inhibiting the activity or expression of the target protein or not. Thus, the term "agonist" is defined in the context of the biological role of the target polypeptide. Preferred agonists herein specifically interact (e.g., bind) with the target, although specifically included within this definition are compounds that initiate or enhance the biological activity of the target polypeptide by interacting with other members of a signal transduction pathway of which the target polypeptide is a member.

[0110] "Signal transduction" is the process by which stimulatory or inhibitory signals are transmitted to and within cells to induce intracellular responses. A modulator of a signal transduction pathway refers to a compound that regulates the activity of one or more cellular proteins that map to the same specific signal transduction pathway. A modulator can increase (agonist) or inhibit (antagonist) the activity of a signal transduction molecule.

[0111] As used herein, "sample" includes and / or refers to any fluid or liquid sample that is analyzed to detect and / or quantify an analyte. In some embodiments, the sample is a biological sample. Examples of samples include, but are not limited to, bodily fluids, extracts, solutions containing protein and / or DNA, cell extracts, cell lysates, or tissue lysates. Non-limiting examples of bodily fluids include urine, saliva, blood, serum, plasma, cerebrospinal fluid, tears, semen, sweat, pleural effusion, liquefied feces, and lacrimal gland secretions.

[0112] The term "in vivo" refers to events that take place inside an individual's body.

[0113] The term "in vitro" refers to an event that occurs outside of an individual's body. For example, an in vitro assay includes any assay performed outside of an individual. In vitro assays include cell-based assays in which live or dead cells are utilized. In vitro assays also include cell-free assays in which intact cells are not utilized.

[0114] As used herein, "comprising" (and any form of "comprising", 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 inclusive or open-ended and does not exclude additional, unrecited elements or process steps. Similarly, as used herein, in any instance or embodiment described herein, "comprising" may be interchanged with "consisting essentially of" and / or "consisting of" as used herein, and in any instance or embodiment described herein, "comprises" may be interchanged with "consists essentially of" and / or "consists of."

[0115] As used herein, the term "and / or" shall be construed as a specific disclosure of each of the two specified features or components, regardless of the presence or absence of the other. For example, "A and / or B" shall be construed as a specific disclosure of each of (i) A, (ii) B, and (iii) A and B, as if each were individually set forth herein.

[0116] As used herein, the term "about," when used in connection with a dose, amount, or weight percent, means a dose, amount, or weight percent that is within 10%, or within 5%, or within 2%, or within 1% of the stated amount.

[0117] When numerical values ​​are used herein, such values ​​can encompass a range that is ±5% of the stated numerical value. [Example]

[0118] The following examples are included for illustrative purposes only and are not intended to limit the scope of the disclosure.

[0119] Example 1 - Analysis of diftomenib concentrations in tissues and plasma in mice Diftomenib concentrations in plasma, bone marrow, heart, and spleen were measured in female C57BL / 6 mice after 8 days of oral gavage administration of diftomenib. Groups of 5 animals were treated with diftomenib 100 mg / kg PO daily for 8 days. Bone marrow, spleen, plasma, and heart were sampled 24 hours after administration on the 8th day. Each sample was treated as follows: Blood collection and processing: Each blood collection (approximately 0.03 mL) was performed from the saphenous vein or other appropriate site of each animal into pre-chilled commercial EDTA-K2 tubes containing 2 μL of 0.05 M EDTA-K2 as an anticoagulant or pre-chilled plastic microcentrifuge tubes and placed on wet ice until centrifugation. Blood samples were processed for plasma by centrifugation at 3,200 × g for 10 minutes at approximately 4 °C. Plasma was collected, transferred to pre-labeled 96-well plates or polypropylene tubes, flash-frozen on dry ice, and kept at -60 °C or below until analysis. Analysis was performed by LC-MS / MS. Tissue processing: Tissue samples for drug level analysis were homogenized on wet ice using homogenization buffer (MeOH / 15 mM PBS 1:2) at a ratio of 1:9 (1 g tissue with 9 mL buffer). Tissue-homogenates were stored below -60°C until analysis. Analysis was performed by LC-MS / MS. As shown in Figure 1, the analysis showed that diftomenib accumulated at higher levels in tissues compared to plasma.

[0120] Example 2 - Plasma protein binding of diftomenib Diftomenib or the positive control warfarin was spiked into plasma from CD-1 mice and humans. Diftomenib was tested at concentrations of 0.2 μM, 2 μM, and 10 μM, and warfarin was tested at 2 μM. Spiked plasma samples were pre-incubated at 37 ± 1°C and then centrifuged to pellet plasma proteins and allow their separation from free compounds in the supernatant. The concentrations of diftomenib and warfarin in the spiked plasma and supernatant samples were determined using LC-MS / MS. The % unbound, % bound, and % remaining were calculated according to the following formula:

[0121]

number

[0122] Example 3 - Phase 1 / 2 First-in-Human Study of the Menin-MLL (KMT2A) Inhibitor KO-539 (Diftomenib) in Patients with Relapsed or Refractory (R / R) Acute Myeloid Leukemia (AML) (NCT04067336) A. Background KO-MEN-001 is a first-in-human, open-label, multi-cohort study of diftomenib in adult patients with relapsed / relapsed AML, including an initial dose-escalation portion (Phase 1a) and a dose-validation / cohort expansion portion (Phase 1b).

[0123] This phase 1a dose-escalation trial enrolled 30 adult patients with relapsed / relapsed AML (all-comer population), regardless of genotype. Patients received diftomenib at 50 mg (1), 100 mg (1), 200 mg (6), 400 mg (5), 600 mg (5), 800 mg (11), or 1000 mg (1) once daily to evaluate safety, tolerability, and antileukemic activity (CR / CRh, CR with and without measurable residual disease (MRD), duration of remission (DOR), event-free survival, and overall survival (OS)) across a range of diftomenib dose levels and determine the maximum tolerated dose (MTD) and / or recommended phase 2 dose (RP2D) (optimal biologically effective dose) of diftomenib in patients with relapsed / relapsed AML, regardless of genotype. The median age of the subjects was 65.5 years (range, 22-85); 33% (10) had KMT2A-r AML and 13% (4) had NPM1-m AML (non-KMT2A-r / NPM1-m, 16 (53%)). Patients were heavily pretreated, with a median of 3.5 prior lines of therapy (range, 1-9); most had received prior venetoclax; and 23% had received more than one prior stem cell transplant (SCT).

[0124] The Phase 1b dose-finding portion of the study investigated the two lowest doses from the Phase 1a portion that demonstrated meaningful activity in KMT2A-r or NPM1-m AML: 200 mg and 600 mg diftomenib once daily, to explore the possibility of optimal biologically active doses for these patient populations. Among the first 36 enrolled subjects (17 NPM1-m, 19 KMT2A-r, plus co-mutations involving IDH and FLT3, which are dominant in NPM1-m patients), the median age was 52 years (range 28-84), and approximately one-third had undergone at least one prior SCT. Patients had a median number of prior lines of therapy of three (range, e.g., 1-12 for KMT2A-r and 2-8 for NPM1-m for the first 24 patients). 66.7% of patients had been treated with venetoclax prior to enrollment, and 30.6% had undergone SCT.

[0125] Demographic and clinical information for the first 53 cumulative patients in Phase 1b is shown in Table 1.

[0126] [Table 1]

[0127] For the 53 patients in Phase 1b, either the patients discontinued the study within the first cycle or received treatment for at least one cycle, and the patient dispositions are shown in Table 2.

[0128] [Table 2] Approximately 30% of patients were still undergoing treatment, and nearly 50% were still under observation. Most patients discontinued, often due to progression of their underlying disease. Six patients discontinued due to adverse events, none of which were considered treatment-related.

[0129] In both parts, diftomenib was administered orally once daily in 28-day cycles.

[0130] Study participants - analyzed population The population for clinical safety and efficacy analyses in this Phase 1 study was the modified intent-to-treat (mITT) population: all patients who received at least one dose of study drug. Patients were grouped into the mITT population according to treatment cohort and tumor genetics. The following subgroups were used for analysis: KMT2A-r subgroup of the 200 mg and 600 mg dose cohorts in the mITT Phase 1a and 1b combination NPM1-m subgroups of the 200mg and 600mg dose cohorts in the mITT Phase 1a and 1b combination trials.

