Methods for identifying patients likely to benefit from treatment with telomerase inhibitors

Abstract

To provide methods of identifying or selecting a patient most likely to benefit from treatment with a telomerase inhibitor, such as imetelstat.SOLUTION: This disclosure provides methods of identifying or selecting a patient most likely to benefit from treatment with a telomerase inhibitor, such as imetelstat, by testing the patient for: a lack of a mutation in each of JAK2, CALR, and MPL; and / or a high-molecular risk (HMR), based on the presence of a mutation in at least one of the following genes: ASXL1, EZH2, SRSF2, and IDH1 / 2. The patient may suffer from myelofibrosis. The disclosure also provides methods of treating myelofibrosis, which include identifying such patients.SELECTED DRAWING: None

Description

[Technical Field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS Pursuant to 35 U.S.C. § 119(e), this application claims the benefit of priority to the filing dates of U.S. Provisional Patent Application No. 62 / 712,841, filed July 31, 2018, and U.S. Provisional Patent Application No. 62 / 772,849, filed November 29, 2018, the disclosures of which are incorporated herein by reference.

[0002] Sequence Listing This application contains a Sequence Listing which has been submitted electronically in ASCII format and is incorporated herein by reference in its entirety. The ASCII copy is named Sequence_Listing.txt and is 356 KB in size.

[0003] The present application relates to methods for identifying patients most likely to benefit from treatment with a telomerase inhibitor by identifying patients who lack mutations in each of JAK2, CALR, and MPL and / or have high molecular weight risk (HMR) based on the presence of a mutation in at least one of the following genes: ASXL1, EZH2, SRSF2, and IDH1 / 2. The present invention also relates to methods for treating myelofibrosis in a subject (i.e., a patient) in need of treatment with a telomerase inhibitor. [Background technology]

[0004] Preface Myelofibrosis (MF) is a classic BCR-ABL1-negative chronic myeloproliferative neoplasm (MPN) characterized by clonal myeloproliferation and dysregulated kinase signaling. (Cervantes, Blood, 124(17):2635-2642 (2014)) It is also characterized by cytopenia, systemic symptoms, and splenomegaly and may transform into acute myeloid leukemia. (Kuykendall et al., Annals of Hematology, 97:435-431 (2018)) MF is a Philadelphia chromosome-negative myeloproliferative neoplasm with a poor prognosis, and the JAK1 / JAK2 inhibitor ruxolitinib is currently the approved treatment. Ruxolitinib, a Janus kinase (JAK)-1 and JAK-2 inhibitor, is the first-in-class drug licensed in the United States for the treatment of high-risk and intermediate-risk MF. Pardanani, et al.; Blood Cancer J.; 4(12):e268(2014). Several other JAK inhibitors are in development, and some are currently undergoing phase 3 clinical trials. Ibid. Other treatment options for MF include allo-SCT, hydroxyurea, interferon, lenalidomide (Revlimid®), and thalidomide. Currently, there are ongoing clinical trials in MF to evaluate selective JAK inhibitors, histone deacetylase / DNA methyltransferase inhibitors, PI3K inhibitors, hedgehog / mammalian target of rapamycin (MTOR) inhibitors, antifibrotic agents, immunomodulators, monoclonal antibodies, and immune checkpoint inhibitors. Shreenivas, et al., Expert Opin Emerg Drugs, 23(1):37-49(2018).

[0005] Other MPNs include essential thrombocythemia (ET) and polycythemia vera (PV). Cervantes (see below). MF can appear de novo (primary MF [PMF]) or after previous ET or PV (post-ET or post-PV MF). Ibid. According to Cervantes, MF is a clonal proliferation of multipotent hematopoietic stem cells; the abnormal cell population releases several cytokines and growth factors into the bone marrow, which cause myelofibrosis and stromal changes, and colonizes extramedullary organs such as the spleen and liver. Ibid. Myelofibrosis has been associated with mutations in the Janus kinase (JAK) 2 gene (e.g., the V617F mutation), mutations in the thrombopoietin receptor gene (MPL), and mutations in the calreticulin gene (CALR). Ibid. It primarily affects the elderly, and according to Cervantes, "currently, there is no curative therapy other than allogeneic hematopoietic stem cell transplantation (allo-SCT), which can be applied to a minority of patients." Ibid.

[0006] Indeed, according to Langabeer, "The majority of patients with the classic myeloproliferative neoplasms (MPNs) of polycythemia vera, essential thrombocythemia, and primary myelofibrosis have distinct disease-driving mutations in the JAK2, CALR, or MPL genes." Langabeer, JAK-STAT, 5:e1248011 (2016). These mutations are so-called driver mutations. Exemplary driver mutations include mutations in JAK2 V617F, JAK2 exon 12, MPL exon 10, and CALR exon 9. Ibid.

[0007] According to Spiegel, in myelofibrosis (MF), driver mutations in JAK2, MPL, or CALR affect survival and progression to the blast phase, with the greatest risk conferred by triple-negative status (i.e., unmutated JAK2, MPL, and CALR) (Spiegel et al., Blood Adv., 1(20):1729-1738 (2017)). Indeed, the absence of JAK2 / MPL / CALR mutations (i.e., triple-negative status) is associated with the most unfavorable outcome (Pardanani, et al., Blood Cancer J.;4(12):e268 (2014) and also Tefferi et al., Blood, 124(16):2507-13 (2014)). Furthermore, mutations in high molecular weight risk (HMR) genes, such as ASXL1, EZH2, IDH1 / 2, and SRSF2, have also been associated with poor prognosis (Spiegel et al.). The presence of an increasing number of prognostically deleterious / "high molecular weight risk" mutations (i.e., ASXL1, EZH2, SRSF2, and / or IDH-1 / 2 genes) conferred progressively worse survival outcomes, independent of traditional risk factors. Guglielmelli et al., Leukemia, 28(9):1804-10(2014).

[0008] Driver mutations in JAK2, MPL, or CALR, alone or in combination with subclonal mutations in genes such as ASXL1, are associated with differences in overall survival (OS). Spiegel et al. Triple-negative patients without canonical mutations in JAK2, MPL, or CALR have an increased risk of leukemic transformation as well as shorter OS. Spiegel observed that for patients with myelofibrosis treated with ruxolitinib or momelotinib (JAK1 / 2 inhibitors), these mutations were associated with a shorter time to treatment failure. Ibid. Similarly, "Comparing the clinical characteristics of JAK2-positive, CALR-positive, MPL-positive, and TN MF patients, patients with CALR mutations had significantly lower hemoglobin (mean 8.6 vs. 10.7 g / dL; P 5.001) and white blood cell count (mean 11.0 vs. 25 g / dL; P 5.033), a trend reported in other MPN cohorts." Patel et al., Blood; 126(6):790-797 (2015). Patel et al. observed that patients treated with ruxolitinib with three or more mutations showed an inverse correlation with splenic response and time to treatment discontinuation. Driver mutations or triple-negative (JAK2, MPL, CALR) status have been found in myelofibrosis patients who discontinue JAK inhibitor treatment. See, for example, Kuykendall et al. [Prior art documents] [Non-patent literature]

[0009] [Non-Patent Document 1] Cervantes,Blood,124(17):2635-2642(2014) Summary of the Invention [Means for solving the problem]

[0010] The present invention provides a method for identifying or selecting patients most likely to benefit from treatment with a telomerase inhibitor, such as imetelstat, by testing the patient for the absence of mutations in each of the Janus kinase 2 (JAK2), calreticulin (CALR), and thrombopoietin receptor (MPL) genes, and / or for high molecular weight risk (HMR) based on the presence of mutations in at least one of the following genes: additional sex comb-like 1 (ASXL1), enhancer of zest homolog 2 (EZH2), serine- and arginine-rich splicing factor 2 (SRSF2), and isocitrate dehydrogenase 1 / 2 (IDH1 / 2). Patients in need of treatment may be suffering from myelofibrosis. The present invention also provides a method for treating myelofibrosis in a patient in need of such treatment, comprising identifying such a patient.

[0011] One embodiment of the present invention is a method for identifying myelofibrosis patients most likely to benefit from treatment with a telomerase inhibitor, comprising: (a) testing the patient for: (i) triple-negative status, based on the absence of mutations in each of the JAK2, CALR, and MPL genes, and / or (ii) a mutation in at least one of the following genes: ASXL1, EZH2, SRSF2, and IDH1 / 2; and (b) selecting the patient if the patient has: (i) triple-negative status, based on the absence of mutations in each of the JAK2, CALR, and MPL genes, and / or (ii) high molecular weight mutation (HMR), based on the presence of a mutation in at least one of the following genes: ASXL1, EZH2, SRSF2, and IDH1 / 2, wherein the selected patient is most likely to benefit from treatment with a telomerase inhibitor.

[0012] An alternative embodiment of the present invention is a method for identifying patients most likely to benefit from treatment with a telomerase inhibitor, comprising: (a) testing the patient for triple-negative status based on the absence of mutations in each of the JAK2, CALR, and MPL genes; and (b) selecting the patient if the patient has triple-negative status, wherein the selected patient is most likely to benefit from treatment with a telomerase inhibitor. An alternative embodiment of the present invention is a method for identifying patients most likely to benefit from treatment with a telomerase inhibitor, comprising: (a) testing the patient for high molecular weight risk (HMR) based on the presence of a mutation in at least one of the following genes: ASXL1, EZH2, SRSF2, and IDH1 / 2; and (b) selecting patients with high molecular weight risk (HMR) based on the presence of a mutation in at least one of the following genes: ASXL1, EZH2, SRSF2, and IDH1 / 2. The present invention further provides methods for treating myelofibrosis in patients with triple-negative and / or high molecular weight risk (HMR) with a telomerase inhibitor, such as imetelstat.

[0013] Another embodiment of the present invention is a method for identifying patients with myelofibrosis who are most likely to benefit from treatment with a telomerase inhibitor, comprising: (a) obtaining a DNA sample from the patient; (b) testing the DNA sample from such patient for (i) triple-negative status, based on the absence of mutations in each of the JAK2, CALR, and MPL genes, and / or (ii) high-molecular-weight risk (HMR), based on the presence of mutations in at least one of the following genes: ASXL1, EZH2, SRSF2, and IDH1 / 2; and (c) selecting patients if they have (i) triple-negative status, based on the absence of mutations in each of the JAK2, CALR, and MPL genes, and / or (ii) high-molecular-weight risk (HMR), based on the presence of mutations in at least one of the following genes: ASXL1, EZH2, SRSF2, and IDH1 / 2, and the selected patients are most likely to benefit from treatment with a telomerase inhibitor. In certain embodiments of the method, the DNA sample is obtained from bone marrow, peripheral blood, or both.

[0014] The DNA sample can be obtained by first obtaining a bone marrow sample, a peripheral blood sample, or both, and then isolating DNA from the bone marrow sample, the peripheral blood sample, or both. In one embodiment, obtaining a DNA sample from a patient includes obtaining a bone marrow sample from the patient, isolating cells from the bone marrow sample, and extracting DNA from the isolated cells. In another embodiment, obtaining a DNA sample from a patient includes obtaining a peripheral blood sample from the patient, isolating cells from the peripheral blood sample (e.g., granulocytes), and extracting DNA from the isolated cells.

[0015] Yet another embodiment of the present invention is a method for identifying patients with myelofibrosis most likely to benefit from treatment with a telomerase inhibitor, comprising testing the patient for (a) triple negative status based on the absence of any mutations in the JAK2, CALR, and MPL genes, (b) high molecular weight risk (HMR) based on the presence of a mutation in at least one of the following genes: ASXL1, EZH2, SRSF2, and IDH1 / 2, or (c) both, wherein the presence of (a), (b), or (c) indicates a patient most likely to benefit from treatment with a telomerase inhibitor.

[0016] In any of these methods, the patient may be suffering from myelofibrosis. The myelofibrosis may be primary myelofibrosis, myelofibrosis occurring after polycythemia vera (post-PV MF), or myelofibrosis occurring after essential thrombocythemia (post-ET MF). In certain embodiments, the patient has not previously received JAK inhibitor therapy. In other embodiments, the patient has previously received JAK inhibitor therapy and has "failed" JAK inhibitor therapy (i.e., the disease was resistant, or the patient was refractory to the therapy, or initially responded to treatment but the disease recurred). In other embodiments, the patient has previously received JAK inhibitor therapy and has discontinued JAK inhibitor therapy due to treatment-related toxicity or intolerance.

[0017] The method may also include administering a telomerase inhibitor to such a patient upon identification. In certain embodiments, the telomerase inhibitor is imetelstat or a pharmaceutically acceptable salt thereof. In other embodiments, the imetelstat is imetelstat sodium.

[0018] When imetelstat is used to treat patients identified by these methods, imetelstat is administered for 1, 2, 3, 4, 5, 6, 7, 8, or more than 8 administration cycles, each cycle comprising intravenously administering about 7-10 mg / kg of imetelstat once every 3 weeks, intravenously administering about 7-10 mg / kg of imetelstat once a week for 3 weeks, intravenously administering about 2.5-10 mg / kg of imetelstat once every 3 weeks, or intravenously administering about 0.5-9.4 mg / kg of imetelstat once every 3 weeks. In one embodiment, each administration cycle comprises intravenously administering about 7-10 mg / kg of imetelstat once every 3 weeks. In another embodiment, each administration cycle comprises intravenously administering about 9.4 mg / kg of imetelstat once every 3 weeks.

[0019] When imetelstat sodium is used to treat patients identified by these methods, imetelstat sodium is administered over 1, 2, 3, 4, 5, 6, 7, 8, or more than 8 administration cycles, each cycle comprising intravenously administering about 7-10 mg / kg of imetelstat sodium once every 3 weeks, intravenously administering about 7-10 mg / kg of imetelstat sodium once a week for 3 weeks, intravenously administering about 2.5-10 mg / kg of imetelstat sodium once every 3 weeks, or intravenously administering about 0.5-9.4 mg / kg of imetelstat sodium once every 3 weeks. In one embodiment, each administration cycle comprises intravenously administering about 7-10 mg / kg of imetelstat sodium once every 3 weeks. In another embodiment, each administration cycle comprises intravenously administering about 9.4 mg / kg of imetelstat sodium once every 3 weeks.

[0020] Another embodiment of the present invention is a method of treating a patient with myelofibrosis with a telomerase inhibitor, such as imetelstat or imetelstat sodium, comprising: (i) screening patients to determine whether such patients have triple negative status based on the absence of mutations in each of JAK2, CALR, and MPL, and / or whether they are at high molecular weight risk (HMR) based on the presence of a mutation in at least one of the following genes: ASXL1, EZH2, SRSF2, and IDH1 / 2; (ii) administering a telomerase inhibitor to the patient if the patient has triple-negative status based on the absence of mutations in any of JAK2, CALR, and MPL, and / or if the patient has high molecular weight risk (HMR) based on the presence of mutations in at least one of the following genes: ASXL1, EZH2, SRSF2, and IDH1 / 2. The myelofibrosis can be primary myelofibrosis, myelofibrosis occurring after polycythemia vera (post-PV MF), or myelofibrosis occurring after essential thrombocythemia (post-ET MF). In certain embodiments, the patient has not previously received JAK inhibitor therapy. In other embodiments, the patient has previously received JAK inhibitor therapy and has failed the JAK inhibitor therapy, or has previously received JAK inhibitor therapy and discontinued the JAK inhibitor therapy due to treatment-related toxicity or intolerance.

[0021] In certain embodiments of the method of treatment, the telomerase inhibitor is imetelstat, and is administered over one, two, three, four, five, six, seven, eight, or more than eight administration cycles, each cycle comprising intravenously administering about 7-10 mg / kg of imetelstat once every three weeks, intravenously administering about 7-10 mg / kg of imetelstat once a week for three weeks, intravenously administering about 2.5-10 mg / kg of imetelstat once every three weeks, or intravenously administering about 0.5-9.4 mg / kg of imetelstat once every three weeks. In certain embodiments, each administration cycle comprises intravenously administering about 7-10 mg / kg of imetelstat once every three weeks. In another embodiment, each administration cycle comprises intravenously administering about 9.4 mg / kg of imetelstat once every three weeks.