[0131] Where appropriate, data are collected in this example from the following patient subgroupings from the analysis population: Group 1: Phase 1a (all patients in the mITT set at the time of data cut) Group 2: Phase 1b (all patients in the mITT set at the time of data cut) Group 3: Phase 1a on-target mutation patients (e.g., patients with KMT2A-r or NPM1-m AML) plus all patients in the Phase 1b mITT set at data cut (all on-target cohorts) Group 4: Phase 1a on-target mutation patients plus the first 24 patients (or up to a total of 53, if identified) enrolled in Phase 1b (on-target maturation cohort) who have been followed for a minimum of 2 months will be presented. These patient groupings allow for evaluation of safety in patients with and without mutational status (Group 1), safety and efficacy in the on-target patient population (Groups 2-4), and efficacy signals in patients who have had sufficient treatment time to support the determination of RP2D and allow for potential clinical benefit / response (Group 4).

[0132] B.Antileukemia activity In the phase 1a portion of the study, clinical benefit was demonstrated across all genotypes evaluated, with 30% of R / R AML patients continuing treatment for at least four cycles. Disease control (e.g., reduction in blast count [BC] or reduced hydroxyurea requirements) and overall improvements in patient performance status were observed across all dose levels. At 100 mg, one complete remission (CR) was observed in a patient with SETD2 and RUNX1 mutations, indicating the possibility that other AML genotypes may be dependent on the menin pathway. At 200 mg, two NPM1-vn patients responded positively, one patient experienced a [MRD-] CR without measurable residual disease for a duration of at least 100 weeks, and one patient with NPM1-m and FLT3-ITD mutations achieved a morphological leukemia-free state [MLFS]. At the 600 mg and 800 mg doses, KMT2A-r patients experienced stable disease with significant reductions in BC and significant duration of improvement in performance status (e.g., one patient's response lasted more than 4 months). Clinical benefits observed from the Phase 1a portion included 29% continuing treatment for at least four cycles, reductions in blast counts (RUNX1 mutations), reduced hydroxyurea (Hydrea) requirements (one KMT2A-r subject), and overall improvement in patient performance status across dose levels.

[0133] Clinical efficacy in the phase 1b segment of the study was dose-dependent. Response rates consistently demonstrated increased antileukemic activity with the 600 mg dose QD compared with the 200 mg dose QD. The complete response (CR) / CR and composite CR rates with partial hematologic recovery (CRh) in on-target patients in phase 1a and the first 24 patients enrolled in phase 1b were 21.4% and 6.7% (1a) and 28.6% vs. 6.7% (1a and 24 1b), respectively, at 600 mg vs. 200 mg. At 200 mg, changes in bone marrow morphology and stable blast counts / decreased blast counts were observed. Three patients were dose-escalated to 600 mg with improvement. One patient achieved a bone marrow blast count <5% and a significant reduction in heavy extramedullary disease, although small foci of disease persisted; one patient achieved a significant reduction in BC and disease control; and one patient achieved minimal hematologic recovery and continued treatment. At 600 mg, 25% of the first 24 Phase 1b patients had a best response of CR or CR with partial hematologic recovery (CRh), and 33.3% of NPM1-m patients achieved CR or CRh. For the first 24 Phase 1b patients, the composite CR was 33%, 75% had measurable residual disease (MRD), and the overall response rate (ORR) was 42%.

[0134] As of October 2022, Phase 1b enrollment was complete (N=53), with 30% (including 21% at the 600 mg dose) still on ongoing treatment with progressive clinical benefit. The 200 mg dose demonstrated some efficacy in the NPM1-m patient population, while KMT2A-r patients did not demonstrate any formal responses. Overall efficacy signals are shown in Table 3, with an overall CR / CRh of 21.2%, CRc of 26.3%, and ORR of 28.9% for the pooled group. Within this expanded patient group, a total of 20 NPM1-m patients were enrolled and treated with diftomenib 600 mg. This subgroup continued to demonstrate strong evidence of antileukemic activity, with CR / CRh of 30.0%, CRc of 35.0%, and ORR of 40.0%. Forty-three percent of patients who achieved CRc also achieved MRD negativity, although only five of these patients were tested for MRD. Of the patients tested, 60% were MRD-negative. Of the CR / CRh patients, two-thirds had IDH and / or FTL3 co-mutations. Overall, of the seven patients with IDH co-mutations, 57% had a CR to diftomenib. Furthermore, across groups, patients who experienced differentiation syndrome had an ORR of 75%. Significant activity was observed in KMT2A-r patients, but a formal CR / CRh response was achieved in only one patient, with an ORR of 16.7%.

[0135] [Table 3]

[0136] C. Safety and Tolerability Safety and tolerability by dose There was no apparent relationship between diftomenib dose and the rate of treatment-emergent adverse events (TEAEs), regardless of severity, seriousness, or relevance. Overall, events were consistent with the effects of the underlying disease. The incidence of TEAEs and serious adverse events (SAEs) is shown in Table 4. Although there were numerical differences between dose groups, there was no significant increase in the rate or severity of AEs with increasing dose.

[0137] [Table 4]

[0138] The most frequent TEAEs reported by >15% of patients across Phase 1a and Phase 1b were similar and included diarrhea, anemia, nausea, increased blood creatinine, fatigue, pneumonia, increased alanine aminotransferase, arthralgia, increased aspartate aminotransferase, decreased appetite, peripheral edema, fever, anemia, epistaxis, febrile neutropenia, hypomagnesemia, and differentiation syndrome.

[0139] TEAEs by severity (grade 3 or higher) In Phase 1a, 11 of 11 patients (100%) reported at least one TEAE, and 26 of 30 patients (86.7%) reported a grade 3 or higher TEAE, regardless of causality. The most common grade 3 or higher TEAEs were anemia (8 patients [26.7%]), pneumonia (7 patients [23.3%]), followed by febrile neutropenia, neutropenia, thrombocytopenia, and decreased appetite (3 patients [10.0%] each). Eight patients (26.7%) reported a grade 3 or higher TEAE considered by the investigator to be related to diftomenib (one each at 50 mg, 100 mg, 200 mg, 400 mg, 600 mg, and 1000 mg, and two at 800 mg). The most common grade ≥3 diftomenib-related TEAE reported in patients in Phase 1a was pulmonary embolism (2 patients [6.7%]). Grade ≥3 TEAEs by dose for Phase 1a are shown in Table 5.

[0140] [Table 5]

[0141] Two dose-limiting toxicities occurred: pneumonitis (400 mg dose) and differentiation syndrome (1000 mg dose). No drug-induced QT / QTc prolongation was reported in Phase 1a. Two DLTs were reported: (1) in the 400 mg cohort (pneumonitis, post-aspiration pneumonia) and (2) in the 1000 mg cohort (differentiation syndrome).

[0142] For the first 24 patients in the phase 1b portion of the study, grade ≥3 TEAEs occurring in ≥10% of all patients (N=24) were anemia, febrile neutropenia, neutropenia, and thrombocytopenia (25% each), differentiation syndrome (DS) and leukocytosis (17% each), and sepsis and leukopenia (13% each). At 200 mg (N=12), grade ≥3 TEAEs occurring in ≥10% of patients were neutropenia and thrombocytopenia (33% each), febrile neutropenia, anemia, and sepsis (25% each), and DS, leukocytosis, and respiratory failure (17% each). At 600 mg (N=12), grade ≥3 TEAEs occurring in ≥10% of patients were febrile neutropenia and anemia (25% each), and DS, leukocytosis, neutropenia, thrombocytopenia, leukopenia, and diarrhea (17% each).

[0143] Among the first 36 patients in Phase 1b, 34 of the 36 patients (94.4%) reported at least one TEAE, and 28 patients (77.8%) reported a grade 3 or higher TEAE regardless of causality. The most common grade 3 or higher TEAEs were febrile neutropenia (seven patients [19.4%]), DS and anemia (six patients [16.7%] each), and pneumonia, sepsis, and decreased platelet count (four patients [11.1%] each). Fifteen patients (41.7%), including seven patients (41.2%) in the 200 mg cohort and eight patients (42.1%) in the 600 mg cohort, reported grade 3 or higher TEAEs considered by the investigator to be related to diftomenib. The most common grade 3 or higher diftomenib-related TEAE reported in Phase 1b patients was DS (six patients [16.7%]). Regardless of causality, SAEs were reported by 23 patients (63.9%), and 11 patients (30.6%) reported SAEs considered by the investigator to be related to diftomenib (5 patients [29.4%] in the 200 mg cohort and 6 patients [31.6%] in the 600 mg cohort). No patients reported TEAEs requiring dose reduction, and the proportion of patients requiring dose interruption or treatment discontinuation was similar between the 200 mg and 600 mg diftomenib dose levels (41.2% vs. 31.6% and 11.8% vs. 10.5%, respectively). Serious adverse events determined to be related to diftomenib included DS (4 patients), increased alanine aminotransferase (1 patient), and pleuritic chest pain (1 patient) at 200 mg, and myocarditis (1 patient), DS (2 patients), febrile neutropenia (2 patients), dyspnea (1 patient), leukocytosis (1 patient), cardiomyopathy (1 patient), diarrhea (1 patient), and dehydration (1 patient) at 600 mg.

[0144] Among the 53 subjects in the Phase 1b portion, grade 3 or higher adverse events, regardless of causality, occurred infrequently in more than 10% of patients. Grade 3 or higher TEAEs occurring in 10% or more of all patients (N=24) were anemia, febrile neutropenia, neutropenia, and thrombocytopenia (25% each), differentiation syndrome and leukocytosis (17% each), and sepsis and leukopenia (13% each). At 200 mg (N=12), grade 3 or higher TEAEs occurring in 10% or more of patients were neutropenia and thrombocytopenia (33% each), febrile neutropenia, anemia, and sepsis (25% each), and differentiation syndrome, leukocytosis, and respiratory failure (17% each). At 600 mg (N=12), grade ≥3 TEAEs occurring in ≥10% of patients were febrile neutropenia and anemia (25% each), and differentiation syndrome, leukocytosis, neutropenia, thrombocytopenia, leukopenia, and diarrhea (17% each).

[0145] No reports of QTc prolongation were noted in Phase 1b, and overall, no clinically meaningful trends in clinical laboratory assessments, vital signs, or ECGs were observed.

[0146] In summary, safety data collected during Phase 1b at the pooled data cutoff of 36 patients were consistent with data from Phase 1a, suggesting comparable safety and tolerability between diftomenib monotherapy doses of 200 mg to 600 mg.

[0147] Genotypic safety and tolerability Safety results for KMT2A-r and NPM1-m AML patients receiving 200 mg or 600 mg across Phase 1a and Phase 1b (Table 6) were generally consistent across study populations, indicating no apparent relationship between diftomenib dose and the overall frequency of TEAEs. The most common grade 3 or higher TEAEs were anemia, febrile neutropenia, pneumonia, sepsis, increased alanine aminotransferase, decreased platelet count, and differentiation syndrome in KMT2A-r patients, and anemia, neutropenia, thrombocytopenia, diarrhea, pneumonia, sepsis, decreased neutrophil count, decreased white blood cell count, and hyperglycemia in NPM1-m patients. However, the risk of serious adverse events was elevated in the KMT2A-r cohort (70.8%) compared with 52.9% in NPM1-r patients. Serious adverse events occurring in ≥10% of all patients included febrile neutropenia (25.0%), pneumonia (12.5%), sepsis (12.5%), and DS (20.8%) with KMT2A-r and Clostridium difficile infection (11.8%), pneumonia (11.8%), and sepsis (11.8%) with NPM1-m, and were not dose-dependent. Dose-limiting toxicities occurred in 30.0% of patients with KMT2A-r and 14.3% of patients with NPM1-m.

[0148] [Table 6]

[0149] For each AE category (TEAEs, grade ≥3 TEAEs, and SAEs), the rate and severity of reported events were consistent across dose groups. Combined with the limited number of TEAE-related dose interruptions and / or discontinuations, the data indicate that diftomenib tolerability did not decrease with increasing dose, with comparable safety and tolerability across diftomenib doses from 200 mg to 600 mg. When assessed by genetic subtype, safety results for KMT2A-r and NPM1-m AML patients receiving 200 mg or 600 mg were generally consistent across the entire study population, indicating no apparent relationship between diftomenib dose and the overall frequency of TEAEs. With the exception of DS, which is discussed further below, reported events appeared to be common in nature or related to the underlying disease.

[0150] ·Differentiation syndrome Given that inhibition of menin activity disrupts gene expression pathways involved in maintaining the pathological dedifferentiated stem cell-like phenotype of KMT2A-r and NPM1-m AML, TEAEs in DS are likely to be treatment-related and result from the effects of diftomenib on cancer cells. DS responses must be carefully managed to ensure patient safety and continued access to treatment.

[0151] In Phase 1a, a single event (grade 4) of on-target DS was reported in a KMT2A-r patient at the 1000 mg dose level, leading to tapering of the 1000 mg dose cohort and completion of Phase 1a enrollment at the 800 mg dose. Among the first 24 patients in Phase 1b, DS occurred in seven patients, including three KMT2A-r patients at 200 mg (two of whom experienced grade 3 or higher events, including one death) and four patients at 600 mg (two of whom experienced grade 3 DS (one KMT2A-r and one NPM1-m) and two of whom experienced grade 2 DS (one KMT2A-r and one NPM1-m)). Development and implementation of DS guidance has reduced the severity of reported DS events.

[0152] A review of all data for potential DS events not identified by the investigator was performed using the Norsworthy algorithm (Norsworthy, 2020). These data, along with investigator-identified cases, are summarized in Table 7. As shown in Table 7, among the first 36 patients in Phase 1b, the rate of investigator-reported DS was consistent between the 200 mg and 600 mg dose levels (29.4% vs. 26.3%, respectively). The rate of severe DS in Phase 1b was higher at the 200 mg dose level, with 23.5% of patients reporting Grade 3 or higher DS compared with 10.5% at the 600 mg dose level, suggesting that DS severity did not increase with dose.

[0153] [Table 7]

[0154] While the overall safety profile for each genetic cohort was consistent across the entire study population and not dose-dependent, a genetic subtype dependency for DS was observed, with KMT2A-r patients being more likely to experience events, as shown in Table 8. Of the 10 total DS cases reported in 36 patients in Phase 1b, 7 (70.0%) occurred in KMT2A-r patients, suggesting that KMT2A-r patients are potentially more susceptible to DS-type events. In addition, KMT2A-r patients reported an increased rate of severe DS compared with the overall NPM1-m subgroup (20.8% in KMT2A-r vs. 5.9% in NPM1-m), as well as at the 200 mg and 600 mg diftomenib dose levels, respectively. When DS occurred in NPM1-m subjects, it was generally moderate in severity and resolved with appropriate intervention.

[0155] [Table 8]

[0156] Grade ≥ 3 TEAEs (regardless of cause assessment) occurring in > 10% of participants in Phase 1b by genetic subtype and dose are shown in Table 9 for the 53-subject group. Few adverse events, regardless of causality, were grade ≥ 3 and occurred in > 10% of patients. None were reported in the NPM1-m population. For KMT2A-r patients, 25% reported differentiation syndrome and 13% reported febrile neutropenia at the 600 mg dose.

[0157] [Table 9]

[0158] The incidence and severity of differentiation syndrome across 53 subjects in Phase 1a / 1b for the 200 mg and 600 mg doses by genotype is shown in Table 10.

[0159] [Table 10] As shown above, 20% of NPM1-m patients treated with 600 mg experienced differentiation syndrome, and one-quarter of those were at least grade 3. For KMT2A-r patients, rates were similar across doses, with approximately 38% of patients experiencing differentiation syndrome and 25-30% experiencing grade 3 or higher events. Patients with differentiation syndrome events at 600 mg had ORR rates of 75% for NPM1-m and 16.7% for KMT2A-r.

[0160] There were additional, less commonly reported adverse events (elevated alanine aminotransferase, pleuritic chest pain, and myocarditis) that may be related to DS events. These adverse events occurred almost exclusively in KMT2A-r patients. Because KMT2A-r disease is often monocytic and therefore invasive, it may be associated with extramedullary disease that is not always recognized at baseline (Fianchi et al., Mediterr. J. Hematol Infect Dis. 2021, 13(1), e2021030.). Because diftomenib concentrates in tissues, it is possible and suspected that these events are related to diftomenib-induced differentiation of leukemic cells in relevant extramedullary sites of disease and are not indicators of disease progression.