[0022] In some embodiments of the method for identifying or selecting patients most likely to benefit from treatment with a telomerase inhibitor, the method further comprises determining an average relative telomere length by analyzing the relative length of telomeric nucleic acids in target cells present in a biological sample from the patient. In some embodiments of the method for identifying or selecting patients most likely to benefit from treatment with a telomerase inhibitor, the method further comprises selecting patients identified as having an average relative telomere length in target cells present in a biological sample from the patient determined to be at or below the 50th percentile of a range of relative telomere lengths determined from one or more known standards. In certain embodiments, the telomerase inhibitor is imetelstat or a pharmaceutically acceptable salt thereof. In other embodiments, the imetelstat is imetelstat sodium.

[0023] The present disclosure provides a method for treating a patient with myelofibrosis with a telomerase inhibitor, comprising administering the telomerase inhibitor to the patient when the patient has a triple-negative status based on the absence of mutations in JAK2, CALR, and MPL. In certain embodiments, the telomerase inhibitor is imetelstat or a pharmaceutically acceptable salt thereof. In other embodiments, the imetelstat is imetelstat sodium.

[0024] The present disclosure provides methods of treating a patient having myelofibrosis with a telomerase inhibitor, comprising administering the telomerase inhibitor to the patient if such patient has triple negative status based on the absence of mutations in each of JAK2, CALR, and MPL, and / or has high molecular weight risk (HMR) based on the presence of a mutation in at least one of the following genes: ASXL1, EZH2, SRSF2, and IDH1 / 2.

[0025] The present disclosure provides a method of treating patients with myelofibrosis with a telomerase inhibitor, wherein such patients have the following characteristics: (a) the average relative telomere length of target cells present in a biological sample from the individual, determined to be less than or equal to the 50th percentile of a range of relative telomere lengths determined from one or more known standards; (b) triple-negative status, based on the absence of mutations in each of JAK2, CALR, and MPL; (c) administering a telomerase inhibitor to a patient if the patient has one or more of high molecular weight mutations (HMR) based on the presence of mutations in at least one of the following genes: ASXL1, EZH2, SRSF2, and IDH1 / 2. In certain embodiments, the telomerase inhibitor is imetelstat or a pharmaceutically acceptable salt thereof. In other embodiments, the imetelstat is imetelstat sodium.

[0026] The present disclosure provides a method for identifying a subject having myelofibrosis (MF) for treatment with a telomerase inhibitor, the method comprising measuring an hTERT expression level in a biological sample obtained from the patient after administration of the telomerase inhibitor and comparing the hTERT expression level in the biological sample with a baseline hTERT expression level before administration of the telomerase inhibitor, wherein a decrease in the hTERT expression level in the biological sample identifies a patient who is likely to benefit from treatment with the telomerase inhibitor.

[0027] The present disclosure provides a method for treating myelofibrosis (MF), comprising administering an effective amount of a telomerase inhibitor to a subject in need thereof and assessing hTERT expression levels in a biological sample obtained from the patient after administration of the telomerase inhibitor. In certain embodiments, the telomerase inhibitor is imetelstat or a pharmaceutically acceptable salt thereof. In other embodiments, the imetelstat is imetelstat sodium.

[0028] The present disclosure provides a method for monitoring the effectiveness of treatment in a subject with myelofibrosis (MF), the method comprising measuring hTERT expression levels in a biological sample obtained from the patient after administration of a telomerase inhibitor and comparing the hTERT expression level in the biological sample with a baseline hTERT expression level before administration of the telomerase inhibitor, wherein a 50% or greater decrease in the hTERT expression level in the biological sample identifies a subject likely to benefit from treatment with the telomerase inhibitor. In certain embodiments, the telomerase inhibitor is imetelstat or a pharmaceutically acceptable salt thereof. In other embodiments, the imetelstat is imetelstat sodium.

[0029] The present disclosure provides a method for selecting patients most likely to benefit from treatment with a telomerase inhibitor, the method comprising: testing a patient for average relative telomere length by analyzing the relative length of telomeric nucleic acid in target cells present in a biological sample from the patient; and selecting a patient if the patient has an average relative telomere length in target cells present in the biological sample from the patient that is determined to be less than or equal to the 50th percentile of a range of relative telomere lengths determined from one or more known standards, wherein the selected patient is most likely to benefit from treatment with a telomerase inhibitor.

[0030] The present disclosure provides a method for identifying patients most likely to benefit from treatment with a telomerase inhibitor, comprising obtaining a biological sample from the patient; determining an average relative length by analyzing the relative length of telomeric nucleic acid in target cells present in the biological sample from the patient; and identifying the patient if the patient has an average relative telomere length in the target cells present in the biological sample from the patient that is determined to be less than or equal to the 50th percentile of a range of relative telomere lengths determined from one or more known standards, wherein the identified patient is most likely to benefit from treatment with a telomerase inhibitor.

[0031] The present disclosure provides a method for treating a patient with myelofibrosis with a telomerase inhibitor, comprising administering the telomerase inhibitor to the patient when the patient has an average relative telomere length in target cells present in a biological sample from the patient that is determined to be equal to or less than the 50th percentile of a relative telomere length range determined from one or more known standards. In certain embodiments, the telomerase inhibitor is imetelstat or a pharmaceutically acceptable salt thereof. In other embodiments, the imetelstat is imetelstat sodium.

[0032] The present disclosure provides a method for monitoring the effectiveness of treatment in a subject with myelofibrosis (MF), the method comprising measuring hTERT expression levels in a biological sample obtained from the patient after administration of a telomerase inhibitor and comparing the hTERT expression level in the biological sample with the baseline hTERT expression level before administration of the telomerase inhibitor, wherein a 50% or greater decrease in the hTERT expression level in the biological sample identifies a subject likely to benefit from treatment with the telomerase inhibitor. In certain embodiments, the measured or assessed hTERT expression level is hTERT RNA expression level. In certain embodiments, the telomerase inhibitor is imetelstat or a pharmaceutically acceptable salt thereof. In other embodiments, the imetelstat is imetelstat sodium.

[0033] The present disclosure provides a method for identifying a patient having myelofibrosis (MF) for treatment with a telomerase inhibitor, the method comprising measuring an hTERT expression level in a biological sample obtained from the patient after administration of a telomerase inhibitor, and comparing the hTERT expression level in the biological sample with a baseline hTERT expression level before administration of the telomerase inhibitor, wherein a decrease in the hTERT expression level in the biological sample identifies patients who are likely to benefit from treatment with the telomerase inhibitor.

[0034] The present disclosure provides a method for monitoring the effectiveness of treatment in a subject with myelofibrosis (MF), the method comprising measuring the level of telomerase activity in a biological sample obtained from the patient after administration of a telomerase inhibitor and comparing the level of telomerase activity in the biological sample with the baseline telomerase activity level before administration of the telomerase inhibitor, wherein a 50% or greater decrease in the level of telomerase activity in the biological sample identifies a subject likely to benefit from treatment with the telomerase inhibitor. In certain embodiments, the telomerase inhibitor is imetelstat or a pharmaceutically acceptable salt thereof. In other embodiments, the imetelstat is imetelstat sodium. The present invention provides, for example, the following items. (Item 1) 1. A method for selecting patients most likely to benefit from treatment with a telomerase inhibitor, comprising: (a) testing the patient for triple-negative status, wherein the triple-negative status comprises the absence of mutations in each of the Janus kinase 2 (JAK2), calreticulin (CALR), and thrombopoietin receptor (MPL) genes; and / or (b) testing the patient to determine whether said patient has HMR, wherein having HMR comprises the presence of a mutation in at least one gene selected from the group consisting of additional sex comb-like 1 (ASXL1), enhancer of zest homolog 2 (EZH2), serine- and arginine-rich splicing factor 2 (SRSF2), and isocitrate dehydrogenase 1 / 2 (IDH1 / 2); and / or (c) testing the patient for average relative telomere length by analyzing the relative length of telomeric nucleic acid of target cells present in a biological sample from the patient; The patient if the patient has triple negative status; or the patient has HMR, or selecting the patient if the patient has a mean relative telomere length of target cells present in a biological sample from the patient that is determined to be less than or equal to the 50th percentile of a range of relative telomere lengths determined from one or more known standards; The method wherein the selected patients are most likely to benefit from treatment with a telomerase inhibitor. (Item 2) 2. The method of claim 1, wherein the method comprises testing the patient for triple-negative status and selecting the patient if the patient does not have a mutation in each of the JAK2, CALR, and MPL genes. (Item 3) 2. The method of claim 1, wherein the method comprises testing the patient to determine whether the patient has HMR, and wherein selecting the patient comprises the presence of a mutation in at least one gene selected from ASXL1, EZH2, SRSF2, and IDH1 / 2. (Item 4) 2. The method of claim 1, wherein the method comprises testing a patient for an average relative telomere length by analyzing the relative length of telomeric nucleic acid of target cells present in a biological sample from the patient; and selecting the patient if the patient has an average relative telomere length of target cells present in the biological sample from the patient that is determined to be less than or equal to the 50th percentile of a range of relative telomere lengths determined from one or more known standards. (Item 5) 5. The method according to any one of items 1 to 4, wherein the myelofibrosis is selected from the group consisting of primary myelofibrosis, myelofibrosis that develops after polycythemia vera (post-PV MF), and myelofibrosis that develops after essential thrombocythemia (post-ET MF). (Item 6) 6. The method of any one of items 1 to 5, wherein the patient has not previously received JAK inhibitor therapy. (Item 7) The patient: have received JAK inhibitor therapy and the patient was resistant to JAK inhibitor therapy; have previously received JAK inhibitor therapy and have relapsed; or 6. The method of any one of items 1 to 5, wherein the patient has received JAK inhibitor therapy and discontinued the JAK inhibitor therapy due to treatment-related toxicity or intolerance. (Item 8) 8. The method according to any one of items 1 to 7, wherein the telomerase inhibitor is imetelstat. (Item 9) 9. The method of any one of items 1 to 8, further comprising obtaining a sample comprising DNA from the patient, wherein the sample comprises bone marrow, peripheral blood, or a combination thereof. (Item 10) 1. Use of a telomerase inhibitor in the treatment of a patient with myelofibrosis, comprising: (a) the patient is determined to have triple-negative status, wherein the triple-negative status comprises the absence of mutations in each of the JAK2, CALR, and MPL genes; and / or (b) the patient is determined to have high molecular weight mutation (HMR), where HMR comprises the presence of a mutation in at least one gene selected from the group consisting of ASXL1, EZH2, SRSF2, and IDH1 / 2; and / or (c) the patient has myelofibrosis, wherein cells present in a biological sample from the patient have been determined to have an average relative telomere length determined to be less than or equal to the 50th percentile of a range of relative telomere lengths determined from one or more known standards. (Item 11) 11. The use of item 10, wherein the patient is determined to have triple-negative status, wherein the triple-negative status comprises the absence of mutations in each of the JAK2, CALR, and MPL genes. (Item 12) 11. The use of item 10, wherein the patient is determined to have high molecular weight mutation (HMR), wherein having HMR comprises the presence of a mutation in at least one gene selected from the group consisting of ASXL1, EZH2, SRSF2, and IDH1 / 2. (Item 13) 11. The use of item 10, wherein the patient has myelofibrosis, and wherein cells present in a biological sample from the patient have a mean relative telomere length determined to be less than or equal to the 50th percentile of a range of relative telomere lengths determined from one or more known standards. (Item 14) 14. The use according to any one of items 10 to 13, wherein the myelofibrosis is selected from the group consisting of primary myelofibrosis, myelofibrosis that develops after polycythemia vera (post-PV MF), and myelofibrosis that develops after essential thrombocythemia (post-ET MF). (Item 15) 15. The use of any one of items 10 to 14, wherein the patient has not previously received JAK inhibitor therapy. (Item 16) The patient: have received JAK inhibitor therapy and the patient was resistant to JAK inhibitor therapy; have previously received JAK inhibitor therapy and have relapsed; or 15. The use of any one of items 10 to 14, in which the patient has received JAK inhibitor therapy and discontinued the JAK inhibitor therapy due to treatment-related toxicity or intolerance. (Item 17) 17. The method according to any one of items 10 to 16, wherein the telomerase inhibitor is imetelstat. [Brief explanation of the drawings]

[0035] The foregoing summary, as well as the following detailed description of the invention, will be better understood when read in conjunction with the accompanying drawings. For the purpose of illustrating the invention, the drawings demonstrate embodiments of the invention. It should be understood, however, that the invention is not limited to the precise arrangements, examples, and apparatus shown.

[0036] [Figure 1]1 shows a waterfall plot of spleen volume reduction (SVR) at week 24 for the 4.7 mg / kg and 9.4 mg / kg treatment groups of Example 1. SVR is presented as percent change from baseline. [Figure 2] 1 shows a waterfall plot of total symptom score reduction (TSS) at week 24 for the 4.7 mg / kg and 9.4 mg / kg treatment groups in Example 1. TSS is presented as percent change from baseline. [Figure 3] Kaplan-Meier plot of overall survival grouped by mutation status of the JAK2 / MPL / CALR genes: TN vs. non-TN (MUT) for the 4.7 mg / kg group. Specifically, Figure 3 shows the survival probability for patients with triple-negative status (TN) and patients with at least one mutation (MUT) as a function of time. [Figure 4] Kaplan-Meier plot of overall survival grouped by mutation status of the JAK2 / MPL / CALR genes: TN vs. non-TN (MUT) for the 9.4 mg / kg group. Specifically, Figure 4 shows the survival probability for patients with triple-negative status (TN) and patients with at least one mutation (MUT) as a function of time for the 9.4 mg / kg group. [Figure 5] 1 shows a Kaplan-Meier plot of overall survival as a function of time grouped according to patients in the 9.4 mg / kg vs. 4.7 mg / kg groups. [Figure 6] Kaplan-Meier plot of overall survival (OS) grouped by JAK2 / MPL / CALR gene mutation status: TN vs. non-TN for the 9.4 mg / kg group. Specifically, Figure 6 shows the survival probability for patients with triple-negative status (TN) and patients with at least one mutation (non-TN) as a function of time for the 9.4 mg / kg group. [Figure 7]Kaplan-Meier plot of overall survival (OS) grouped by JAK2 / MPL / CALR gene mutation status: TN vs. non-TN for the 4.7 mg / kg group. Specifically, Figure 7 shows the survival probability for patients with triple-negative status (TN) and patients with at least one mutation (non-TN) as a function of time for the 4.7 mg / kg group. DETAILED DESCRIPTION OF THE INVENTION

[0037] This application is based on the discovery that patients with myelofibrosis who are triple-negative (i.e., lacking mutations in JAK2, CALR, and MPL) and / or in the high-risk (HMR) category based on the presence of mutations in at least one of the following genes: ASXL1, EZH2, SRSF2, and IDH1 / 2, can benefit from treatment with a telomerase inhibitor, such as imetelstat or imetelstat sodium. Patients with mutations in the ASXL1, EZH1, IDH1 / 2, and SRSF2 genes are at increased risk of early death or leukemic transformation. These patients typically do not benefit from treatment with conventional therapies, such as JAK inhibitors. Gisslinger et al., Blood, 128:1931 (2016). Therefore, the fact that these patients benefit from treatment with a telomerase inhibitor is unexpected and surprising.

[0038] Thus, the present application provides a method for identifying patients most likely to benefit from treatment with a telomerase inhibitor, such as imetelstat. The method includes testing or identifying a patient to determine whether the patient has triple-negative status based on the absence of mutations in each of JAK2, CALR, and MPL, and / or whether the patient has high molecular weight risk (HMR) based on the presence of a mutation in at least one of the following genes: ASXL1, EZH2, SRSF2, and IDH1 / 2. The present application also provides a method for treating myelofibrosis with a telomerase inhibitor, such as imetelstat, which includes identifying a patient as having triple-negative status based on the absence of mutations in each of JAK2, CALR, and MPL, and / or having high molecular weight risk (HMR) based on the presence of a mutation in at least one of the following genes: ASXL1, EZH2, SRSF2, and IDH1 / 2. Such patients are most likely to benefit from treatment with a telomerase inhibitor. A telomerase inhibitor (e.g., imetelstat) is then administered to the patient. For clarity of disclosure, and not by way of limitation, the detailed description of the invention is divided into subsections that describe or illustrate particular features, embodiments or applications of the invention.