[0161] These data suggest that differentiation syndrome represents the most significant adverse effect associated with diftomenib therapy. While this does not show evidence of dose dependency, there is a clear association with KMT2A-r disease versus NPM1-m AML. In the genetically enriched phase 1b population, DS was observed in a genetic subtype-dependent manner, rather than a dose-dependent manner, with KMT2A-r patients being more likely to experience events. Due to the monocytic and therefore invasive nature of KMT2A-r leukemia, these patients are suspected to have a high degree of undiagnosed extramedullary disease and associated organ involvement (including the heart, liver, and other organs). Menin inhibitor-associated DS, such as that induced by diftomenib, has not been previously reported and may manifest as bone pain, transient pain in the extremities, initial elevations in liver function tests, cardiac inflammatory events, and other transient changes in organ function, all of which are potentially related to the differentiation of previously unrecognized extramedullary disease. This concept is supported by the well-known aggressiveness of monocytic disease and high levels of extramedullary involvement in KMT2A-r patients versus AML patients with other genotypes (Kapur et al., Leuk. Res. Rep. 2022, 18, 100349.).

[0162] At the time of filing, Phase 1b enrollment was complete (N=53), with 21% (all at the 600 mg dose) still on treatment with progressive clinical benefit. The overall safety profile in the expansion group was consistent with that described above. No new DS cases were reported or detected via the DS algorithm since the last data cutoff (N=3 datasets / 4 safety databases) among NPM1-m subjects.

[0163] Discoveries made during the study contributed to the development of treatment strategies to maximize patient time on treatment. Notably, leukocytosis was often misinterpreted as disease progression or DS, and changes in peripheral blast counts were similarly unpredictable. Control of DS episodes was associated with a higher likelihood of clinical benefit. While rare, when this event occurred in NPM1-m patients, it was manageable with modified DS monitoring and treatment implementation, resolved rapidly, and often predicted leukemia clearance, in part by allowing patients to continue treatment for at least two cycles (approximately 75% of NPM1-m patients with DS events achieved CR / CRh / CRi (ORR)). The emergence of specific DS monitoring and treatment strategies helped identify the unique menin inhibitor-associated presentation of DS and address it in a way that allowed patients to continue treatment.

[0164] D. Blood chemistry The mean changes from baseline in cycle 1 (28 days) for peripheral blasts and white blood cells for the on-target populations (Phase 1a and Phase 1b) at 600 mg showed a transient increase in these blood chemistry markers over the first 1 to 2 weeks, followed by a decrease in these markers for the remainder of the first cycle, as shown in Figure 2. While not always associated with DS events or predictive of progression, the early rise in white blood cell (WBC) count and peripheral blasts appears to correlate with drug activity and provides context for the rapid onset of DS-type symptoms after treatment initiation. The recognition that DS symptoms were indicative of drug efficacy led to the implementation of mitigation procedures that allowed patients to continue treatment. Managed DS can predict patient response, with 75% and 17% of NPM1-m and KMT2A-r patients with DS, respectively, ultimately achieving ORR events, although differences in the severity of DS between genetic subtypes were observed. The different responses may be due to the disseminated nature of KMT2A-r disease, with extensive infiltration into the extramedullary space. As shown in Example 1, diftomenib accumulates in tissues at high levels. This high tissue penetration may lead to the diffuse and unpredictable DS symptoms (e.g., diffuse pain) reported in KMT2A-r patients. The implementation of DS guidance has led to better identification of DS events and earlier intervention.

[0165] E. Exposures and Biomarkers Intensive or sparse pharmacokinetic sampling was performed after the first dose (Cycle 1, Day 1) and at steady state (Cycle 2, Day 1). Steady-state trough samples (pre-dose) were collected on Days 8 and 15 of Cycle 1 and, when possible, in subsequent cycles. Concentrations of diftomenib and specific active metabolites were measured by validated liquid chromatography / mass spectrometry assays. The following parameters were measured: AUC 0-24-ss (area under the plasma concentration-time curve over the dosing interval at steady state), C trough-ss (trough plasma concentration measured at the end of the dosing interval at steady state), and C max-ss(maximum concentration at steady state) was determined. Exposure-response analyses were performed for subjects who received at least 21 doses in the first cycle, using ORR (defined as MLFS, Cri [including CRp], CRh, or CR+ / -MRD-) as the efficacy endpoint to determine the exposure / efficacy relationship. As shown in Figure 3, panels A-C, a dose-dependent increase in exposure was observed with the 600 mg dose compared with the 200 mg dose. However, diftomenib exposure at the 600 mg and 800 mg doses was comparable. Based on these studies, any increase in dose beyond 600 mg QD is not expected to increase the probability or extent of response.

[0166] Changes in MEIS1 expression on day 28 of cycle 1 were measured as a pharmacodynamic biomarker of diftomenib inhibition of menin transcriptional signaling. Evaluation of MEIS1 expression levels was performed in bone marrow aspirate samples collected during screening and at the end of cycle 1 treatment with diftomenib. Changes in MEIS1 expression were determined using quantitative ribonucleic acid sequencing. Patients who achieved a CR by the time of bone marrow biopsy were not evaluated because MEIS1 expression is expected to be low due to the absence of leukemic cells. As shown in Figure 4A, MEIS1 expression decreased by more than 85% at the end of cycle 1 in four patients treated with 600 mg of diftomenib, but no decrease was detected in two patients treated with 200 mg. As shown in Figure 4B, MEIS1 expression was 6-fold and 8-fold lower on day 28 of cycle 1 in KMT2A-r and NPM1-m patients treated with 600 mg compared to 200 mg, respectively. Target gene expression at the 800 mg dose provided no evidence of further knockdown.

[0167] Furthermore, expression of other menin target genes, such as HOXA9, HOXA10, and MEF2C, was 2- to 6-fold lower at 600 mg than at 200 mg. RNAseq data showing stronger inhibition of the menin pathway at 600 mg provides supportive evidence for the 600 mg dose over the 200 mg dose.

[0168] Population pharmacokinetic analysis determined that concomitant administration of CYP3A4 inhibitors, renal or hepatic status, patient mutations, and ECOG status were not significant covariates affecting pharmacokinetics.

[0169] F. Supplemental Clinical Study Data Supplementary note 1.

[0170] This report provides an update on duration of remission (DoR) for Ph1 NPM1-m patients treated with 600 mg RP2D (n=20) and those treated with 200 mg as of January 31, 2023. The median age of patients treated with 600 mg RP2D was 70.5 years (range, 22-86 years). FLT3 and IDH1 / 2 mutations were common (35% for FLT3 and 30% for IDH1 / 2, respectively), with 20% having simultaneous FLT3 and IDH1 / 2 co-mutations. The median number of prior lines of therapy was 2.5 (range, 1-8), with 15% having one or more prior stem cell transplants (SCTs) and 60% having prior venetoclax.

[0171] The cumulative safety profile of diftomenib 600 mg RP2D was consistent with previous reports, with no new signals observed. Most subjects (85%) experienced at least one grade 3 or higher treatment-emergent adverse event (TEAE), with 30% of TEAEs considered potentially treatment-related. The most frequent (>10%) grade 3 or higher TEAEs were anemia (25%), pneumonia (20%), thrombocytopenia, neutropenia, and hyperglycemia (15% each). Among NPM1-m patients, 20% reported differentiation syndrome (DS) events of any grade, and 5% (n=1) experienced grade 3 DS.

[0172] As of January 31, 2023, the complete remission (CR) rate for patients treated with 600 mg of NPM1-m was 30% (6 patients, 95% CI 12-54%); the CR and CR / CRh rates were both 30% (6 patients, 95% CI 12-54%); the combined CR rate (CRc, CR+CRh+CRi) was 35% (7 patients, 95% CI 15-59%); the MRD-negativity rate was 43% (3 patients, 95% CI 10-82%); five of the seven patients who achieved CRc were evaluated for MRD, and 60% of those evaluated were MRD-negative; and the ORR rate was 40% (8 patients, 95% CI 19-64%). The median DoR for patients achieving a maturing CRc was 8.2 months per Kaplan-Meier (KM) estimate (95% CI 1.5-NE). One CR was noted at the 200 mg dose, with an ongoing DoR of 32 cycles. The median time to CR for NPM1-m patients at RP2D was 70 days (r: 26-89). Two patients (one CR and one CR with incomplete hematologic recovery [CRi]) proceeded to SCT, and both remained in remission at the time of the cutoff. The median overall survival for NPM1-m patients treated with 600 mg was 5.1 months (95% CI: 2.1-NE), with a median follow-up of 8.0 months. With respect to the cutoff, 57.1% of patients who achieved CRc at RP2D were continuing treatment or were undergoing follow-up after SCT, and patients continued to show evidence of progressive response during treatment.

[0173] Diftomenib 600 mg continued to demonstrate significant clinical activity in heavily pretreated and co-mutated R / R NPM1-m AML patients. The safety profile remained consistent with previous reports, and the on-target effects of diftomenib remained manageable. Data suggest sustained remission as DoR continues to mature in five of eight patients with ongoing CRc at the cutoff (four of seven achieved CR at 600 mg and one at 200 mg). A single-arm, enrollment-oriented phase 2 trial is currently underway to further evaluate diftomenib monotherapy in R / R NPM1-m AML.

[0174] Supplement 2. This report provides an update of NPM1-m patients from Phase 1a and Phase 1b, with emphasis on patients receiving the 600 mg RP2D (N=20) as of April 12, 2023. The median age of patients receiving the 600 mg RP2D was 70.5 years (range, 22-86 years). Co-mutations in FLT3 (30%) and IDH1 / 2 (40%) were common (20% had both co-mutations). The median number of prior therapies was 3.0 (range, 1-10), and 20% had received one or more prior stem cell transplants (SCTs). Phase 1B patient baseline characteristics are shown in Table 11.

[0175] [Table 11]

[0176] Based on the latest results, the complete remission (CR) rate for patients receiving 600 mg of NPM1-m was 35%, with 40% achieving a combined CR (CRc) and an overall ORR of 45%. The median time to first response was 51 days (r: 26-225). One CR at the 200 mg dose had an ongoing DoR of 35 cycles. The median DoR for all NPM1-m patients who achieved CRc was 8.2 months per Kaplan-Meier estimate (95% CI: 1.0-NE). Two patients (one CR and one CRi) underwent SCT and remained in remission at the cutoff, and one received diftomenib maintenance therapy after SCT. The median remission duration was 8.2 months.

[0177] Because at least two patients with FLT3 and IDH1 co-mutations present at baseline were undetectable after two cycles, molecular analysis suggests that diftomenib results in measurable residual disease (MRD) clearance of targeted mutations and co-mutations (e.g., FLT3 and IDH1) such as NPM1, possibly by targeting founder clonal and subclonal events or by targeting aberrant gene expression. The data are shown in Table 12.

[0178] [Table 12]

[0179] Most patients (85%) had at least one grade 3 or higher treatment-emergent adverse event (TEAE), with 30% potentially treatment-related. The most common (>20%) grade 3 or higher TEAEs were anemia (25%) and thrombocytopenia (20%). Differentiation syndrome (DS) of any grade was reported in 20%, with most (n=3) being grade 2.

[0180] Resistance profile was investigated and the resistance mutation MEN1-M327I was found to develop in 1 of 29 patients (3.4%) and was detected at C4D28, with the patient maintaining stable disease through cycle 7.

[0181] In summary, diftomenib continued to demonstrate significant clinical activity in heavily pretreated and co-mutated R / RNPM1-m AML patients, with 35% achieving a complete response. The safety profile was consistent with previous reports, and episodes of DS were clinically manageable. Data revealed durable remissions with MRD clearance of NPM1 and significant co-mutations. Resistance mutations were rarely detected, indicating that diftomenib remains effective against common menin gatekeeper mutations.

[0182] Supplement 3. This report provides an update on the phase 1 study of diftomenib in patients with AML as of August 30, 2023 (cutoff date). Patient demographics remained the same as those reported in Table 11. No patients continued treatment as of the cutoff date, and four patients continued on the study. Treatment was discontinued due to adverse events (5, 25%), death (1, 5%), physician's decision (1, 5%), investigator-assessed disease progression (9, 45%), acceptance of alternative anticancer therapy (1, 5%), and other (2, 10%). Patient participation in the study was discontinued due to completion (1, 5%) or death (15, 75%).

[0183] In 20 patients with relapsed / refractory NPM1-m AML treated with diftomenib 600 mg daily across phase 1a and 1b (a median of three prior lines of therapy), the complete remission (CR) rate at the cutoff date was 35% (95% confidence interval [CI], 15.4-59.2), the combined remission (CR / CRh) rate was 40%, and the overall response rate (ORR) was 45% (95% CI, 23.1-68.5). CR / CRh responses were durable (5.6 months, 7.7 months for CRc responders) and associated with myeloblast reduction, count recovery, transfusion independence, and negative tests for measurable residual disease (MRD). The median overall survival (OS) for the 20 NPM1-m patients treated with diftomenib 600 mg was 5.6 months, and 12.1 months for CR / CRh responders.

[0184] The known menin resistance mutation MEN1-M327I was detected in 1 of 29 (3.4%) treated patients tested by RNA next-generation sequencing of serial bone marrow aspirates. Even among patients who received at least two cycles of diftomenib and continued to have measurable leukemia, no additional resistance mutations were detected, suggesting that disease progression in these patients was not due to MEN1 mutations. In contrast, 39% (12 of 31) of patients treated with revumenib for at least two cycles were found to have one or more MEN1 mutations, often concomitant with clinical progression (Perner et al., MEN1 mutations mediate clinical resistance to menin inhibition, Nature 2023, 615, 913-919).

[0185] Across all NPM1-m patients in phase 1a and 1b, the median time to response was 51 days (range, 26-225). For all patients with NPM1-m across phase 1a and 1b who achieved a combined complete response (CR), the median remission duration was 7.7 months, with a median follow-up period of 13.4 months. One patient with a CR on the 200 mg dose had an ongoing remission lasting 35 cycles. Two patients (one in CR and one in CRh) proceeded to stem cell transplant and remained in remission as of the cutoff date, and one patient remained on diftomenib maintenance therapy after transplantation. The 200 mg dose had a lower CR / CRh recovery rate (6.7%) and a lower overall response rate (13.3%) compared with the 600 mg dose.

[0186] QTc prolongation due to diftomenib was not detected in this patient population. Even in patients treated with the menin inhibitor revumenib, a 53% QT prolongation rate was observed as a treatment-related adverse event, and no significant drug-drug interactions were observed (Issa et al., The menin inhibitor revumenib in KMT2A-rearranged or NPM1-mutant leukemia, Nature 2023, 615, 920-924). Cases of grade 3 or higher differentiation syndrome were rare and effectively managed. In patients treated with 200 mg or 600 mg diftomenib, the rate of differentiation syndrome was lower in NPM1-m patients (15%) than in KMT2A-r patients (41%). Clinically significant cytopenias were not observed in patients achieving CR or CRh in the study.

[0187] Tabulated data are shown in Tables 13 and 14.

[0188] [Table 13]

[0189] [Table 14]

[0190] G. Summary In summary, diftomenib was safe and tolerable. Reported adverse events were mostly consistent with the characteristics and manifestations of the underlying disease. No evidence of drug-induced QTc prolongation was observed. The on-target effect of differentiation syndrome was managed with monitoring and / or interventional mitigation. Clinical activity of diftomenib monotherapy was found to be optimal at the 600 mg dose. A favorable NPM1-m benefit / risk balance was achieved with significant activity and a 30% CR rate (n=20), which increased to 35% as the study progressed, a 35% CR / CRh rate (n=40), and an ORR of 40%, which increased to 45% as the study progressed. High levels of diftomenib tissue penetration can facilitate clearance of extramedullary disease in patients with AML. Increased exposure above the 600 mg dose did not significantly correlate with an increased probability of clinical response for any genetic subgroup and for NPM1-m subjects; specifically, increased exposure did not correlate with a higher risk of grade 3 or higher AEs (i.e., safety risks do not increase with increased exposure). While an initial cut of data focused on the 200 mg dose, the expanded data set combined with pharmacokinetic analysis demonstrates the superiority of the 600 mg dose over the 200 mg dose in providing peak exposure without a significantly increased risk of grade 3 or higher adverse events. Notably, in the NPM1-m patient population, the 600 mg QD dose is comparable to the 200 mg dose.

[0191] The 600 mg once-daily dose was identified as the optimal biologic dose and approved by the FDA as the recommended phase 2 dose of diftomenib for further testing in the NPM1-m AML population.