[0039] A.Definition As used herein, mutations in additional sex comb-like 1 (ASXL1), enhancer of zest homolog 2 (EZH2), serine- and arginine-rich splicing factor 2 (SRSF2), and isocitrate dehydrogenase 1 / 2 (IDH1 / 2) are intended to include any mutations in these genes that affect survival and disease progression in patients with myelofibrosis. Furthermore, as used herein, IDH1 / 2 is intended to include IDH1 and IHD2. Exemplary mutations can be found in the following publications, the disclosures of each of which are incorporated herein for their relevance to the disclosure of genetic mutations associated with myelofibrosis: Langabeer, JAK-STAT,5:e1248011(2016), Cervantes,Blood;124(17):2635-2642(2014), Patel et al.,Blood;126(6):790-797(2015), Spiegel et al.,Blood Adv.,1(20):1729-1738(2017), Newburry et al.,Blood,130(9):1125-1131(2017), Kuykendall et al.Annals of Hematology,97:435-431(2018). Exemplary sequences are as follows: High molecular weight risk (HMR) can be determined based on the presence of a mutation in at least one of the following genes: for example, the ASXL1 gene having the nucleic acid sequence of SEQ ID NO: 5, for example, the EZH2 gene having the nucleic acid sequence of SEQ ID NO: 6, for example, the SRSF2 gene having the nucleic acid sequence of SEQ ID NO: 7, for example, the IDH1 gene having the nucleic acid sequence of SEQ ID NO: 8, the IDH2 gene having the nucleic acid sequence of SEQ ID NO: 9, and combinations thereof.

[0040] In some embodiments, mutations of interest in the ASXL1 gene include Q575, Q588, Y591, Q592, S604, L614, Q623, A627, E635, T638, A640, G646, G658, R678, C687, D690, R693, Y700, G704, E705, Q708, G710, L721, E727, V751, P763, Q780, These mutations include W796, V807, T822, K825, S846, D855, C856, L857, L885, L890, S903, S970, Y974, R965, G967, V962, L992, S1028, Q1039, R1073, E1102, H1153, S1209, S1231, A1312, F1305, P1377, R1415, and I1436. In some embodiments, the mutations are a Q575X mutation, a Q588X mutation, a Y591X mutation, a Y591N mutation, a Q592X mutation, a S604F mutation, a L614F mutation, a Q623X mutation, a A627G mutation, a E635R mutation, a T638V mutation, a A640G mutation, a G646W mutation, a G658X mutation, a R678K mutation, a C687R mutation, a C687V mutation, a D690G mutation, a R693X mutation, a Y700X mutation, a G704R mutation, a G704W mutation, an E705X mutation, a Q708X mutation, a G710E mutation, a L721C mutation, an E727X mutation, a V751L mutation, a P763R mutation, a Q780X mutation, a W796X mutation, a mutations, W796G mutation, V807F mutation, T822H mutation, K825X mutation, S846Q mutation, D855A mutation, C856X mutation, L857R mutation, L885X mutation, L890F mutation, S903I mutation, S970N mutation, Y974X mutation, R965X mutation, G967del mutation, V962A mutation, L992Q mutation, S1028R mutation, Q1039L mutation, R1073C mutation, E1102D mutation, H1153R mutation, S1209I mutation, S1231F mutation, A1312V mutation, F1305W mutation, P1377S mutation, R1415Q mutation, and / or I1436M mutation.

[0041] In some embodiments, mutations of interest in the EZH2 gene include mutations at W60, R63, P312, F145, N182, R288, Q328, Q553, R566, T573, R591, R659, D677, V679, R690, A702, V704, E726, D730, and / or Y733. In some embodiments, the mutation is a W60X mutation, an R63X mutation, a P312S mutation, an F145S mutation, an N182D mutation, an R288Q mutation, a Q328X mutation, a Q553X mutation, an R566H mutation, a T573I mutation, an R591H mutation, an R659K mutation, a D677H mutation, a V679M mutation, an R690H mutation, an A702V mutation, a V704L mutation, an E726V mutation, a D730X mutation, and / or a Y733X mutation.

[0042] In some embodiments, the mutation of interest in the SRSF2 gene comprises a mutation in P95. In some embodiments, the mutation is a P95H mutation, a P95L mutation, or a P95R mutation.

[0043] In some embodiments, the mutation of interest in the IDH1 / 2 gene comprises a mutation at R132 and / or R140. In some embodiments, the mutation is an R132G mutation, an R132H mutation, or an R140Q mutation.

[0044] In certain embodiments, mutations of interest include those set forth below: [Table A]

[0045] As used herein, "triple-negative status," "triple-negative," or "TN" refers to a patient who lacks mutations in each of the Janus kinase 2 (JAK2), calreticulin (CALR), and thrombopoietin receptor (MPL) genes. Triple-negative status can be determined based on the absence of mutations in, for example, the JAK2 gene having the nucleic acid sequence of SEQ ID NO: 2, the CALR gene having the nucleic acid sequence of, for example, SEQ ID NO: 3, and the MPL gene having the nucleic acid sequence of, for example, SEQ ID NO: 4.

[0046] In certain embodiments, the triple-negative status includes the absence of mutations in the JAK2 gene, such as mutations at G335, F556, G571, V617, and / or V625. For example, the triple-negative status may include the absence of G335D, F556V, G571S, V617F, and / or V625S mutations in the JAK2 gene.

[0047] In certain embodiments, triple-negative status includes the absence of mutations in the MPL gene, such as mutations at T119, S204, P222, E230, V285, R321, S505, W515, Y591, and / or R592. For example, triple-negative status can include the absence of a T119I mutation, a S204F mutation, a S204P mutation, a P222S mutation, an E230G mutation, a V285E mutation, a R321W mutation, a S505N mutation, a W515R mutation, a W515L mutation, a Y591N mutation, and / or a R592Q mutation in the MPL gene.

[0048] In certain embodiments, the triple-negative status includes the absence of mutations in the CALR gene, such as mutations at L367, K368, E381, K385, and / or E396. For example, the triple-negative status may include the absence of L367T, K368R, K385N, E381A, and / or E396del mutations in the CALR gene.

[0049] In certain embodiments, mutations of interest include those set forth below: [Table B]

[0050] As used herein, a patient may have "failed" JAK inhibitor therapy when the disease was resistant or when the patient was refractory to treatment, or when the disease recurred after initially responding to treatment.

[0051] As used herein, when referring to a measurable value such as an amount, time duration, etc., the term "about" is meant to encompass a variation of ±20% to ±0.1%, preferably ±20% to ±10%, more preferably ±5%, even more preferably ±1%, and even more preferably ±0.1% from the specified value, as appropriate for performing the disclosed methods.

[0052] The term "pharmaceutically acceptable salt" refers to a salt that is acceptable for administration to a patient, such as a mammal (a salt having a counterion that is acceptable for a given dosage regimen and safe for the mammal). Such salts can be derived from pharmaceutically acceptable inorganic or organic bases and pharmaceutically acceptable inorganic or organic acids. "Pharmaceutically acceptable salt" refers to a pharmaceutically acceptable salt of a compound, which salt is derived from a variety of organic and inorganic counterions well known in the art, including, by way of example only, sodium, and, when the molecule contains a basic functional group, includes salts of organic or inorganic acids, such as hydrochloride. Pharmaceutically acceptable salts of interest include, but are not limited to, aluminum, ammonium, arginine, barium, benzathine, calcium, cholinate, ethylenediamine, lysine, lithium, magnesium, meglumine, procaine, potassium, sodium, tromethamine, N-methylglucamine, N,N'-dibenzylethylenediamine, chloroprocaine, diethanolamine, ethanolamine, piperazine, zinc, diisopropylamine, diisopropylethylamine, triethylamine, and triethanolamine salts.

[0053] The term "salt(s) thereof" refers to a compound formed when an acid proton is replaced with a cation, such as a metal cation or an organic cation. Preferably, the salt is a pharmaceutically acceptable salt. By way of example, salts of the present compounds include those in which the compound is protonated with an inorganic or organic acid to form a cation, with the conjugate base of the inorganic or organic acid serving as the anionic component of the salt. Salts of interest include, but are not limited to, aluminum, ammonium, arginine, barium, benzathine, calcium, cesium, cholinate, ethylenediamine, lithium, magnesium, meglumine, procaine, N-methylglucamine, piperazine, potassium, sodium, tromethamine, zinc, N,N'-dibenzylethylenediamine, chloroprocaine, diethanolamine, ethanolamine, piperazine, diisopropylamine, diisopropylethylamine, triethylamine, and triethanolamine salts. It should be understood that for any of the oligonucleotide structures depicted herein, including a backbone of internucleoside linkages, such oligonucleotides may include any convenient salt form. In some embodiments, the acidic form of the internucleoside bond is shown for simplicity. In some cases, the salt of the subject compound is a monovalent cation salt. In certain cases, the salt of the subject compound is a divalent cation salt. In some cases, the salt of the subject compound is a trivalent cation salt. A "solvate" refers to a complex formed by the combination of a solvent molecule with a solute molecule or ion. The solvent can be an organic compound, an inorganic compound, or a mixture of both. Some examples of solvents include, but are not limited to, methanol, N,N-dimethylformamide, tetrahydrofuran, dimethyl sulfoxide, and water. When the solvent is water, the solvate formed is a hydrate.

[0054] "Stereoisomer" and "stereoisomer" refer to compounds that have the same atomic connectivity but different atomic arrangements in space. Stereoisomers include, for example, cis-trans isomers, E and Z isomers, enantiomers, and diastereomers. With respect to any of the groups disclosed herein that contain one or more substituents, it is understood that such groups do not contain any substitutions or substitution patterns that are sterically impractical and / or synthetically infeasible. All stereoisomers are intended to be included within the scope of this disclosure.

[0055] Those skilled in the art will recognize that other tautomeric configurations of the groups described herein are possible, and it is understood that all tautomeric forms of the subject compounds are encompassed by the depicted structures, even if not specifically shown, where one possible tautomeric configuration of the groups of the compounds is encompassed by the depicted structures.

[0056] Pharmaceutically acceptable salt solvates of the stereoisomeric tautomers of the subject compounds are intended to be included and are intended to be included within the scope of this disclosure.

[0057] Before certain embodiments are described in more detail, it is to be understood that the invention is not limited to particular embodiments described, as such may, of course, vary. It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only, and is not intended to be limiting, since the scope of the present invention will be limited only by the appended claims.

[0058] Where a range of values ​​is provided, unless the context clearly dictates otherwise, it is understood that each intervening value, to the tenth of the unit of the lower limit, between the upper and lower limit of that range and any other stated or intervening value in that range, is encompassed within the invention. The upper and lower limits of these smaller ranges may independently be included in the smaller ranges and are also encompassed within the invention, subject to any specifically excluded limit in the stated range. Where the stated range includes one or both of the limits, ranges excluding either or both of those included limits are also included in the invention.

[0059] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Although any methods and materials similar or equivalent to those described herein can also be used in the practice or testing of the present invention, representative exemplary methods and materials are described herein.

[0060] All publications and patents cited herein are incorporated by reference to the same extent as if each individual publication or patent was specifically and individually indicated to be incorporated by reference, and are incorporated by reference herein to disclose and describe the methods and / or materials in connection with the cited publications. The citation of a publication is for its disclosure prior to the filing date and should not be construed as an admission that the present invention is not entitled to antedate such publication by virtue of prior invention. Further, the dates of publication provided may be different from the actual publication dates, which may need to be independently confirmed.

[0061] It should be noted that, as used in this specification and the appended claims, the singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise. It should be further noted that the claims may be drafted to exclude any optional element. As such, this statement is intended to serve as a predicate for the use of exclusive terminology such as "solely," "only," or the use of a "negative" limitation in connection with the recitation of claim elements.

[0062] Each of the individual embodiments described and illustrated herein has distinct components and features which may be readily separated from or combined with the features of any of the other embodiments without departing from the scope or spirit of the invention. Any recited method may be carried out in the order of events recited or in any other order which is logically possible.

[0063] B. Identifying Patients Most Likely to Benefit from Telomerase Inhibitor Treatment In one aspect, the present disclosure provides a method for identifying or selecting myelofibrosis patients who are most likely to benefit from treatment with a telomerase inhibitor. The method relies on identifying triple-negative status patients (patients who lack mutations in each of the JAK2, CALR, and MPL genes) or patients with high molecular weight risk (HMR) based on the presence of mutations in at least one of the following genes: ASXL1, EZH2, SRSF2, and IDH1 / 2. These triple-negative or HMR patients are most likely to benefit from treatment with a telomerase inhibitor, such as imetelstat or imetelstat sodium.

[0064] The myelofibrosis can be primary myelofibrosis, myelofibrosis occurring after polycythemia vera (post-PV MF), or myelofibrosis occurring after essential thrombocythemia (post-ET MF). In certain embodiments, the patient has not previously received JAK inhibitor therapy. In other embodiments, the patient has previously received JAK inhibitor therapy and has failed JAK inhibitor therapy (i.e., the disease was resistant, or the patient was refractory to the therapy, or initially responded to treatment but the disease relapsed). In other embodiments, the patient has previously received JAK inhibitor therapy and has discontinued JAK inhibitor therapy due to treatment-related toxicity or intolerance. In yet alternative embodiments, the patient has previously received JAK inhibitor therapy and has discontinued JAK inhibitor therapy.

[0065] In one embodiment, the patient has received JAK inhibitor therapy and the myelofibrosis is resistant to JAK inhibitor therapy. In another embodiment, the patient has received JAK inhibitor therapy and the patient is resistant to JAK inhibitor therapy. In another embodiment, the patient has received JAK inhibitor therapy and the patient has relapsed. In an alternative embodiment, the patient has received JAK inhibitor therapy and discontinued JAK inhibitor therapy due to treatment-related toxicity or intolerance.

[0066] In one embodiment, the present invention provides a method for selecting patients most likely to benefit from treatment with a telomerase inhibitor by testing for one or more of triple-negative status based on the absence of mutations (i.e., the absence of any mutations) in each of the JAK2, CALR, and MPL genes. In that embodiment, patients may also be tested for high molecular weight risk (HMR) based on the presence of mutations in at least one of the following genes: ASXL1, EZH2, SRSF2, and IDH1 / 2. In another embodiment, the present invention provides a method for selecting patients most likely to benefit from treatment with a telomerase inhibitor by testing for triple-negative status based on the absence of mutations (i.e., the absence of any mutations) in each of the JAK2, CALR, and MPL genes and / or high molecular weight risk (HMR) based on the presence of mutations in at least one of the following genes: ASXL1, EZH2, SRSF2, and IDH1 / 2. In another embodiment, the present invention provides a method for identifying patients most likely to benefit from treatment with a telomerase inhibitor, comprising testing the patient for (a) triple-negative status based on the absence of any mutations in the JAK2, CALR, and MPL genes, (b) high molecular weight risk (HMR) based on the presence of a mutation in at least one of the following genes: ASXL1, EZH2, SRSF2, and IDH1 / 2, or (c) both. In this embodiment, the presence of (a), (b), or (c) indicates a patient most likely to benefit from treatment with a telomerase inhibitor.

[0067] Another embodiment of the present invention is a method for identifying patients most likely to benefit from treatment with a telomerase inhibitor, comprising: a. Examining the patient for: i. Triple-negative status based on the absence of mutations in each of the JAK2, CALR, and MPL genes, and / or ii. Testing for high molecular weight risk (HMR), based on the presence of a mutation in at least one of the following genes: ASXL1, EZH2, SRSF2, and IDH1 / 2; b. If the patient has: i. Triple-negative status based on the absence of mutations in each of the JAK2, CALR, and MPL genes, and / or ii. selecting patients as having high molecular weight risk (HMR) based on the presence of a mutation in at least one of the following genes: ASXL1, EZH2, SRSF2, and IDH1 / 2; The selected patients are those most likely to benefit from treatment with a telomerase inhibitor.

[0068] Yet another embodiment of the present invention is a method for identifying patients most likely to benefit from treatment with a telomerase inhibitor, comprising: testing a patient for triple-negative status based on the absence of mutations in each of the JAK2, CALR, and MPL genes; and selecting the patient if the patient has triple-negative status based on the absence of mutations in each of the JAK2, CALR, and MPL genes, wherein the selected patient is most likely to benefit from treatment with a telomerase inhibitor. In one embodiment, the method also comprises testing the patient for high molecular weight risk (HMR) based on the presence of a mutation in at least one of the following genes: ASXL1, EZH2, SRSF2, and IDH1 / 2; and selecting the patient if the patient has HMR.

[0069] In certain embodiments of any of these methods, the triple-negative patient lacks mutations in the coding regions (exons) of the JAK2, CALR, and MPL genes.

[0070] Additionally, in other embodiments of any of these methods, high molecular weight risk (HMR) is determined by the presence of a mutation in the coding region (exon) of at least one of the ASXL1, EZH2, SRSF2, and IDH1 / 2 genes.

[0071] In certain embodiments, high molecular weight risk (HMR) is determined by detecting the presence of a mutation in ASXL1, EZH2, SRSF2, or IDH1 / 2, or a combination thereof. In some embodiments, the method comprises detecting the presence of a mutation in ASXL1. In some embodiments, the method comprises detecting the presence of a mutation in EZH2. In some embodiments, the method comprises detecting the presence of a mutation in SRSF2. In some embodiments, the method comprises detecting the presence of a mutation in IDH1 / 2. In some embodiments, the method comprises detecting the presence of a mutation in ASXL1 and EZH2. In some embodiments, the method comprises detecting the presence of a mutation in ASXL1 and SRSF2. In some embodiments, the method comprises detecting the presence of a mutation in ASXL1 and IDH1 / 2. In some embodiments, the method comprises detecting the presence of a mutation in EZH2, SRSF2. In some embodiments, the method comprises detecting the presence of a mutation in EZH2 and IDH1 / 2. In some embodiments, the method comprises detecting the presence of a mutation in SRSF2 and IDH1 / 2. In some embodiments, the method comprises detecting the presence of a mutation in ASXL1, EZH2, and SRSF2. In some embodiments, the method comprises detecting the presence of mutations in ASXL1, EZH2, and IDH1 / 2. In some embodiments, the method comprises detecting the presence of mutations in EZH2, SRSF2, and IDH1 / 2. In some embodiments, the method comprises detecting the presence of mutations in ASXL1, EZH2, SRSF2, and IDH1 / 2. In yet another embodiment, the present invention provides a method for identifying or selecting patients in a patient population who are most likely to benefit from treatment with a telomerase inhibitor. In this method, the patient population is screened for patients with mutations in each of the JAK2, CALR, and MPL genes, and triple-negative patients within the population are identified. In an alternative embodiment, the method relies on identifying triple-negative patients who lack canonical mutations in each of JAK2, MPL, and CALR.

[0072] In certain embodiments, the method also includes a step of collecting a patient DNA sample. The patient sample may be collected from a DNA sample obtained from bone marrow, peripheral blood, or both. Thus, in certain embodiments, the method includes obtaining a patient blood sample and isolating (extracting) DNA from the patient blood sample. The method may also include a step of isolating cells (e.g., granulocytes) from the patient blood sample. Similarly, the method may include obtaining a bone marrow sample and isolating (extracting) DNA from the bone marrow sample. The method may also include a step of isolating cells from a patient bone sample.

[0073] The patient DNA sample is tested for the presence or absence of mutations in each of the JAK2, CALR, and MPL genes using conventional techniques. Alternatively, the patient DNA sample is tested for the presence of mutations in at least one of the following genes: ASXL1, EZH2, SRSF2, and IDH1 / 2 using conventional techniques. In certain embodiments, the patient DNA sample is tested for (i) the presence or absence of mutations in each of the JAK2, CALR, and MPL genes, and (ii) the presence of mutations in at least one of the following genes: ASXL1, EZH2, SRSF2, and IDH1 / 2.

[0074] In certain embodiments, testing of the DNA sample may be a next-generation sequencing assay using the Illumina MiSeq platform, as described in Patel et al., Blood;126(6):790-797(2015), the disclosure of which relating to DNA sample testing is incorporated herein.

[0075] C. Pharmacodynamics (PD) The present disclosure is based in part on pharmacodynamic effects demonstrating an association between response to telomerase inhibitor therapy in subjects with myelofibrosis and a decrease in telomerase hTERT expression levels in the subjects from baseline levels. In some cases, among subjects who achieve a clinical response (spleen or symptoms) to telomerase inhibitor therapy at 24 weeks, a higher proportion of subjects achieve a 50% or greater decrease in hTERT RNA expression levels than subjects who do not achieve a response.

[0076] The present disclosure provides for the stratification and selection of patients likely to benefit from telomerase inhibition therapy for myelofibrosis, and provides methods for monitoring response, relapse, and prognosis in treated subjects.

[0077] Aspects of the present disclosure include a method for selecting a subject with myelofibrosis (MF) for treatment with a telomerase inhibitor, and a method for treating MF. Also provided is a method for monitoring the effectiveness of treatment in a subject with MF. In some cases, the pharmacodynamic effect that the subject method embodiment is based on is a reduction of 50% or more, for example, 60% or more, 70% or more, 80% or more, or 90% or more of hTERT RNA expression.

[0078] The telomerase ribonucleoprotein consists of components or subunits, two of which are the telomerase RNA template (hTR) and the telomerase reverse transcriptase protein (hTERT). hTERT expression levels can be assessed, determined, and / or measured using any convenient method. Various methods can be applied to amplify, detect, and measure the mRNA of telomerase components or related proteins in body fluids. Methods and assays of interest that may be adapted for use in the subject methods include, but are not limited to, TaqMan fluorescence-based, real-time quantitative RT-PCR assays, immunohistochemistry for protein expression, and methods described in U.S. Pat. No. 6,607,898, Bieche et al., Clin. Cancer Res February 1 2000(6)(2)452-459, Terrin et al. (“Telomerase expression in B-cell chronic lymphocytic leukemia predicts survival and delineates subgroups of patients with the same igVH mutation status and different outcome.” Leukemia 2007;21:965-972), and Palma et al. (“Telomere length and expression of human telomerase reverse transcriptase splice variants in chronic lymphocytic leukemia.” Experimental Hematology 2013;41:615-626).

[0079] hTERT expression level can be evaluated or measured in any suitable target cell or biological sample.Target cell can be any suitable cell of patient, including but not limited to the bone marrow or peripheral blood cell of patient.In some cases, target cell is isolated from the bone marrow sample of patient.In some cases, target cell is isolated from the peripheral blood sample of patient.Target cell can be granulocyte.

[0080] The hTERT RNA expression level can be assessed or measured in an RNA sample using any convenient method. The RNA sample can be obtained by first obtaining a bone marrow sample, a peripheral blood sample, or both, and then isolating RNA from the bone marrow sample, the peripheral blood sample, or both. In one embodiment, obtaining a sample from a patient includes obtaining a bone marrow sample from the patient, isolating cells from the bone marrow sample, and extracting RNA and / or DNA from the isolated cells. In another embodiment, obtaining an RNA sample from a patient includes obtaining a peripheral blood sample from the patient, isolating cells from the peripheral blood sample (e.g., granulocytes), and extracting RNA and / or DNA from the isolated cells.

[0081] D. Treatment Embodiments of the present disclosure relate to a patient with a triple-negative status based on the absence of any mutations in the JAK2, CALR, and MPL genes (i.e., no mutations in these genes or these genes lacking mutations), and / or ... a patient with a triple-negative status based on the absence of any mutations in these genes), and / or a patient with a triple-negative status based on the absence of any mutations in the JAK2 The present invention includes a method for treating myelofibrosis in a subject (i.e., a patient) in need of treatment for myelofibrosis who has high molecular weight risk (HMR) based on the presence of a mutation in at least one of ASXL1, EZH2, SRSF2, and IDH1 / 2. One embodiment of the present invention is a method for treating myelofibrosis in a subject (i.e., a patient) in need of treatment for myelofibrosis who has triple-negative status based on the absence of any mutations in the JAK2, CALR, and MPL genes (i.e., no mutations or lack of mutations in these genes). In one embodiment, the myelofibrosis is primary myelofibrosis. In another embodiment, the myelofibrosis is myelofibrosis that develops after polycythemia vera (post-PV MF). In an alternative embodiment, the myelofibrosis is myelofibrosis that develops after essential thrombocythemia (post-ET MF).

[0082] In certain embodiments of the method of treatment, the patient has not previously received JAK inhibitor therapy. In other embodiments, the patient has previously received JAK inhibitor therapy and has "failed" JAK inhibitor therapy (i.e., the disease was resistant, or the patient was refractory to the therapy, or initially responded to treatment but the disease recurred). In alternative embodiments of the method of treatment, the patient has previously received JAK inhibitor therapy and discontinued JAK inhibitor therapy due to treatment-related toxicity or intolerance. In certain embodiments, the method of treatment further includes premedication with diphenhydramine (25-50 mg) and hydrocortisone (100-200 mg), or their equivalents.

[0083] The subject is a mammal in need of cancer treatment. Generally, the subject is a human patient. In some embodiments of the present invention, the subject may be a non-human mammal, such as a non-human primate, an animal model (e.g., an animal such as a rat used for drug screening, characterization, and evaluation), and other mammals. As used herein, the terms "patient," "subject," and "individual" are used interchangeably.

[0084] As used herein and as is well understood in the art, "treatment" refers to an approach to obtain beneficial or desired results, including clinical results. For the purposes of the present invention, beneficial or desired clinical results include, but are not limited to, the alleviation or amelioration of one or more symptoms, whether detectable or undetectable, a reduction in the extent of the disease, a stabilized (i.e., non-worsening) state of the disease, the prevention of the spread of the disease, the delay or slowing of the progression of the disease, the improvement or alleviation of the disease state, and remission (whether partial or total). "Treatment" can also mean prolonging survival compared to the expected survival if not receiving treatment.

[0085] E. Telomerase inhibitors The method of the present invention can be used to identify patients who are most likely to benefit from treatment with any suitable telomerase inhibitor.Furthermore, any suitable telomerase inhibitor can be used in a subject treatment method.In some embodiments, the telomerase inhibitor is an oligonucleotide having telomerase inhibitory activity, particularly the oligonucleotide defined in WO2005 / 023994 and / or WO2014 / 088785, the disclosures of which are incorporated herein by reference in their entirety.In some cases, one or more telomerase inhibitors (e.g., two or three telomerase inhibitors) can be administered to a mammal to treat hematological malignancies.

[0086] Imetelstat In certain embodiments, the telomerase inhibitor is imetelstat, including its tautomers and salts thereof, e.g., pharmaceutically acceptable salts. Imetelstat is a novel, first-in-class telomerase inhibitor with clinical activity in hematological malignancies (Baerlocher et al., NEJM 2015;373:920-928, Tefferi et al., NEJM 2015;373:908-919) (shown below): [ka] where "nps" represents a thiophosphoramidate bond, --NH--P(.dbd.O)(SH)--O--, linking the 3'-carbon of one nucleoside to the 5'-carbon of an adjacent nucleoside.

[0087] In certain cases, the telomerase inhibitor is imetelstat sodium, including its tautomers. Imetelstat sodium is the sodium salt of imetelstat, a synthetic lipid-conjugated 13-mer oligonucleotide N3'→P5'-thio-phosphoramidate. Imetelstat sodium is a telomerase inhibitor that is a covalently lipidated 13-mer oligonucleotide (shown below) complementary to the human telomerase RNA (hTR) template region. The chemical name for imetelstat sodium is DNA, d(3'-amino-3'-deoxy-P-thio)(TAGGGTTAGACAA), 5'-[O-[2-hydroxy-3-(hexadecanoylamino)propyl]phosphorothioate], sodium salt (1:13) (SEQ ID NO: 1). Imetelstat sodium does not function through an antisense mechanism and therefore does not have the side effects commonly observed with such therapies. [ka]

[0088] Unless otherwise indicated or clear from the context, a reference herein to imetelstat also includes its tautomers and salts thereof, e.g., pharmaceutically acceptable salts. As mentioned above, imetelstat sodium is, in particular, the sodium salt of imetelstat. Unless otherwise indicated or clear from the context, a reference herein to imetelstat sodium also includes all of its tautomers.

[0089] Imetelstat and imetelstat sodium can be manufactured, formulated, or obtained as described elsewhere (see, for example, Asai et al., Cancer Res., 63:3931-3939 (2003), Herbert et al., Oncogene, 24:5262-5268 (2005), and Gryaznov, Chem. Biodivers., 7:477-493 (2010)). Unless otherwise indicated or clear from the context, a reference to imetelstat herein also includes its salts. As noted above, imetelstat sodium is, specifically, the sodium salt of imetelstat.

[0090] Imetelstat targets the telomerase RNA template and inhibits telomerase activity and cell proliferation in various cancer cell lines and tumor xenografts in mice. Phase 1 studies in patients with breast cancer, non-small cell lung cancer, and other solid tumors, multiple myeloma, or chronic lymphocytic leukemia have provided information on the drug's pharmacokinetics and pharmacodynamics. A subsequent phase 2 study in patients with essential thrombocythemia demonstrated platelet-lowering activity associated with significant reductions in JAK2 V617F and CALR mutant allele burden. Imetelstat sodium is administered intravenously on a daily basis. It is contemplated that other administration routes, such as intrathecal administration, intratumoral injection, and oral administration, can also be used in the practice of the subject method. Imetelstat sodium can be administered at doses equivalent to those routinely used clinically. In certain embodiments, imetelstat sodium is administered as described elsewhere herein.

[0091] Certain embodiments are according to any one of the other embodiments, wherein the imetelstat is limited to imetelstat sodium.

[0092] F. Pharmaceutical Compositions For ease of administration, telomerase inhibitors (e.g., those described herein) can be formulated into various pharmaceutical forms for administration purposes. In some cases, the telomerase inhibitor is administered as a pharmaceutical composition. The carrier or diluent of a pharmaceutical composition must be "acceptable" in the sense of being compatible with the other ingredients of the composition and not harmful to the recipient. The pharmaceutical composition may be in a unitary dosage form suitable for administration, particularly oral, rectal, transdermal, parenteral injection, or inhalation. In some cases, administration can be via intravenous injection. For example, when preparing a composition in oral dosage form, any of the usual pharmaceutical media, such as water, glycols, oils, alcohols, etc., may be employed for oral liquid preparations such as suspensions, syrups, elixirs, emulsions, and solutions; or solid carriers, such as starches, sugars, kaolin, diluents, lubricants, binders, disintegrants, etc., may be employed for powders, pills, capsules, and tablets. Tablets and capsules are the most advantageous oral dosage unit forms due to their ease of administration; in these cases, solid pharmaceutical carriers are obviously employed. For parenteral compositions, the carrier typically comprises, at least in large part, sterile water, although other ingredients, for example, to aid solubility, may be included. For example, injectable solutions may be prepared in which the carrier comprises saline, glucose solution, or a mixture of saline and glucose solution. For example, injectable solutions may be prepared in which the carrier comprises saline, glucose solution, or a mixture of saline and glucose solution. Injectable solutions containing the telomerase inhibitors described herein may be formulated in oil for long-acting action. Suitable oils for this purpose include, for example, peanut oil, sesame oil, cottonseed oil, corn oil, soybean oil, synthetic glycerol esters of long-chain fatty acids, and mixtures thereof with other oils. Injectable suspensions may also be prepared, in which case appropriate liquid carriers, suspending agents, etc. may be employed. Also included are solid form preparations intended to be converted to liquid form preparations immediately prior to use.In compositions suitable for transdermal administration, the carrier optionally contains a penetration enhancer and / or a suitable wetting agent, optionally combined in small proportions with suitable additives of any nature, which do not cause significant adverse effects on the skin. Such additives may facilitate application to the skin and / or may be useful in formulating the desired composition. The composition may be administered in various ways, for example, as a transdermal patch, as a spot-on, or as an ointment.