[0192] Although both NPM1-m and KMT2A-r AML are menin-dependent, it can be assumed that the same dosing regimen is optimal for both groups at a safe and effective dose. Indeed, although efficacy data generated in the combined population were promising, the safety profile differentiated response rates and benefit-risk profiles for the two genotypes, likely due to more extensive extramedullary disease in the KMT2A-r subtype.

[0193] In Phase 1a, there was significant overlap in exposure between the 200 mg and 400 mg dose levels and between the 400 mg and 600 mg dose levels, suggesting that clinical efficacy was similar across all dose levels. Nevertheless, clinically meaningful differences in response rates were observed in NPM1-m patients in the combined Phase 1a / 1b study, with a 16.7% CR rate at 200 mg and a rapid and durable 35% CR rate at 600 mg. Treatment benefits associated with CR include rapid recovery of hematological parameters, which contribute to transfusion independence.

[0194] The safety profile of diftomenib for both genotypes combined is influenced by the experience of KMT2A-r patients and distinguishes it from that observed in NPM1-m patients. Regarding differentiation syndrome specifically, no fatal or life-threatening cases of DS have been reported in NPM1-m patients, and all cases in this population have been tolerable, reversible, and usually low-grade. Additionally, DS cases in NPM1-m patients were associated with clinical responses, occurring in three of four NPM1-m patients who experienced DS. The severity of DS in KMT2A-r patients may be related to the extensive extramedullary disease specific to this genotype.

[0195] H. Resistance Mutation Test Various somatic resistance mutations in MEN1, including MEN1-M327I, MEN1-T349M, MEN1-G331R, and MEN1-G331D, have been reported after menin inhibitor therapy. For example, the T349 mutation was detected in the majority of patients who acquired menin gatekeeper mutations in another recent menin inhibitor clinical trial (Perner, F., Stein, EM, Wenge, DV et al. MEN1 mutations mediate clinical resistance to menin inhibition. Nature 615, 913-919 (2023)). In the diftomenib phase 1 trial described herein, mutation analysis (RNA sequencing) of clinical samples after treatment with diftomenib revealed that one resistance mutation, MEN1-M327I, developed in 1 of 29 patients (3.4%) tested. In one patient, the mutation was detected at C4D28, but the patient maintained stable disease through cycle 7. It has been reported that binding of diftomenib to M327I-mutated MEN1 in a menin-MLL binding assay was reduced compared to binding to the wild-type protein, but diftomenib retained an IC50 binding activity of less than 100 nM against the T349M variant. The data are shown in Table 15 (Grembecka, Development of new targeted therapeutics for AML, Presentation at 3rd Biennial Miami Leukemia Symposium, March 31-April 2, 2023).

[0196] [Table 15]

[0197] In summary, from the data available to date, resistance mutations appear to emerge infrequently after treatment with diftomenib, and diftomenib retains binding activity against common menin gatekeeper mutations observed in response to exposure to menin inhibitors.

[0198] Thus, provided herein is a method of inhibiting MEN1-MLL interaction, comprising contacting MEN1 with diftomenib, wherein MEN1 comprises a resistance mutation, optionally selected from mutations at M327, T349, S160, and G331, and combinations thereof; optionally, the mutations are selected from M327I, M327V, T349M, S160T, G331R, and G331D, and combinations thereof. Furthermore, in some embodiments of the methods provided herein, the acute leukemia, leukemia cells, or AML comprises MEN1 comprising a resistance mutation, optionally selected from mutations at M327, T349, S160, and G331, and combinations thereof; optionally, the mutation is selected from M327I, M327V, T349M, S160T, G331R, and G331D, and combinations thereof. In some embodiments, the mutation is T349M or G331D. In some embodiments, the MEN1 resistance mutation is a de novo mutation. In some embodiments, the MEN1 resistance mutation is an acquired mutation that develops, for example, after exposure to a menin inhibitor.

[0199] While several embodiments of the present invention have been shown and described herein, it will be apparent to those skilled in the art that such embodiments are provided by way of example only. It is not intended that the present invention be limited by the specific examples provided within the specification. While the present invention has been described with reference to the foregoing specification, the description and illustration of the embodiments herein are not meant to be construed in a limiting sense. Numerous variations, changes, and substitutions will occur to those skilled in the art without departing from the invention. Furthermore, it should be understood that all aspects of the present invention are not limited to the specific depictions, configurations, or relative proportions set forth herein, which depend upon a variety of conditions and variables. It should be understood that various alternatives to the embodiments of the present invention described herein may be utilized in practicing the invention. It is therefore contemplated that the present invention also encompasses such alternatives, modifications, variations, or equivalents. The following claims define the scope of the invention, and it is intended that methods and structures within the scope of the claims and their equivalents be covered thereby.

Claims

1. 1. A method of treating acute leukemia in an individual, or inhibiting proliferation and / or inducing apoptosis in leukemia cells in an individual, comprising administering to the individual 600 milligrams of diftomenib or a pharmaceutically acceptable form thereof daily.

2. 1. A method for treating extramedullary leukemia in an individual with acute leukemia, comprising administering to said individual an effective amount of a menin inhibitor, particularly 600 milligrams of diftomenib or a pharmaceutically acceptable form thereof, daily.

3. 3. The method of claim 1 or 2, wherein the acute leukemia or leukemia cells are menin dependent.

4. 4. The method of any one of claims 1 to 3, wherein the acute leukemia or leukemia cells comprise a mutation selected from an NPM1 mutation, a KMT2A rearrangement, a KMT2A-PTD mutation, a SETD2 mutation, a RUNX1 mutation, a FLT3-ITD mutation, a FLT3-TKD mutation, an IDH mutation (e.g., an IDH1 mutation or an IDH2 mutation), a TERT mutation, or a BRAF mutation, or any combination thereof, and optionally the acute leukemia or leukemia cells comprise an NPM1 mutation, optionally in combination with a FLT3 mutation (e.g., a FLT3-internal tandem duplication (ITD) mutation or a FLT3 mutation in the tyrosine kinase domain (FLT3-TKD mutation)) or an IDH mutation (e.g., an IDH1 mutation or an IDH2 mutation), or a combination thereof.

5. 5. The method of any one of claims 1 to 4, wherein the acute leukemia or leukemia cells comprise a nucleophosmin 1 (NPM1) mutation, a lysine methyltransferase 2a (KMT2A) rearrangement, a SET domain-containing 2 (SETD2) mutation, or a runt-related transcription factor 1 (RUNX1) mutation.

6. 6. The method of any one of claims 1-5, wherein the acute leukemia is acute myeloid leukemia (AML), optionally wherein the AML is refractory AML, relapsed AML, or both relapsed and refractory AML, and optionally wherein the AML is acute promyelocytic leukemia, acute myeloblastic leukemia, or acute megakaryoblastic leukemia.

7. The method of any one of claims 1 to 6, wherein the leukemic cells, acute leukemia, or AML comprise an NPM1 mutation.

8. The method of any one of claims 1 to 6, wherein the leukemic cells, acute leukemia, or AML comprise a KMT2A rearrangement.

9. 7. The method of any one of claims 1 to 6, wherein the acute leukemia is acute lymphocytic leukemia (ALL), optionally wherein the ALL is refractory ALL, relapsed ALL, or both relapsed and refractory ALL, and optionally wherein the ALL is precursor B acute lymphoblastic leukemia, precursor T acute lymphoblastic leukemia, Burkitt's leukemia, or acute mixed lineage leukemia.

10. 10. The method of claim 9, wherein the ALL comprises an NPM1 mutation.

11. 10. The method of claim 9, wherein the ALL comprises a KMT2A rearrangement.

12. 12. The method of any one of claims 1-11, wherein the administration of diftomenib or a pharmaceutically acceptable form thereof comprises administering diftomenib or a pharmaceutically acceptable form thereof to the individual daily for at least 3 days, or at least 5 days, or at least 7 days, or at least 10 days, or at least 14 days, or at least 21 days, or at least 28 days, or over a cycle comprising at least 28 days, or over a cycle comprising 28 days.

13. 13. The method of claim 12, wherein the administering comprises administering diftomenib or a pharmaceutically acceptable form thereof to the individual daily for N cycles, wherein a cycle comprises 28 days and N is at least 1.