[0093] It is particularly advantageous to formulate the aforementioned pharmaceutical compositions into unit dosage forms for ease of administration and uniformity of dosage.As used herein, unit dosage refers to a physically discrete unit suitable for single administration, each unit containing a predetermined amount of active ingredient calculated to produce a desired therapeutic effect in association with the necessary pharmaceutical carrier.Examples of such unit dosage forms are tablets (including scored tablets or coated tablets), capsules, pills, powder packets, wafers, suppositories, injection solutions or suspensions, etc., and separate multiple doses thereof.

[0094] To enhance the solubility and / or stability of the drugs described herein in the pharmaceutical composition, it may be advantageous to employ α-, β-, or γ-cyclodextrin or their derivatives, particularly hydroxyalkyl-substituted cyclodextrins, such as 2-hydroxypropyl-β-cyclodextrin or sulfobutyl-β-cyclodextrin. Also, cosolvents such as alcohols may improve the solubility and / or stability of the telomerase inhibitor in the pharmaceutical composition.

[0095] Depending on the mode of administration, the pharmaceutical composition preferably comprises 0.05 to 99% by weight, more preferably 0.1 to 70% by weight, even more preferably 0.1 to 50% by weight of a telomerase inhibitor described herein, and 1 to 99.95% by weight, more preferably 30 to 99.9% by weight, even more preferably 50 to 99.9% by weight of a pharmaceutically acceptable carrier, all percentages being based on the total weight of the composition.

[0096] G. Administration and Dosing Regimen The administration frequency can be any frequency that reduces the severity of myelofibrosis symptoms without causing significant toxicity to the subject. For example, the administration frequency can be about once every two months to about once per week, alternatively about once per month to about twice per month, alternatively about once per six weeks, about once per five weeks, alternatively about once per four weeks, alternatively about once per three weeks, alternatively about once per two weeks, or alternatively about once per week. The administration frequency can remain constant or can vary over the course of treatment. A course of treatment with a composition containing one or more telomerase inhibitors can include a rest period. For example, a composition containing a telomerase inhibitor can be administered weekly for three weeks, followed by a two-week rest period, and such a regimen can be repeated multiple times. As with the effective amount, various factors can affect the actual administration frequency used for a particular application. For example, the effective amount, duration of treatment, use of multiple therapeutic agents, route of administration, and the severity of myelofibrosis and associated symptoms may require increased or decreased frequency of administration.

[0097] The effective period for administering a composition containing a telomerase inhibitor (e.g., imetelstat or imetelstat sodium) can be any period that reduces the severity of myelofibrosis symptoms (e.g., as described herein) without causing significant toxicity to the subject. Thus, the effective period can vary from one month to several months or years (e.g., one month to two years, one month to one year, three months to two years, three months to ten months, or three months to 18 months). Generally, the effective period for treating myelofibrosis can range from two months to 20 months. In some cases, the effective period can last as long as the individual subject lives. Several factors can affect the actual effective period used for a particular treatment. For example, the effective period can vary depending on the frequency of administration, the effective amount, the use of multiple therapeutic agents, the route of administration, and the severity of myelofibrosis and associated symptoms.

[0098] In certain cases, the progress of treatment and the severity of one or more symptoms associated with myelofibrosis can be monitored.Any method can be used to determine whether the severity of myelofibrosis symptoms is reduced.For example, the severity of myelofibrosis symptoms (for example, as described herein) can be evaluated using biopsy technology.

[0099] The telomerase inhibitors used in the subject methods can be administered at any therapeutically effective dose, such as doses equivalent to those routinely used clinically. Specific dosage regimens (e.g., recommended effective doses) for known and approved anticancer drugs are known to physicians and are listed in the product descriptions found, for example, in PHYSICIANS' DESK REFERENCE, 2003, 57th Ed., Medical Economics Company, Inc., Oradell, NJ, Goodman & Gilman's "THE PHARMACOLOGICAL BASIS OF THERAPEUTICS" 2001, 10th Edition, McGraw-Hill, New York, and / or are available from the Federal Drug Administration and / or are discussed in medical literature.

[0100] In some embodiments, the dose of the telomerase inhibitor imetelstat sodium administered to a subject is about 1.0 mg / kg to about 13.0 mg / kg. In other embodiments, the dose of the telomerase inhibitor is about 4.5 mg / kg to about 11.7 mg / kg, or about 6.0 mg / kg to about 11.7 mg / kg, or about 6.5 mg / kg to about 11.7 mg / kg. In some embodiments, the dose of the telomerase inhibitor is at least about 4.5 mg / kg, 4.6 mg / kg, 4.7 mg / kg, 4.8 mg / kg, 4.9 mg / kg, 5.0 mg / kg, 5.5 mg / kg, 6.0 mg / kg, 6.1 mg / kg, 6.2 mg / kg, 6.3 mg / kg, 6.4 mg / kg, 6.5 mg / kg, 6.6 mg / kg, 6.7 mg / kg, 6.8 mg / kg, 6.9 mg / kg, 7 mg / kg, or 8 mg / kg. g, 7.1mg / kg, 7.2mg / kg, 7.3mg / kg, 7.4mg / kg, 7.5mg / kg, 7.6mg / kg, 7.7mg / kg, 7.8mg / kg, 7.9mg / kg, 8mg / kg, 8.1mg / kg, 8.2mg / kg, 8.3mg / kg, 8.4mg / kg, 8.5mg / kg, 8.6mg / kg, 8.7mg / kg, 8.8mg / kg, 8.9mg / kg, 9mg / kg, 9.1mg / kg, 9.2m g / kg, 9.3mg / kg, 9.4mg / kg, 9.5mg / kg, 9.6mg / kg, 9.7mg / kg, 9.8mg / kg, 9.9mg / kg, 10mg / kg, 10.1mg / kg, 10.2mg / kg , 10.3mg / kg, 10.4mg / kg, 10.5mg / kg, 10.6mg / kg, 10.7mg / kg, 10.8mg / kg, 10.9mg / kg, 11mg / kg, 11.1mg / kg, 11.2mg / kg, 11.3mg / kg, 11.4mg / kg, 11.5mg / kg, 11.6mg / kg, 11.7mg / kg, 11.8mg / kg, 11.9mg / kg, 12mg / kg, 12.1mg / kg, 12.2mg / kg, 12.3mg / kg, 12.4mg / kg, 12.5mg / kg, 12.6mg / kg, 12.7mg / kg, 12.8mg / kg, 12.9mg / kg, or 13mg / kg.

[0101] In some embodiments, the effective amount of the telomerase inhibitor administered to an individual comprises at least about any of 1 mg / kg, 2.5 mg / kg, 3.5 mg / kg, 4.7 mg / kg, 5 mg / kg, 5.5 mg / kg, 6.0 mg / kg, 6.5 mg / kg, 7.0 mg / kg, 7.5 mg / kg, 8.0 mg / kg, 8.5 mg / kg, 9.0 mg / kg, 9.4 mg / kg, 10 mg / kg, 15 mg / kg, or 20 mg / kg. In some embodiments, the effective amount of the telomerase inhibitor administered to an individual is at least about any of 1 mg / kg, 2.5 mg / kg, 3.5 mg / kg, 4.7 mg / kg, 5 mg / kg, 6.5 mg / kg, 7.5 mg / kg, 9.4 mg / kg, 10 mg / kg, 15 mg / kg, or 20 mg / kg. In various embodiments, the effective amount of telomerase inhibitor administered to an individual comprises less than about any of 350 mg / kg, 300 mg / kg, 250 mg / kg, 200 mg / kg, 150 mg / kg, 100 mg / kg, 50 mg / kg, 30 mg / kg, 25 mg / kg, 20 mg / kg, 10 mg / kg, 7.5 mg / kg, 6.5 mg / kg, 5 mg / kg, 3.5 mg / kg, 2.5 mg / kg, 1 mg / kg, or 0.5 mg / kg of telomerase inhibitor.

[0102] Exemplary dosing frequencies for pharmaceutical compositions comprising a telomerase inhibitor include, but are not limited to, daily, every other day, twice a week, three times a week, weekly without a break, weekly, three weeks out of four, once every three weeks, once every two weeks, or weekly for two weeks out of three weeks. In some embodiments, the pharmaceutical composition is administered approximately once a week, once every two weeks, once every three weeks, once every four weeks, once every five weeks, once every six weeks, once every seven weeks, or once every eight weeks. In some embodiments, the composition is administered at least once, twice, three times, four times, five times, six times, or seven times a week (i.e., daily), or three times a day or twice a day. In some embodiments, the interval between each administration is less than about 6 months, 3 months, 1 month, 20 days, 15 days, 12 days, 10 days, 9 days, 8 days, 7 days, 6 days, 5 days, 4 days, 3 days, 2 days, or 1 day. In some embodiments, the interval between each administration is more than about 1 month, 2 months, 3 months, 4 months, 5 months, 6 months, 8 months, or 12 months. In some embodiments, there are no breaks in the dosing schedule. In some embodiments, the interval between each administration is about 1 week or less.

[0103] A telomerase inhibitor, such as imetelstat (e.g., imetelstat sodium), can be administered using any suitable method. For example, a telomerase inhibitor, such as imetelstat (e.g., imetelstat sodium), can be administered intravenously over a period of time (e.g., 1 hour, 2 hours, 3 hours, 4 hours, or 5 hours) once every four weeks. In some embodiments, imetelstat is administered intravenously at 7-10 mg / kg over approximately two hours once every week. In certain embodiments, imetelstat is administered intravenously at about 0.5-9.4 mg / kg over approximately two hours once every three weeks. In one embodiment, imetelstat is administered intravenously at 0.5-5 mg / kg over approximately two hours once every four weeks. In one embodiment, imetelstat is administered intravenously at about 2.5-10 mg / kg over approximately two hours once every three weeks. Alternatively, imetelstat is administered intravenously at about 0.5 to 9.4 mg / kg over about two hours once every four weeks.

[0104] In certain embodiments of the method, imetelstat is administered for 1, 2, 3, 4, 5, 6, 7, 8, or more than 8 administration cycles, each cycle comprising intravenously administering about 7-10 mg / kg of imetelstat once every 3 weeks, intravenously administering about 7-10 mg / kg of imetelstat once a week for 3 weeks, intravenously administering about 2.5-10 mg / kg of imetelstat once every 3 weeks, or intravenously administering about 0.5-9.4 mg / kg of imetelstat once every 3 weeks. In certain instances, each administration cycle comprises intravenously administering about 7-10 mg / kg of imetelstat once every 3 weeks. In some instances, each administration cycle comprises intravenously administering about 9.4 mg / kg of imetelstat once every 3 weeks.

[0105] In one embodiment of the present invention, imetelstat is administered intravenously at a dose of about 7-10 mg / kg of imetelstat once every three weeks after premedication with an antihistamine, a corticosteroid, or both. In another embodiment, imetelstat is administered intravenously at a dose of about 9.4 mg / kg, alternatively about 7.0 mg / kg to about 9.8 mg / kg of imetelstat once every three weeks after premedication with an antihistamine, a corticosteroid, or both.

[0106] In certain embodiments, imetelstat is administered at a dose of about 7.5 mg / kg, alternatively about 7.0 mg / kg to about 7.7 mg / kg, once every three weeks for at least three cycles, with the dose escalating thereafter. In certain embodiments, the dose of imetelstat is such that the ANC and platelet nadir are each about 1.5 x 10 9 / L~Approx. 75×10 9 The dose may be increased to about 9.4 mg / kg, or to about 8.8 mg / kg to about 9.6 mg / kg, provided that the blood pressure is not reduced by 100 mg / L and there is no Grade ≥ 3 non-hematologic toxicity.

[0107] It will be understood that cancer treatment sometimes involves multiple "rounds" or "cycles" of drug administration, with each cycle comprising one or more drug administrations according to a designated schedule (e.g., three consecutive days every three weeks; once a week, etc.). For example, anticancer drugs can be administered for one to eight cycles, or for a longer period. When a subject is administered two or more drugs (e.g., two drugs), each can be administered according to its own schedule (e.g., weekly, once every three weeks, etc.). It will be apparent that drug administration, even for drugs administered at different cycles, can be coordinated so that both drugs are administered at least some time on the same day, or alternatively, so that the drugs are administered at least some time on consecutive days.

[0108] In certain embodiments, imetelstat may be administered via a dose-reducing regimen. In one embodiment, a patient is initially administered about 9.4 mg / kg every 3 weeks, then the dose is changed to about 7.5 mg / kg every 3 weeks, and then the dose is changed to about 6.0 mg / kg every 3 weeks.

[0109] As is understood in the art, if toxicity is observed or for the convenience of the patient, treatment with a cancer therapeutic agent can be temporarily interrupted and then resumed without departing from the scope of the present invention.

[0110] An embodiment of the subject method involves identifying or selecting patients most likely to benefit from treatment based on the relative telomere length in the patient's target cells (e.g., as described herein). The target cells can be any convenient cell in the patient, including, but not limited to, cells from the patient's bone marrow or peripheral blood. In some cases, the target cells are isolated from the patient's bone marrow sample. In some cases, the target cells are isolated from the patient's peripheral blood sample. The target cells can be granulocytes. In some cases, the patient lacks mutations in the Janus kinase 2 (JAK2), calreticulin (CALR), and thrombopoietin receptor (MPL) genes, resulting in a specific short telomere length in the patient's target cells. As used herein, a short telomere length is one that is below the median or average telomere length compared to a suitable control, such as one or more known standards described herein. Thus, the subject methods can further include determining relative telomere length by analyzing the relative length of telomeric nucleic acid of target cells present in a biological sample from the individual, and selecting an individual who would benefit from treatment with a telomere enzyme inhibitor if the average relative telomere length among target cells present in the biological sample from the individual is determined to be below the 50th percentile of a range of relative telomere lengths determined from one or more known standards, e.g., below the 45th percentile, below the 40th percentile, below the 35th percentile, below the 30th percentile, below the 25th percentile, below the 20th percentile, or below the range of relative telomere lengths determined from one or more known standards.

[0111] In some examples of the method, one or more known standards are telomere length ranges established from multiple naturally occurring target cells (for example, as described herein) from multiple individuals diagnosed with the disease. In a specific example of the method, one or more known standards are characterized cell lines. By "characterized cell lines," it is meant that the relative telomere nucleic acid of cells in the cell line is known and relatively constant.

[0112] In some embodiments, the telomere length of cancer cells present in the biological sample is determined to be equal to or less than the median or mean telomere length. In some embodiments, the telomere length of cancer cells present in the biological sample is determined to be equal to or less than the 50th percentile, 40th percentile, 35th percentile, 30th percentile, 25th percentile, 20th percentile, 15th percentile, 10th percentile, or 5th percentile of a range of relative telomere lengths determined from one or more known standards.

[0113] Telomere length of target cells can be determined using any convenient assay, including, but not limited to, qPCR, telo-FISH, or Southern blot assays, such as those described by Bassett et al. in U.S. Patent No. 9,200,327. In one embodiment, telomere length can be determined by measuring the average length of terminal restriction fragments (TRFs). TRFs are defined as the length (usually the average length) of fragments resulting from complete digestion of genomic DNA with a restriction enzyme that does not cleave nucleic acids within the telomeric sequence. In some cases, DNA is digested with a restriction enzyme that frequently cleaves within genomic DNA but not within the telomeric sequence. In some cases, restriction enzymes have four-base recognition sequences (e.g., AluI, HinfI, RsaI, and Sau3A1) and are used alone or in combination. The resulting terminal restriction fragments contain both telomeric repeats and subtelomeric DNA. Subtelomeric DNA is a DNA sequence adjacent to the tandem repeats of telomere sequences, containing telomere repeat sequences interspersed with variable telomere-like sequences. The digested DNA is separated by electrophoresis and blotted onto a support such as a membrane. Fragments containing telomere sequences are detected by hybridizing a probe, i.e., a labeled repeat sequence, to the membrane. Visualization of telomere-containing fragments allows the average length of terminal restriction fragments to be calculated (Harley, C.B. et al. Nature. 345(6274):458-60(1990), incorporated herein by reference). TRF estimation by Southern blotting shows the distribution of telomere length within cells or tissues, and therefore indicates the median and average telomere length of all cells.