14. 14. The method of claim 13, wherein N is 2, 3, 4, 5, or 6, or is at least 2, at least 3, at least 4, at least 5, or at least 6.

15. 15. The method of claim 13 or 14, wherein the N cycles are consecutive (i.e., 0 days between cycles).

16. 16. The method of any one of claims 1-15, wherein the risk of the individual developing any Grade 3 or higher treatment-emergent adverse event (TEAE) regardless of causality following administration is less than about 80%, or less than about 75%, or about 71%.

17. 16. The method of any one of claims 1-15, wherein the risk of the individual developing any serious adverse event, regardless of causality following administration, is less than about 65%, or less than about 60%, or less than about 55%, or is about 53%.

18. 16. The method of any one of claims 1-15, wherein the risk to the individual of developing any adverse event suspected of being a differentiation syndrome following administration is less than about 80%, or less than about 75%, or less than about 70%, or less than about 65%, or less than about 60%, or less than about 55%, or less than about 50%, or is about 47%.

19. 18. The method of any one of claims 1-17, wherein the risk of the individual developing differentiation syndrome following administration is less than about 25%, or less than about 20%, or less than about 19%, or less than about 18%, or is about 18%.

20. 20. The method of claim 19, wherein the probability of differentiation syndrome, including severe differentiation syndrome, is less than about 50%, or less than about 45%, or less than about 40%, or less than about 35%, or about 33%.

21. 18. The method of any one of claims 1-17, wherein the risk of the individual developing severe differentiation syndrome following administration is less than about 20%, or less than about 15%, or less than about 10%, or less than about 7%, or about 6%.

22. 18. The method of any one of claims 1-17, wherein the probability that the individual develops a differentiation syndrome following administration and that the differentiation syndrome developed by said individual is not a severe differentiation syndrome (e.g., is grade 1 or 2) is greater than about 50%, or greater than about 55%, or greater than about 60%, or greater than about 65%, or about 67%.

23. 23. The method of any one of claims 1-22, wherein the administration results in a CR or CR / CRh rate that is at least about 20%, or at least about 21%, or at least about 22%, or at least about 23%, or at least about 24%, or at least about 25%, or at least about 26%, or at least about 27%, or at least about 28%, or at least about 29%, or at least about 30%, or at least about 31%, or at least about 32%, or at least about 33%, or at least about 34%, or at least about 35%, or about 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, or 35%, or about 35%.

24. Administration may result in at least about 20%, or at least about 21%, or at least about 22%, or at least about 23%, or at least about 24%, or at least about 25%, or at least about 26%, or at least about 27%, or at least about 28%, or at least about 29%, or at least about 30%, or at least about 31%, or at least about 32%, or at least about 33%, or at least about 34% 24. The method of any one of claims 1 to 23, wherein the method results in a CRc rate that is at least about 35%, or at least about 36%, or at least about 37%, or at least about 38%, or at least about 39%, or at least about 40%, or is about 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, or 40%.

25. Administration may result in at least about 20%, or at least about 21%, or at least about 22%, or at least about 23%, or at least about 24%, or at least about 25%, or at least about 26%, or at least about 27%, or at least about 28%, or at least about 29%, or at least about 30%, or at least about 31%, or at least about 32%, or at least about 33%, or at least about 34%, or at least about 35%, or at least about 36%, or at least about 37%, or 25. The method of any one of claims 1-24, wherein the method results in an ORR that is at least about 38%, or at least about 39%, or at least about 40%, or at least about 41%, or at least about 42%, or at least about 43%, or at least about 44%, or at least about 45%, or about 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, 40%, 41%, 42%, 43%, 44%, or 45%.

26. 26. The method of any one of claims 7 or 12-25, wherein the administration results in a duration of remission (DoR) in individuals who achieve a CRc that is at least 2 months, or at least 3 months, or at least 4 months, or at least 5 months, or at least 6 months, or at least 7 months, or at least 8 months, or at least 9 months, or at least 10 months, or at least 11 months, or at least 12 months, or at least about 18 months, or at least about 24 months, or at least about 8.2 months, or about 8.2 months.

27. 27. The method of any one of claims 1-26, wherein the individual develops differentiation syndrome, wherein the differentiation syndrome is a severe differentiation syndrome (e.g., grade 3, 4, or 5), and wherein the individual is likely to achieve an ORR of at least about 60%, or at least about 65%, or at least about 70%, or at least about 75%, or about 65%, 66%, 67%, 68%, 69%, 70%, 71%, 72%, 73%, 74%, or 75%.

28. 28. The method of any one of claims 1 to 27, wherein the acute leukemia or leukemia cells comprise an NPM1 mutation, including an A, B, C, or D mutation.

29. 28. The method of any one of claims 1 to 27, wherein the acute leukemia or leukemia cell comprises an NPM1 mutation comprising an insertion (eg, a four nucleotide insertion) in exon 12 of the NPM1 gene.

30. 1. A method for increasing the level of myeloid blasts in the blood of an individual with AML, comprising administering to said individual an effective amount of a menin inhibitor, particularly 600 mg of diftomenib or a pharmaceutically acceptable form thereof, daily.

31. 31. The method of claim 30, wherein the AML is menin-dependent.

32. 32. The method of claim 30 or 31, wherein the AML comprises a mutation selected from an NPM1 mutation, a KMT2A rearrangement, a KMT2A-PTD mutation, a SETD2 mutation, a RUNX1 mutation, a FLT3-ITD mutation, a FLT3-TKD mutation, an IDH mutation (e.g., an IDH1 mutation or an IDH2 mutation), a TERT mutation, or a BRAF mutation, or any combination thereof, and optionally the AML comprises an NPM1 mutation, optionally in combination with a FLT3 mutation (e.g., a FLT3-internal tandem duplication (ITD) mutation or a FLT3 mutation in the tyrosine kinase domain (FLT3-TKD mutation)) or an IDH mutation (e.g., an IDH1 mutation or an IDH2 mutation), or a combination thereof.

33. The method of any one of claims 30 to 32, wherein the AML comprises an NPM1 mutation, a KMT2A rearrangement, a SETD2 mutation, or a RUNX1 mutation.

34. 34. The method of claim 33, wherein the AML comprises an NPM1 mutation or a KMT2A rearrangement, preferably the AML comprises an NPM1 mutation.

35. 35. The method of claim 33 or 34, wherein the relative level of myeloid blasts in the individual's blood is assessed by analysis of two separated blood samples from the individual, optionally the blood samples being taken, for example, (a) at a first time point and a second time point (both time points during administration), or (b) at a first time point before the start of the administration and a second time point during the administration (e.g., the first time point is before cycle 1, day 1 and the second time point is at least 7, 14, 21, 28 days later, e.g., from cycle 1, day 2 to cycle 1, day 28).

36. 36. The method of claim 35, wherein each analysis is a complete blood count (CBC) optionally with differential.

37. 37. The method of claim 35 or 36, wherein the first and second time points are separated by 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, 1 week, 2 weeks, 3 weeks, 4 weeks, at least 1 week, at least 2 weeks, at least 3 weeks, or at least 4 weeks, or the duration of one cycle (such as cycle 1), or the second time point is the time when the individual achieves complete remission (CR).

38. 38. The method of any one of claims 33 to 37, wherein an increased level of myeloid blasts in the blood of the individual does not correlate with progression of AML.

39. 39. The method of any one of claims 33 to 38, wherein the increased level of myeloid blasts occurs in the extramedullary space.

40. 40. The method of claim 39, wherein an increased level of myeloid blasts indicates that the individual is sensitive to a menin inhibitor, particularly diftomenib or a pharmaceutical form thereof.

41. 1. A method of identifying acute leukemia (particularly AML) in an individual as being susceptible to administration of a menin inhibitor, particularly diftomenib, comprising: administering to said individual daily an effective amount of said menin inhibitor, particularly 600 mg of diftomenib or a pharmaceutically acceptable form thereof; receiving an identification of the level of myeloid blasts in a first blood sample taken from the individual at a first time point, either before the start of administration or during said administration; receiving an identification of the level of myeloid blasts in a second blood sample taken from the individual at a second time point after the first time point and during the administration; determining that the acute leukemia is susceptible to said administration if the level of myeloid blasts at said second time point is greater than the level at said first time point.

42. 1. A method of treating a differentiation disorder, particularly tumor lysis syndrome, in an individual diagnosed with a differentiation disorder in an individual with acute leukemia, or a method of reducing the risk of developing a severe differentiation disorder in an individual with acute leukemia, optionally comprising: (a) administering to said individual daily an effective amount of a menin inhibitor, particularly 600 mg of diftomenib or a pharmaceutically acceptable form thereof; (b) administering IV hydration to said individual; and optionally, (c) administering to said individual an effective amount of a xanthine oxidase inhibitor, optionally wherein said xanthine oxidase inhibitor is allopurinol; and optionally administering allopurinol at a dose of about 200-400 mg / m ２ and administering daily at a dose of 1 to 3 divided doses up to a maximum of about 800 mg / day.

43. administering the xanthine oxidase inhibitor, wherein prior to administering the xanthine oxidase inhibitor, the individual: (1) Approximately 25 x 10 ９ a white blood cell count below L and a lactate dehydrogenase level less than two times the upper limit of normal, or (2) Approximately 25 to approximately 100 x 10 ９ / L white blood cell count, or (3) Approximately 25 x 10 ９ 43. The method of claim 42, comprising diagnosing the individual as having a low or moderate risk of having or developing the differentiation disorder based on receiving identification of the individual as having a white blood cell count of less than 1 / L and a lactate dehydrogenase level greater than two times the upper limit of normal.

44. 1. A method of treating differentiation disorders in an individual with acute leukemia or reducing the risk of developing a severe differentiation disorder in an individual with acute leukemia, comprising: (a) administering to said individual daily an effective amount of a menin inhibitor, particularly 600 mg of diftomenib or a pharmaceutically acceptable form thereof; (b) administering IV hydration to the individual; (c) administering a therapeutically effective amount of rasburicase daily, optionally at a dose of about 0.2 mg / kg, optionally as an intravenous infusion over about 30 minutes, for up to about 5 days.

45. Prior to administering rasburicase, the individual (i) Approximately 100 × 10 ９ have a white blood cell count level of 1 / L or greater, or (ii) (i) about 25 to about 100 × 10 ９ / L, or (ii) a white blood cell count level of about 25 x 10 ９ have a white blood cell count level of less than 1 / L and a lactate dehydrogenase level of more than two times the upper limit of normal, For each of (i) and (ii), the individual has impaired renal function or uric acid, potassium, and / or phosphate levels above the applicable upper normal limit; 45. The method of claim 44, comprising receiving an identification that the individual has or is at high risk of developing the differentiation disorder.

46. 1. A method of treating differentiation disorders or reducing the risk of developing severe differentiation disorders in an individual with acute leukemia, comprising: (a) administering to said individual a prophylactic and / or effective amount of a corticosteroid, optionally wherein the corticosteroid is prednisone at a dose of about 0.5 mg / kg (or an equivalent dose of an alternative corticosteroid); (b) administering to said individual a therapeutically effective amount of a menin inhibitor.

47. 47. The method of claim 46, wherein the corticosteroid and the menin inhibitor are administered daily, and sequentially or simultaneously, optionally comprising administering a first dose of each of the corticosteroid and the menin inhibitor on the same or about the same day, or administering a first dose of the corticosteroid on a day before or after the first administration of the menin inhibitor.

48. Prior to administering the corticosteroid or the menin inhibitor, the individual has about 5×10 ９ 48. The method of claim 46 or 47, wherein the patient has one or more of: a white blood cell count greater than 1 / L, an elevated serum creatinine level, significant extramedullary disease, and proliferative acute leukemia.

49. 49. The method of any one of claims 46-48, comprising administering the corticosteroid daily starting on day 1, administering the menin inhibitor daily starting on the same or a subsequent day, and decreasing the dose of the corticosteroid after about 28 days of administration of the menin inhibitor if the individual has not been diagnosed with the differentiation disorder (in methods of reducing risk) or if the individual develops the differentiation disorder and the differentiation disorder is ameliorated and bone marrow blasts are at a level of less than about 5% (in methods of reducing risk or methods of treating).

50. 49. The method of any one of claims 46-48, comprising administering intravenously to the individual a dose of about 5 mg, 10 mg, or 15 mg, or about 5-10 mg, preferably about 10 mg, of dexamethasone every 12 hours (or an equivalent dose of an alternative oral or IV corticosteroid) for 1, 2, or 3 days.

51. The individual's white blood cell count or leukocyte count is about 10 x 10 ９ 51. The method of any one of claims 46-50, comprising administering to the individual a therapeutically effective amount of hydroxyurea if the IL-16 expression level increases to greater than 1 / L or doubles within about 24-48 hours, and optionally administering to the individual a therapeutically effective amount of cytarabine, idarubicin, or gemtuzumab.

52. 52. The method of claim 51, comprising tapering the dose of and / or discontinuing administration of the corticosteroid, hydroxyurea, cytarabine, idarubicin, or gemtuzumab upon improvement of the differentiation disorder.

53. interrupting administration of the menin inhibitor during all or part of the administration of one or more of IV hydration, allopurinol, rasburicase, prednisone, dexamethasone, hydroxyurea, cytarabine, idarubicin, and gemtuzumab; and administering the menin inhibitor to a patient after the differentiation disorder has improved (e.g., when the white blood cell count reaches about 20×10 ９ and resuming administration when the serum saturation level drops below 1 / L.

54. The method of any one of claims 42 to 53, wherein the differentiation disorder is differentiation syndrome with or without leukocytosis or tumor lysis syndrome.

55. The method of any one of claims 41 to 54, wherein the acute leukemia comprises AML.

56. 55. The method of any one of claims 41 to 54, wherein the acute leukemia comprises ALL.

57. 57. The method of claim 55 or 56, wherein the acute leukemia comprises an NPM1 mutation.

58. 58. The method of any one of claims 1 to 57, wherein the individual is aged 18 years or older.

59. 59. The method of any one of claims 41 to 58, wherein the menin inhibitor is diftomenib or a pharmaceutically acceptable form thereof.

60. 60. The method of any one of claims 1 to 59, wherein daily administration is once-daily administration.

61. The method of any one of claims 1 to 60, wherein diftomenib or a pharmaceutically acceptable salt thereof is diftomenib or a pharmaceutically acceptable salt thereof, or a solvate thereof, optionally diftomenib free base or a solvate thereof.

62. 62. The method of any one of claims 1-61, wherein 600 mg of diftomenib or a pharmaceutically acceptable form thereof is the optimal biological dose, the recommended Phase 2 dose, the safe and effective dose, or the submaximal tolerated dose of diftomenib or a pharmaceutically acceptable form thereof.

63. 63. The method of any one of claims 41 to 62, wherein the acute leukemia is menin dependent.

64. 64. The method of any one of claims 41-63, wherein the acute leukemia comprises a mutation selected from an NPM1 mutation, a KMT2A rearrangement, a KMT2A-PTD mutation, a SETD2 mutation, a RUNX1 mutation, a FLT3-ITD mutation, a FLT3-TKD mutation, an IDH mutation (e.g., an IDH1 mutation or an IDH2 mutation), a TERT mutation, or a BRAF mutation, or any combination thereof.

65. The method of any one of claims 41 to 64, wherein the acute leukemia comprises an NPM1 mutation, a KMT2A rearrangement, a SETD2 mutation, or a RUNX1 mutation, preferably an NPM1 mutation or a KMT2A rearrangement.

66. A pharmaceutical composition comprising an optimal biological dose, a recommended Phase 2 dose, a safe and effective dose, or a submaximal tolerated dose of diftomenib or a pharmaceutically acceptable form thereof, and a pharmaceutically acceptable carrier.

67. 67. The pharmaceutical composition of claim 66, wherein the optimal biological dose, the recommended Phase 2 dose, the safe and effective dose, or the submaximal tolerated dose is 600 mg.

68. 68. The pharmaceutical composition of claim 66 or 67, wherein the diftomenib or a pharmaceutically acceptable form thereof has breakthrough therapy designation.

69. 69. The pharmaceutical composition of any one of claims 66-68, wherein the pharmaceutical composition comprises one or more dosage forms, such as one or more oral dosage forms, optionally each oral dosage form comprising 50-600 mg of diftomenib, or optionally each oral dosage form comprising 50 mg, 100 mg, 200 mg, 300 mg, 400 mg, 500 mg, or 600 mg of diftomenib, wherein the total diftomenib in the one or more oral dosage forms is 600 mg.

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