[0114] In another embodiment, telomere length can be measured by flow cytometry (Hultdin, M. et al., Nucleic Acids Res. 26:3651-3656 (1998); Rufer, N. et al., Nat. Biotechnol. 16:743-747 (1998), incorporated herein by reference). Flow cytometry is a variant of the FISH technique. When the starting material is tissue, a cell suspension is generally prepared by mechanical separation and / or treatment with protease. The cells are fixed with a fixative and hybridized with a telomere sequence-specific probe, preferably a PNA probe, labeled with a fluorescent label. After hybridization, the cells are washed and then analyzed by FACS. After appropriate subtraction of background fluorescence, the fluorescent signal is measured for cells in Go / G1. This technique is suitable for rapid estimation of telomere length for a large number of samples. Similar to TRF, telomere length is the average length of telomeres in a cell.

[0115] In other embodiments, the median or average telomere length from cells in a biological sample is determined via quantitative PCR (qPCR) or telomere fluorescence in situ hybridization (telo-FISH). In qPCR, a DNA-binding dye binds to all double-stranded DNA, emitting fluorescence. An increase in DNA product during the PCR reaction results in an increase in fluorescence intensity, which is measured at each cycle of the PCR reaction. This allows for quantification of DNA concentration. The relative concentration of DNA present during the exponential phase of the reaction is determined by plotting the fluorescence level against the PCR cycle number on a semi-logarithmic scale. A threshold for detecting fluorescence above background is determined. The cycle at which fluorescence from a sample exceeds the threshold is called the cycle threshold (Ct). Because the amount of DNA theoretically doubles with each cycle during the exponential phase, the relative amount of DNA can be calculated. The baseline is the early cycle of PCR, during which there is little change in the fluorescence signal.

[0116] In some embodiments, telomere length is determined using telo-FISH. In this method, cells are fixed and hybridized with probes conjugated to fluorescent labels, such as Cy-3, fluorescein, or rhodamine. The probes are oligonucleotides designed to specifically hybridize to telomere sequences. Generally, probes are 8 nucleotides or longer, e.g., 12-20 nucleotides or longer. In one embodiment, the probes are oligonucleotides containing naturally occurring nucleotides. In one embodiment, the probes are peptide nucleic acids, which have a higher Tm than similar naturally occurring sequences, allowing for more stringent hybridization conditions. Cells can be treated with agents such as colcemid to induce cell cycle arrest at metaphase and provide metaphase chromosomes for hybridization and analysis. In some embodiments, cellular DNA can also be stained with the fluorescent dye 4',6-diamidino-2-phenylindole (DAPI).

[0117] Digital images of intact metaphase chromosomes are acquired, and the fluorescence intensity of the probe hybridized to the telomere is quantified. This allows for the measurement of telomere length for individual chromosomes in addition to the average or median telomere length within cells, avoiding problems associated with the presence of subtelomeric DNA (Zjilmans, J. Met. al., Proc. Natl. Acad. Sci. USA 94:7423-7428 (1997); Blasco, M. A. et al., Cell 91:25-34 (1997), incorporated by reference). The intensity of the fluorescent signal correlates with telomere length, with brighter fluorescent signals indicating longer telomeres.

[0118] In certain embodiments, the present invention relates to a telomerase inhibitor for use in a method for treating myelofibrosis, the method comprising: To identify patients most likely to benefit from treatment with a telomerase inhibitor, said patients being selected from: (a) triple-negative status, based on the absence of any mutations in the JAK2, CALR, and MPL genes; (b) high molecular weight risk (HMR) based on the presence of a mutation in at least one of the following genes: ASXL1, EZH2, SRSF2, and IDH1 / 2; or (c) both, including inspecting The presence of (a), (b), or (c) indicates that the patient is most likely to benefit from treatment with a telomerase inhibitor and administering to the patient an effective amount of the telomerase inhibitor. In certain embodiments, the invention relates to a telomerase inhibitor for use in the method defined in any of the other embodiments.

[0119] Yet another embodiment of the present invention is a telomerase inhibitor for use in treating myelofibrosis, the use comprising: (a) screening patients to determine whether such patients have triple-negative status based on the absence of mutations in each of JAK2, CALR, and MPL, and / or are at high risk (HMR) based on the presence of a mutation in at least one of the following genes: ASXL1, EZH2, SRSF2, and IDH1 / 2; and (b) administering the telomerase inhibitor to the patient if such patients have triple-negative status based on the absence of mutations in each of JAK2, CALR, and MPL, and / or are at high risk (HMR) based on the presence of a mutation in at least one of the following genes: ASXL1, EZH2, SRSF2, and IDH1 / 2. In one embodiment, the use comprises screening patients for triple-negative status based on the absence of mutations in each of JAK2, CALR, and MPL. Yet another embodiment of the present invention is a telomerase inhibitor for use in the treatment of myelofibrosis, the use comprising (a) screening patients to determine whether such patients have triple negative status based on the absence of mutations in each of JAK2, CALR, and MPL, and (b) administering the telomerase inhibitor to the patient if such patient has triple negative status.

[0120] Yet another embodiment of the present invention is the use of a telomerase inhibitor for the treatment of myelofibrosis, comprising: (a) screening patients to determine whether such patients have triple-negative status based on the absence of mutations in each of JAK2, CALR, and MPL, and / or are at high risk (HMR) based on the presence of a mutation in at least one of the following genes: ASXL1, EZH2, SRSF2, and IDH1 / 2; and (b) administering a telomerase inhibitor to the patient if such patients have triple-negative status based on the absence of mutations in each of JAK2, CALR, and MPL, and / or are at high risk (HMR) based on the presence of a mutation in at least one of the following genes: ASXL1, EZH2, SRSF2, and IDH1 / 2. In one embodiment, the use comprises screening patients for triple-negative status based on the absence of mutations in each of JAK2, CALR, and MPL. Yet another embodiment of the present invention is the use of a telomerase inhibitor for the treatment of myelofibrosis, comprising: (a) screening a patient to determine whether such patient has triple-negative status based on the absence of mutations in each of JAK2, CALR, and MPL; and (b) administering a telomerase inhibitor to the patient if such patient has triple-negative status.

[0121] In certain embodiments of the present invention, triple-negative status can be determined based on the absence of mutations in each of the JAK2 gene having the nucleic acid sequence of SEQ ID NO: 2, the CALR gene having the nucleic acid sequence of SEQ ID NO: 3, and the MPL gene having the nucleic acid sequence of SEQ ID NO: 4. In other embodiments, triple-negative status can be determined based on the absence of mutations in each of SEQ ID NO: 2, CALR, and MPL. In alternative embodiments, triple-negative status can be determined based on the absence of mutations in each of JAK2, SEQ ID NO: 3, and MPL. In alternative embodiments, triple-negative status can be determined based on the absence of mutations in each of JAK2, CALR, and SEQ ID NO: 4.

[0122] In other embodiments of the present invention, high molecular weight risk (HMR) may be determined based on the presence of a mutation in at least one of the following genes: the ASXL1 gene having the nucleic acid sequence of SEQ ID NO: 5, the EZH2 gene having the nucleic acid sequence of SEQ ID NO: 6, the SRSF2 gene having the nucleic acid sequence of SEQ ID NO: 7, the IDH1 gene having the nucleic acid sequence of SEQ ID NO: 8, the IDH2 gene having the nucleic acid sequence of SEQ ID NO: 9, and combinations thereof.

[0123] In another embodiment of the present invention, telomerase activity and hTERT expression levels can be determined in a biological sample obtained from a patient to evaluate the pharmacodynamic effects and / or monitor patients being treated with telomerase inhibition. Telomerase activity can be measured using a TRAP (Telomeric Repeat Amplification Protocol) telomerase activity assay. hTERT expression levels can be determined by measuring hTERT RNA expression levels in cells in the biological sample using Northern blot, serial analysis of gene expression (SAGE), or other methods.

[0124] In certain embodiments, the present invention relates to a telomerase inhibitor for use in the treatment of myelofibrosis as defined in any of the other embodiments.

[0125] In certain embodiments, the present invention relates to the use of a telomerase inhibitor for the treatment of myelofibrosis as defined in any of the other embodiments.

[0126] Additional Embodiments Additional embodiments of the object are described in the following clauses:

[0127] Clause 1. A method for identifying patients most likely to benefit from treatment with a telomerase inhibitor, comprising: (a) testing patients for triple-negative status based on the absence of mutations in each of the Janus kinase 2 (JAK2), calreticulin (CALR), and thrombopoietin receptor (MPL) genes; and (b) selecting the patient if the patient has triple negative status based on the absence of mutations in each of the JAK2, CALR, and MPL genes, wherein the selected patient is most likely to benefit from treatment with a telomerase inhibitor.

[0128] Clause 2. A method for identifying patients most likely to benefit from treatment with a telomerase inhibitor, comprising: (c) Examining the patient for: i. Triple-negative status based on the absence of mutations in each of the JAK2, CALR, and MPL genes, and / or ii. Testing for high molecular weight risk (HMR) based on the presence of a mutation in at least one of the following genes: ASXL1, EZH2, SRSF2, and IDH1 / 2; (d) selecting a patient having: i. Triple-negative status based on the absence of mutations in each of the JAK2, CALR, and MPL genes, and / or ii. selecting patients with high molecular weight risk (HMR) based on the presence of a mutation in at least one of the following genes: ASXL1, EZH2, SRSF2, and IDH1 / 2; The selected patients are most likely to benefit from treatment with a telomerase inhibitor, method.

[0129] Clause 3. A method for identifying patients most likely to benefit from treatment with a telomerase inhibitor, comprising: (e) obtaining a DNA sample from the patient; (f) testing a DNA sample from the patient for triple-negative status based on the absence of mutations in each of the JAK2, CALR, and MPL genes; (g) selecting the patient if the patient has triple negative status based on the absence of a mutation in each of the JAK2, CALR, and MPL genes; The selected patients are most likely to benefit from treatment with a telomerase inhibitor, method.

[0130] Clause 4. A method for identifying patients most likely to benefit from treatment with a telomerase inhibitor, comprising: (h) obtaining a DNA sample from the patient; (i) a DNA sample from a patient, i. Triple-negative status based on the absence of mutations in each of the JAK2, CALR, and MPL genes, and / or ii. Testing for high molecular weight risk (HMR) based on the presence of a mutation in at least one of the following genes: ASXL1, EZH2, SRSF2, and IDH1 / 2; (j) The patient: i. Triple-negative status based on the absence of mutations in each of the JAK2, CALR, and MPL genes, and / or ii. selecting patients as having high molecular weight risk (HMR) based on the presence of a mutation in at least one of the following genes: ASXL1, EZH2, SRSF2, and IDH1 / 2; The selected patients are most likely to benefit from treatment with a telomerase inhibitor, method.

[0131] Clause 5. The use of a telomerase inhibitor in the treatment of a patient with myelofibrosis, wherein the patient is determined to have triple negative status, Use of telomerase inhibitors, where triple-negative status includes the absence of mutations in each of the Janus kinase 2 (JAK2), calreticulin (CALR), and thrombopoietin receptor (MPL) genes.

[0132] Clause 6. The use of a telomerase inhibitor in the treatment of a patient with myelofibrosis, wherein the patient is determined to have high molecular weight risk (HMR), Use of telomerase inhibitors, where having HMR includes the presence of a mutation in at least one gene selected from the group consisting of additional sex comb-like 1 (ASXL1), enhancer of zest homolog 2 (EZH2), serine and arginine-rich splicing factor 2 (SRSF2), and isocitrate dehydrogenase 1 / 2 (IDH1 / 2).

[0133] Clause 7. Use of a telomerase inhibitor in the treatment of a patient having myelofibrosis, wherein cells present in a biological sample from the patient have been determined to have a mean relative telomere length determined to be less than or equal to the 50th percentile of a range of relative telomere lengths determined from one or more known standards.

[0134] Clause 8. Use of a telomerase inhibitor in the manufacture of a medicament for the treatment of a patient with myelofibrosis, wherein the patient is determined to have triple-negative status, wherein triple-negative status comprises the absence of mutations in each of the Janus kinase 2 (JAK2), calreticulin (CALR), and thrombopoietin receptor (MPL) genes.

[0135] Clause 9. Use of a telomerase inhibitor in the manufacture of a medicament for the treatment of a patient with myelofibrosis, wherein the patient is determined to have high molecular weight risk (HMR), and wherein having HMR comprises the presence of a mutation in at least one gene selected from the group consisting of additional sex comb-like 1 (ASXL1), enhancer of zest homolog 2 (EZH2), serine- and arginine-rich splicing factor 2 (SRSF2), and isocitrate dehydrogenase 1 / 2 (IDH1 / 2).

[0136] Clause 10. Use of a telomerase inhibitor in the manufacture of a medicament for the treatment of a patient having myelofibrosis, wherein cells present in a biological sample from the patient have been determined to have a mean relative telomere length determined to be less than or equal to the 50th percentile of a range of relative telomere lengths determined from one or more known standards.

[0137] The following examples are offered by way of illustration and not by way of limitation. [Example]

[0138] Example 1: Imetelstat sodium is an effective treatment for patients with intermediate-2 (int-2) or high-risk myelofibrosis (MF) that has relapsed or is refractory to Janus kinase (JAK) inhibitor therapy. introduction Imetelstat, a 13-mer oligonucleotide that specifically targets the RNA template of human telomerase, is a potent competitive inhibitor of telomerase enzyme activity (Asai et al. Cancer Res 2003; Herbert, Oncogene 2005). Clinical activity and an acceptable safety profile were reported in a pilot study of 33 patients with intermediate-grade 2 (int-2) or high-risk myelofibrosis (MF), 48% of whom had previously been treated with a Janus kinase inhibitor (JAKi) (Tefferi, N Engl J Med 2015). This example provides results from a phase 2 clinical trial of imetelstat sodium at two dose levels in patients with MF.

[0139] method A randomized, multicenter, phase 2 trial of two doses of imetelstat sodium (9.4 mg / kg or 4.7 mg / kg IV once every 3 weeks) was conducted in adults with a Dynamic International Prognostic Scoring System (DIPSS) score of int-2 or high-risk MF relapsed / refractory to prior JAKi therapy (i.e., either no reduction in splenomegaly after 12 weeks or worsening of splenomegaly at any time after initiation of JAK inhibitor ("JAKi") therapy). A diagnosis of primary, post-essential thrombocythemia, or post-polycythemia vera MF was required. Other eligibility criteria included measurable splenomegaly (by magnetic resonance imaging [MRI]), active MF-related systemic symptoms, and a platelet count ≥ 75 × 10 9 / L. Primary endpoints were splenic response rate (% achieving ≥35% spleen volume reduction [SVR] by MRI at week 24) and symptom response rate (% achieving ≥50% reduction in total symptom score [TSS] according to Myelofibrosis Symptom Assessment Form (MFSAF) v2 at week 24). Secondary endpoints included safety, overall survival (OS), treatment response, molecular response, and pharmacokinetic and pharmacodynamic relationships.

[0140] result 107 patients were enrolled at 55 centers (48 on 4.7 mg / kg and 59 on 9.4 mg / kg). Baseline characteristics are shown in Table 1 below. Additionally, median time on JAKi was 23 months (range 0.9-89.7), and median platelet count was 147 x 10 9 Triple-negative (TN, i.e., no JAK2, MPL, or CALR mutations) accounted for 24.8% of patients, and 67.6% were considered high-molecular-risk (HMR, i.e., ≥1 ASXL1, EZH2, SRSF2, or IDH1 / 2 mutation). [Table 1]

[0141] At the time of the primary clinical cutoff, the median time on study was 22.6 months (range, 0.2-27.4 months) and the median time on treatment was 6.2 months (range, 0.0-27.2 months). Six (10.2%) patients in the 9.4 mg / kg group had a splenic response per MRI confirmed by IRC.

[0142] At the time of clinical cutoff, patients were followed for a median of 22.6 months (range, 0.2-27.4 months), with a median treatment duration of 6.2 months (range, 0.0-27.2 months). The median treatment duration was longer in the 9.4 mg / kg group (7.7 months) than in the 4.7 mg / kg group. Six patients (10.2%) in the 9.4 mg / kg group had a splenic response per MRI, compared with none in the 4.7 mg / kg group (see Figure 1). Nineteen patients (32%) in the 9.4 mg / kg group and three patients (6%) in the 4.7 mg / kg group had a symptomatic response (TSS reduction ≥ 50%) (see Figure 2).

[0143] At the first clinical cutoff, median OS was not reached in the 9.4 mg / kg group, whereas median OS was 19.9 months in the 4.7 mg / kg group. The 18-month survival rates were 76.7% and 62.9% in the 9.4 mg / kg and 4.7 mg / kg groups, respectively. In a sensitivity analysis, censoring patients at the time of subsequent dose escalation for JAKi therapy or stem cell transplant yielded similar results. In the 9.4 mg / kg group, an association was observed between TN and OS (median OS was not reached in TN patients and 23.6 months in non-TN patients). The splenic response rate was higher in patients with one HMR mutation (ASXL1, EZH2, SRSF2, or IDH1 / 2).

[0144] The most common adverse events (all grades) with 9.4 mg / kg treatment were thrombocytopenia (49%), anemia (44%), neutropenia (36%), and nausea (34%), compared with diarrhea (38%), nausea (31%), anemia (31%), and thrombocytopenia (23%) at 4.7 mg / kg. Grade 3 / 4 neutropenia and thrombocytopenia were more frequent at 9.4 mg / kg (34% and 42%, respectively) than at 4.7 mg / kg (13% and 29%, respectively), with most cytopenias resolving within 4 weeks. Grade 3 / 4 LFT elevations were observed in seven patients on study. No imetelstat-related hepatotoxicity was observed as confirmed by an independent Hepatic Review Committee.

[0145] At the time of the second clinical cutoff, patients were followed up for 27.4 months (range, 0.2-33.0 months), with a median treatment duration of 26.9 weeks (range, 0.1-118.1 weeks). The median treatment duration was longer in the 9.4 mg / kg group (33.3 weeks) than in the 4.7 mg / kg group (23.9 weeks). Early closure of the 4.7 mg / kg group affected treatment duration. The median OS with 95% confidence interval (CI) in the 9.4 mg / kg group was 29.9 months (22.8, NE) (NE is not estimable) in the 9.4 mg / kg group, reaching the second clinical cutoff.

[0146] Triple negative vs OS Subjects were grouped by JAK2 / MPL / CALR gene mutation status: triple-negative (TN, lacking mutations in either of the JAK2 / MPL / CALR genes) and non-TN (having mutations in either of the JAK2 / MPL / CALR genes). In the 9.4 mg / kg group, median OS was not estimable (NE) for TN subjects with a 95% confidence interval (23.2, NE) and 23.6 months for non-TN subjects with a 95% confidence interval (20.7, NE). In the 4.7 mg / kg group, median OS with a 95% confidence interval (95% confidence interval) was 22.3 (17, NE) and 20.3 (18.3, NE) for TN and non-TN subjects, respectively. In the 9.4 mg / kg group, lower mortality was observed in the triple-negative (TN) group compared with the non-TN group (see Table 2, Figure 3, and Figure 4). [Table 2]

[0147] At the second clinical cutoff, lower mortality was seen in the triple negative (TN) group at 9.4 mg / kg compared to the non-TN group (see Table 3, Figures 6 and 7). [Table 3]

[0148] Triple negative vs. 24-week response In the 9.4 mg / kg group, a higher response rate (SVR or TSS) was seen in the TN group compared to the non-TN group (see Table 4 below). [Table 4]

[0149] Molecular risk versus response at 24 weeks In the 9.4 mg / kg group, higher response rates (SVR or TSS) were observed in the low molecular risk (LMR) group or high molecular risk (HMR) group with only one mutation (mut) compared to the HMR group with more than one mutation (see Table 5). [Table 5]

[0150] For 9.4 mg / kg subjects, an association between the following factors and clinical response or OS was observed: Triple-negative (TN): Responses (SVR or TSS) were robust in TN subjects. Median OS not estimable for TN, non-TN = 23.6 months, and Molecular Risk: Response (SVR or TSS) was enhanced in subjects with HMR with only one mutation, and responses were observed in patients with HMR with more than one mutation treated with 9.4 mg / kg imetelstat.

[0151] Example 2 Baseline telomere length (TL) versus overall survival (OS) Subjects are grouped by median baseline TL. In the 9.4 mg / kg group, median OS was not estimable (NE) at the first clinical cutoff and was (23.2, NE) and 22.8 (16.2, NE) months with 95% confidence intervals for subjects with shorter baseline TL (<=median) and longer TL (>median), respectively (Table 6). In the 4.7 mg / kg group, median OS with 95% confidence intervals was 20.3 (17.2, NE) and 22.3 (16.6, NE) months for subjects with shorter and longer baseline TL, respectively (Table 6).

[0152] In the 9.4 mg / kg group, a trend toward better OS was observed for subjects with shorter baseline TL, i.e., subjects with a baseline TL equal to or less than the median TL. [Table 6]

[0153] Baseline TL vs. response at week 24 Baseline telomere length (TL): SVR or TSS response at week 24 was enhanced in subjects with shorter baseline TL (<=median). 17.3% (5 / 29) of subjects with shorter baseline TL and 4.2% (1 / 24) of subjects with longer baseline TL had a spleen response, respectively. 34.5% (10 / 29) of subjects with shorter baseline TL and 25% (6 / 24) of subjects with longer baseline TL had a TSS response, respectively.

[0154] In the 9.4 mg / kg group, subjects with shorter baseline TL had enhanced higher response rates (SVR or TSS) at week 24 compared with subjects with longer TL (Table 7). [Table 7]

[0155] Example 3 Dose-dependent pharmacodynamic (PD) effects Telomerase activity and hTERT were analyzed to evaluate the pharmacodynamic effects of imetelstat. Among subjects with available baseline and post-treatment data, 23 subjects (51.1%) in the 9.4 mg / kg group and 10 subjects (29.4%) in the 4.7 mg / kg group achieved a telomerase activity reduction of >=50% from baseline, a PD effect that correlated with antitumor activity in vivo in preclinical xenograft models. In addition, 35 subjects (61.4%) in the 9.4 mg / kg group and 20 subjects (47.7%) in the 4.7 mg / kg group, respectively, achieved a PD effect of >=50% from baseline in hTERT RNA levels (Table 8). Thus, a dose-dependent PD effect was demonstrated, indicating target engagement. [Table 8]

[0156] Example 4 Association between PD efficacy and response at 24 weeks A higher proportion of subjects who were splenic responders (83.3%) achieved at least a 50% decrease in hTERT RNA expression levels than subjects who were non-splenic responders (55.6%), and a higher proportion of subjects with TSS responses achieved at least a 50% decrease in hTERT RNA expression levels than non-TSS responders (Table 9). hTERT RNA expression levels were measured from whole blood samples collected from patients before and after treatment. [Table 9]

[0157] A higher percentage of subjects who had a spleen or TSS response achieved at least a >=30% or >=50% decrease in telomerase activity than subjects who did not have a spleen or TSS response (Table 10). [Table 10]

[0158] Aspects, including embodiments, of the subject matter described herein may be useful alone or in combination with one or more other aspects or embodiments. Without limiting the description, certain non-limiting aspects of the present disclosure are provided below. As will be apparent to those skilled in the art upon reading this disclosure, each individually numbered aspect can be used or combined with any of the preceding or subsequent individually numbered aspects. This is intended to provide support for all such combinations of aspects, and is not limited to the combinations of aspects explicitly provided below. 1. The use of a telomerase inhibitor in the treatment of a patient with myelofibrosis, wherein the patient is determined to have triple-negative status; Use where triple-negative status comprises the absence of mutations in each of the Janus kinase 2 (JAK2), calreticulin (CALR), and thrombopoietin receptor (MPL) genes. 2. The use according to aspect 1, wherein the myelofibrosis is primary myelofibrosis. 3. The use according to aspect 2, wherein the myelofibrosis is myelofibrosis that develops after polycythemia vera (post-PV MF). 4. The use according to aspect 2, wherein the myelofibrosis is myelofibrosis that develops after essential thrombocythemia (post-ET MF). 5. The use of any one of aspects 1-4, wherein the patient has not previously received JAK inhibitor therapy. 6. The use of any one of aspects 1 to 4, wherein the patient has received JAK inhibitor therapy and the patient was resistant to JAK inhibitor therapy. 7. The use of any one of aspects 1 to 4, wherein the patient has received JAK inhibitor therapy and has relapsed. 8. The use of any one of aspects 1-4, wherein the patient has received JAK inhibitor therapy and discontinued the JAK inhibitor therapy due to treatment-related toxicity or intolerance. 9. The use according to any one of aspects 1 to 8, wherein the telomerase inhibitor is imetelstat. 10. The use of aspect 9, wherein the imetelstat is imetelstat sodium. 11. The telomerase inhibitor is imetelstat and is administered for 1, 2, 3, 4, 5, 6, 7, 8, or more than 8 administration cycles, each cycle comprising: Approximately 7 to 10 mg / kg of imetelstat is administered intravenously once every 3 weeks. Approximately 7 to 10 mg / kg of imetelstat will be administered intravenously once a week for 3 weeks. Approximately 2.5 to 10 mg / kg of imetelstat administered intravenously once every 3 weeks, or Approximately 0.5 to 9.4 mg / kg of imetelstat is administered intravenously once every 3 weeks. 11. The use according to embodiment 10, comprising 12. The use of aspect 11, wherein each administration cycle comprises intravenously administering about 7-10 mg / kg of imetelstat once every three weeks. 13. The use of aspect 12, wherein each administration cycle comprises intravenously administering about 9.4 mg / kg of imetelstat once every three weeks. 14. The use according to any one of aspects 1 to 13, wherein the average relative telomere length is determined by analyzing the relative length of telomeric nucleic acid in target cells present in a patient-derived biological sample. 15. The use of any one of aspects 1-14, further comprising selecting a patient identified as having an average relative telomere length in target cells present in a biological sample from the patient that is determined to be less than or equal to the 50th percentile of a range of relative telomere lengths determined from one or more known standards. 16. The use of any one of aspects 1 to 15, further comprising screening the patient to determine whether the patient has high molecular weight risk (HMR), wherein having HMR comprises the presence of a mutation in at least one gene selected from the group consisting of ASXL1, EZH2, SRSF2, and IDH1 / 2. 17. The use according to any one of aspects 1 to 16, further comprising assessing the level of hTERT expression in a biological sample obtained from the patient after administration of the telomerase inhibitor. 18. The use according to aspect 17, wherein the hTERT expression level is reduced by 50% or more compared to the baseline hTERT expression level before administration of the telomerase inhibitor. 19. The use of aspect 17 or 18, further comprising altering the dosage, frequency of administration, or course of treatment of the telomerase inhibitor administered to the subject. 20. Use of a telomerase inhibitor in the treatment of a patient with myelofibrosis, wherein the patient is determined to have high molecular weight risk (HMR), Use wherein having HMR includes the presence of a mutation in at least one gene selected from the group consisting of additional sex comb-like 1 (ASXL1), enhancer of zest homolog 2 (EZH2), serine and arginine-rich splicing factor 2 (SRSF2), and isocitrate dehydrogenase 1 / 2 (IDH1 / 2). 21. The use according to aspect 20, wherein the myelofibrosis is primary myelofibrosis. 22. The use according to aspect 21, wherein the myelofibrosis is myelofibrosis that develops after polycythemia vera (post-PV MF). 23. The use according to aspect 21, wherein the myelofibrosis is myelofibrosis that develops after essential thrombocythemia (post-ET MF). 24. The use of any one of aspects 20-23, wherein the patient has not previously received JAK inhibitor therapy. 25. The use of any one of aspects 20-23, wherein the patient has previously received JAK inhibitor therapy and the patient was resistant to JAK inhibitor therapy. 26. The use of any one of aspects 20-23, wherein the patient has received JAK inhibitor therapy and has relapsed. 27. The use of any one of aspects 20-23, wherein the patient has received JAK inhibitor therapy and discontinued the JAK inhibitor therapy due to treatment-related toxicity or intolerance. 28. The use according to any one of aspects 20 to 27, wherein the telomerase inhibitor is imetelstat. 29. The use according to aspect 28, wherein the imetelstat is imetelstat sodium. 30. The telomerase inhibitor is imetelstat and is administered for 1, 2, 3, 4, 5, 6, 7, 8, or more than 8 dosing cycles, each cycle comprising: Approximately 7 to 10 mg / kg of imetelstat is administered intravenously once every 3 weeks. Approximately 7 to 10 mg / kg of imetelstat will be administered intravenously once a week for 3 weeks. Approximately 2.5 to 10 mg / kg of imetelstat administered intravenously once every 3 weeks, or 29. The use of embodiment 28, comprising intravenously administering about 0.5 to 9.4 mg / kg of imetelstat once every three weeks. 31. The use of aspect 30, wherein each administration cycle comprises intravenously administering about 7-10 mg / kg of imetelstat once every three weeks. 32. The use of aspect 31, wherein each administration cycle comprises intravenously administering about 9.4 mg / kg of imetelstat once every three weeks. 33. The use according to any one of aspects 20 to 32, further comprising determining the average relative telomere length by analyzing the relative length of telomeric nucleic acid in target cells present in a biological sample from a patient. 34. The use of any one of aspects 20-33, further comprising selecting a patient identified as having an average relative telomere length in target cells present in a patient-derived biological sample that is determined to be less than or equal to the 50th percentile of a range of relative telomere lengths as determined from one or more known standards. 35. The use of any one of aspects 20 to 34, further comprising screening the patient to determine whether the patient has triple-negative status, wherein triple-negative status comprises the absence of a mutation in each of the genes selected from the group consisting of JAK2, CALR, and MPL. 36. The use according to any one of aspects 20 to 35, further comprising assessing the level of hTERT expression in a biological sample obtained from the patient after administration of a telomerase inhibitor. 37. The use according to aspect 36, wherein the hTERT expression level is reduced by 50% or more compared to the baseline hTERT expression level before administration of the telomerase inhibitor. 38. The use of any one of aspects 36-37, further comprising altering the dosage, frequency of administration, or course of treatment of the telomerase inhibitor administered to the subject. 39. Use of a telomerase inhibitor in treating a patient with myelofibrosis, wherein cells present in a biological sample from the patient have been determined to have a mean relative telomere length determined to be less than or equal to the 50th percentile of a range of relative telomere lengths determined from one or more known standards. 40. The use according to aspect 39, wherein the myelofibrosis is primary myelofibrosis. 41. The use according to aspect 40, wherein the myelofibrosis is myelofibrosis that develops after polycythemia vera (post-PV MF). 42. The use according to aspect 40, wherein the myelofibrosis is myelofibrosis that develops after essential thrombocythemia (post-ET MF). 43. The use of any one of aspects 39-42, wherein the patient has not previously received JAK inhibitor therapy. 44. The use of any one of aspects 39-42, wherein the patient has received JAK inhibitor therapy and the patient was resistant to JAK inhibitor therapy. 45. The use of any one of aspects 39-42, wherein the patient has received JAK inhibitor therapy and has relapsed. 46. ​​The use of any one of aspects 39-42, wherein the patient has received JAK inhibitor therapy and discontinued the JAK inhibitor therapy due to treatment-related toxicity or intolerance. 47. The use of any one of aspects 39 to 46, wherein the telomerase inhibitor is imetelstat. 48. The use according to aspect 47, wherein the imetelstat is imetelstat sodium. 49. The telomerase inhibitor is imetelstat and is administered for 1, 2, 3, 4, 5, 6, 7, 8, or more than 8 dosing cycles, each cycle comprising: Approximately 7 to 10 mg / kg of imetelstat is administered intravenously once every 3 weeks. Approximately 7 to 10 mg / kg of imetelstat will be administered intravenously once a week for 3 weeks. Approximately 2.5 to 10 mg / kg of imetelstat administered intravenously once every 3 weeks, or 48. The use of embodiment 47, comprising intravenously administering about 0.5 to 9.4 mg / kg of imetelstat once every three weeks. 50. The use of aspect 49, wherein each administration cycle comprises intravenously administering about 7-10 mg / kg of imetelstat once every three weeks. 51. The use of aspect 50, wherein each administration cycle comprises intravenously administering about 9.4 mg / kg of imetelstat once every three weeks. 52. The use according to any one of aspects 39 to 51, further comprising determining the average relative telomere length by analyzing the relative length of telomeric nucleic acid in cells present in the patient-derived biological sample. 53. The use according to any one of aspects 39 to 52, further comprising assessing the level of hTERT expression in a biological sample obtained from the patient after administration of a telomerase inhibitor. 54. The use according to aspect 53, wherein the hTERT expression level is reduced by 50% or more compared to the baseline hTERT expression level before administration of the telomerase inhibitor. 55. The use of any one of aspects 53-54, further comprising altering the dosage, frequency of administration, or course of treatment of the telomerase inhibitor administered to the subject. 56. A method for selecting patients most likely to benefit from treatment with a telomerase inhibitor, comprising: testing the patient for triple-negative status, wherein triple-negative status comprises the absence of mutations in each of the JAK2, CALR, and MPL genes; selecting the patient if the patient has triple negative status; The selected patients are most likely to benefit from treatment with a telomerase inhibitor, method. 57. The method of embodiment 56, wherein the patient has myelofibrosis. 58. The method of aspect 57, wherein the myelofibrosis is primary myelofibrosis. 59. The method of aspect 57, wherein the myelofibrosis is myelofibrosis that develops after polycythemia vera (post-PV MF). 60. The method of aspect 57, wherein the myelofibrosis is myelofibrosis that develops after essential thrombocythemia (post-ET MF). 61. The method of any of aspects 56-60, wherein the patient has not previously received JAK inhibitor therapy. 62. The patient: have received previous JAK inhibitor therapy, have previously received JAK inhibitor therapy and have failed JAK inhibitor therapy; or The method of any one of aspects 56-60, wherein the patient has previously received JAK inhibitor therapy and discontinued the JAK inhibitor therapy due to treatment-related toxicity or intolerance. 63. The method of any one of aspects 56-60, wherein the patient has received JAK inhibitor therapy, and the patient was resistant to JAK inhibitor therapy. 64. The method of any one of aspects 56-60, wherein the patient has received JAK inhibitor therapy and has relapsed. 65. The method of any one of aspects 56-60, wherein the patient has received JAK inhibitor therapy and discontinued the JAK inhibitor therapy due to treatment-related toxicity or intolerance. 66. The method of any one of aspects 56-65, further comprising administering to the patient a telomerase inhibitor. 67. The method of embodiment 66, wherein the telomerase inhibitor is imetelstat. 68. The method of embodiment 67, wherein the imetelstat is imetelstat sodium. 69. The method of any one of aspects 56-68, further comprising obtaining a sample comprising DNA from the patient. 70. The method of embodiment 69, wherein the sample comprises bone marrow, peripheral blood, or a combination thereof. 71. The step of obtaining a sample from a patient comprises: Obtaining a bone marrow sample, a peripheral blood sample, or a combination thereof; and isolating DNA from the bone marrow sample, the peripheral blood sample, or a combination thereof. 72. The step of obtaining a sample from a patient comprises: obtaining a bone marrow sample from a patient; isolating cells from the bone marrow sample; extracting DNA from the isolated cells. 73. The step of obtaining a sample from a patient comprises: Obtaining a peripheral blood sample from a patient; Isolating cells from a peripheral blood sample; extracting DNA from the isolated cells. 74. A method for selecting patients most likely to benefit from treatment with a telomerase inhibitor, comprising: testing the patient to determine whether the patient has HMR, where having HMR comprises the presence of a mutation in at least one gene selected from the group consisting of ASXL1, EZH2, SRSF2, and IDH1 / 2; selecting the patient if the patient has HMR; The selected patients are most likely to benefit from treatment with a telomerase inhibitor, method. 75. The method of embodiment 74, wherein the patient has myelofibrosis. 76. The method of aspect 75, wherein the myelofibrosis is primary myelofibrosis. 77. The method of aspect 75, wherein the myelofibrosis is myelofibrosis that develops after polycythemia vera (post-PV MF). 78. The method of aspect 75, wherein the myelofibrosis is myelofibrosis that develops after essential thrombocythemia (post-ET MF). 79. The method of any of aspects 74-78, wherein the patient has not previously received JAK inhibitor therapy. 80. The patient: have received previous JAK inhibitor therapy, have previously received JAK inhibitor therapy and have failed JAK inhibitor therapy, or 79. The method of any one of aspects 74-78, wherein the patient has previously received JAK inhibitor therapy and discontinued the JAK inhibitor therapy due to treatment-related toxicity or intolerance. 81. The method of any one of aspects 74-78, wherein the patient has received JAK inhibitor therapy, and the patient was resistant to JAK inhibitor therapy. 82. The method of any one of aspects 74-78, wherein the patient has received JAK inhibitor therapy and has relapsed. 83. The method of any one of aspects 74-78, wherein the patient has received JAK inhibitor therapy and discontinued the JAK inhibitor therapy due to treatment-related toxicity or intolerance. 84. The method of any one of aspects 74-83, further comprising administering to the patient a telomerase inhibitor. 85. The method of embodiment 84, wherein the telomerase inhibitor is imetelstat. 86. The method of embodiment 85, wherein the imetelstat is imetelstat sodium. 87. The method of any one of aspects 74-86, further comprising obtaining a sample comprising DNA from the patient. 88. The method of embodiment 87, wherein the sample comprises bone marrow, peripheral blood, or a combination thereof. 89. The step of obtaining a sample from a patient comprises: Obtaining a bone marrow sample, a peripheral blood sample, or a combination thereof; and isolating DNA from the bone marrow sample, the peripheral blood sample, or a combination thereof. 90. The step of obtaining a sample from a patient comprises: obtaining a bone marrow sample from a patient; isolating cells from the bone marrow sample; and extracting DNA from the isolated cells. 91. The step of obtaining a sample from a patient comprises: Obtaining a peripheral blood sample from a patient; Isolating cells from a peripheral blood sample; and extracting DNA from the isolated cells. 92. A method for selecting patients most likely to benefit from treatment with a telomerase inhibitor, comprising: testing the patient for average relative telomere length by analyzing the relative length of telomeric nucleic acid of target cells present in a biological sample from the patient; selecting a patient if the patient has a mean relative telomere length of target cells present in a biological sample from the patient that is determined to be less than or equal to the 50th percentile of a range of relative telomere lengths determined from one or more known standards; The selected patients are most likely to benefit from treatment with a telomerase inhibitor, method. 93. The method of embodiment 92, wherein the patient has myelofibrosis. 94. The method of aspect 93, wherein the myelofibrosis is primary myelofibrosis. 95. The method of aspect 93, wherein the myelofibrosis is myelofibrosis that develops after polycythemia vera (post-PV MF). 96. The method of aspect 93, wherein the myelofibrosis is myelofibrosis that develops after essential thrombocythemia (post-ET MF). 97. The method of any of aspects 92-96, wherein the patient has not previously received JAK inhibitor therapy. 98. The patient: have received previous JAK inhibitor therapy, have previously received JAK inhibitor therapy and have failed JAK inhibitor therapy; or 97. The method of any one of aspects 92-96, wherein the patient has previously received JAK inhibitor therapy and discontinued the JAK inhibitor therapy due to treatment-related toxicity or intolerance. 99. The method of any one of aspects 92-96, wherein the patient has previously received JAK inhibitor therapy, and the patient was resistant to JAK inhibitor therapy. 100. The method of any one of aspects 92-96, wherein the patient has received JAK inhibitor therapy and has relapsed. 101. The method of any one of aspects 92-96, wherein the patient has received JAK inhibitor therapy and discontinued the JAK inhibitor therapy due to treatment-related toxicity or intolerance. 102. The method of any one of aspects 92-101, further comprising administering to the patient a telomerase inhibitor. 103. The method of embodiment 102, wherein the telomerase inhibitor is imetelstat. 104. The method of aspect 103, wherein the imetelstat is imetelstat sodium. 105. The method of any one of aspects 92-104, further comprising obtaining a sample comprising DNA from the patient. 106. The method of embodiment 105, wherein the sample comprises bone marrow, peripheral blood, or a combination thereof. 107. The step of obtaining a sample from a patient comprises: Obtaining a bone marrow sample, a peripheral blood sample, or a combination thereof; and isolating DNA from a bone marrow sample, a peripheral blood sample, or a combination thereof. 108. The step of obtaining a sample from a patient comprises: obtaining a bone marrow sample from a patient; isolating cells from the bone marrow sample; extracting DNA from the isolated cells. 109. The step of obtaining a sample from a patient comprises: Obtaining a peripheral blood sample from a patient; Isolating cells from a peripheral blood sample; extracting DNA from the isolated cells. 110. A method for monitoring therapeutic efficacy in a subject with myelofibrosis (MF), the method comprising: measuring the level of hTERT expression in a biological sample obtained from the patient after administration of a telomerase inhibitor; comparing the hTERT expression level in the biological sample to a baseline hTERT expression level before administration of the telomerase inhibitor; A method wherein a 50% or greater decrease in hTERT expression levels in the biological sample identifies a subject likely to benefit from treatment with a telomerase inhibitor. 111. The method of embodiment 110, wherein the hTERT expression level measured or assessed is the hTERT RNA expression level. 112. A method for identifying a patient with myelofibrosis (MF) for treatment with a telomerase inhibitor, the method comprising: measuring the level of hTERT expression in a biological sample obtained from the patient after administration of a telomerase inhibitor; comparing the hTERT expression level in the biological sample to a baseline hTERT expression level before administration of the telomerase inhibitor; A method in which a decrease in hTERT expression levels in a biological sample identifies patients who are likely to benefit from treatment with a telomerase inhibitor. 113. The method of embodiment 112, wherein the decrease in hTERT expression level is 50% or more.

[0159] Although particular embodiments have been described in some detail by way of illustration and example for purposes of clarity of understanding, it will be readily apparent that certain changes and modifications can be made in light of the teachings of the present invention without departing from the spirit or scope of the appended claims.

[0160] Thus, the foregoing merely illustrates the principles of the present invention. Various arrangements may be devised that embody the principles of the present invention and are within its spirit and scope, although not explicitly described or illustrated herein. Furthermore, all examples and conditional language recited herein are intended primarily to aid the reader in understanding the principles of the present invention and the concepts the inventors have contributed to furthering the art, and should not be construed as being limited to such specifically recited examples and conditions. Furthermore, all statements herein reciting principles, aspects, and embodiments of the present invention, as well as specific examples thereof, are intended to encompass both structural and functional equivalents thereof. Additionally, such equivalents are intended to include both currently known equivalents and future-developed equivalents, i.e., any elements developed to perform the same function, regardless of structure. Thus, the scope of the present invention is not intended to be limited to the exemplary embodiments shown and described herein. Rather, the scope and spirit of the present invention are embodied by the appended claims.

Claims

1. 1. A method for providing an index for selecting patients with myelofibrosis who are more likely to benefit from treatment with a telomerase inhibitor, comprising: (a) testing a patient to determine whether said patient has high molecular weight risk (HMR), wherein having HMR comprises the presence of a mutation in at least one gene selected from the group consisting of additional sex comb-like 1 (ASXL1), enhancer of zest homolog 2 (EZH2), serine- and arginine-rich splicing factor 2 (SRSF2), isocitrate dehydrogenase 1 (IDH1), and isocitrate dehydrogenase 2 (IDH2); and / or (b) testing the patient for average relative telomere length by analyzing the relative length of telomeric nucleic acid in target cells present in a biological sample from said patient; and providing an index for selecting the patient with myelofibrosis who is more likely to benefit from treatment with the telomerase inhibitor based on the HMR or the average relative telomere length, the patient has the HMR, or if the patient has a mean relative telomere length in target cells present in a biological sample from the patient determined to be less than or equal to the 50th percentile of a range of relative telomere lengths determined from one or more known standards; and providing a method for treating a patient with a telomerase inhibitor comprising administering to said patient a therapeutically effective amount of a telomerase inhibitor. wherein the telomerase inhibitor is imetelstat or a pharmaceutically acceptable salt thereof.

2. 2. The method of claim 1, wherein the method comprises testing the patient to determine whether the patient has HMR, wherein having HMR comprises the presence of a mutation in at least one gene selected from ASXL1, EZH2, SRSF2, IDH1, and IDH2, and wherein if the patient has the HMR, it indicates that the patient is more likely to benefit from treatment with the telomerase inhibitor.

3. 2. The method of claim 1, wherein the method comprises testing a patient for an average relative telomere length by analyzing the relative length of telomeric nucleic acid in target cells present in a biological sample from the patient, wherein if the patient has an average relative telomere length in target cells present in the biological sample from the patient that is determined to be at or below the 50th percentile of a range of relative telomere lengths determined from one or more known standards, it indicates that the patient is more likely to benefit from treatment with the telomerase inhibitor.

4. The method according to any one of claims 1 to 3, wherein the myelofibrosis is selected from the group consisting of primary myelofibrosis, myelofibrosis that develops after polycythemia vera (post-PV MF), and myelofibrosis that develops after essential thrombocythemia (post-ET MF).

5. The method of any one of claims 1 to 4, wherein the patient has not previously received a JAK inhibitor therapy.

6. The patient: had received JAK inhibitor therapy, and the patient was resistant to JAK inhibitor therapy; have previously received JAK inhibitor therapy and have relapsed; or The method of any one of claims 1 to 4, wherein the patient has previously received JAK inhibitor therapy and discontinued the JAK inhibitor therapy due to treatment-related toxicity or intolerance.

7. The method of any one of claims 1 to 6, wherein the telomerase inhibitor is imetelstat sodium.

8. 8. The method of any one of claims 1 to 7, further comprising obtaining a sample comprising DNA from the patient, wherein the sample comprises bone marrow, peripheral blood, or a combination thereof.

9. 1. A composition for the treatment of myelofibrosis in patients with myelofibrosis and a mean relative telomere length at or below the 50th percentile of a high molecular weight risk (HMR) or relative telomere length range determined from one or more known criteria, said composition comprising a telomerase inhibitor; (a) the patient is determined to have HMR, wherein having HMR comprises the presence of a mutation in at least one gene selected from the group consisting of ASXL1, EZH2, IDH1, and IDH2; and / or (b) cells present in the patient-derived biological sample are determined to have an average relative telomere length determined to be less than or equal to the 50th percentile of a range of relative telomere lengths determined from one or more known standards; The composition, wherein the selected patient is more likely to benefit from treatment with a telomerase inhibitor, and the telomerase inhibitor is imetelstat or a pharmaceutically acceptable salt thereof.

10. 10. The composition of claim 9, wherein the patient is determined to have HMR, and having HMR comprises the presence of a mutation in at least one gene selected from the group consisting of ASXL1, EZH2, IDH1, and IDH2.

11. 10. The composition of claim 9, wherein cells present in the patient-derived biological sample have been determined to have an average relative telomere length determined to be less than or equal to the 50th percentile of a range of relative telomere lengths determined from one or more known standards.

12. The composition according to any one of claims 9 to 11, wherein the myelofibrosis is selected from the group consisting of primary myelofibrosis, myelofibrosis that develops after polycythemia vera (post-PV MF), and myelofibrosis that develops after essential thrombocythemia (post-ET MF).

13. The composition of any one of claims 9 to 12, wherein the patient has not previously received JAK inhibitor therapy.

14. The patient: had received JAK inhibitor therapy, and the patient was resistant to JAK inhibitor therapy; have previously received JAK inhibitor therapy and have relapsed; or The composition of any one of claims 9 to 12, wherein the patient has undergone JAK inhibitor therapy and discontinued the JAK inhibitor therapy due to treatment-related toxicity or intolerance.

15. The composition of any one of claims 9 to 14, wherein the telomerase inhibitor is imetelstat sodium.

Images