Subcutaneous telomerase inhibitor composition and method of using the same

Abstract

Aspects of the present disclosure include telomerase inhibitor compositions formulated for subcutaneous administration. A composition according to a specific embodiment comprises a hyaluronidase enzyme and a telomerase inhibitor having an oligonucleotide and a lipid moiety connected to the 5' and / or 3' ends of the oligonucleotide. A method of subcutaneously administering a telomerase inhibitor composition, such as in the treatment of neoplasms, is also described. A kit with or without a subcutaneous injection device is also provided.

Description

Technology Field

[0001] Cross-reference regarding related applications

[0002] This application claims the benefit of U.S. Provisional Application No. 63 / 053,455 filed July 17, 2020 and U.S. Provisional Application No. 63 / 128,708 filed December 21, 2020, the entire contents of which are incorporated herein by reference. Background Technology

[0003] Hematological malignancies are a form of cancer that originates in cells of blood-forming tissues, such as bone marrow, or in cells of the immune system. Examples of blood cancers include acute and chronic leukemia, lymphoma, multiple myeloma, and myelodysplastic syndrome.

[0004] Myeloproliferative neoplasms (MPNs) are a group of blood disorders that occur when the body produces too many white blood cells, red blood cells, or platelets. This overproduction of blood cells in the bone marrow can cause problems with blood flow and lead to various symptoms. MPNs can originate from myeloid lineage precursors in the bone marrow. MPNs are characterized by myeloproliferation without dysplasia, myelohypercellularity, and a predisposition to thrombosis, hemorrhage, and myelofibrosis. Various types of chronic myeloproliferative disorders exist. The MPN disease spectrum includes essential thrombocytosis (ET), polycythemia vera (PV), chronic myeloid leukemia (CML), myelofibrosis (MF), chronic neutrophilic leukemia, chronic eosinophilic leukemia, and acute myeloid leukemia (AML). Myelodysplastic syndrome (MDS) is a group of conditions involving cancers of the blood and bone marrow. Myelodysplastic syndrome (MDS) includes diseases such as refractory anemia, refractory anemia with excessive blast cells, refractory cytopenia with multiple lineage dysplasia, refractory cytopenia with single lineage dysplasia, and chronic myelomonocytic leukemia (CMML).

[0005] Imetelstat or sodium imetelstat is a telomerase inhibitor that binds with high affinity to the template region of the RNA component of telomerase. Studies have shown that imetelstat or sodium imetelstat inhibits telomerase activity and is effective against cell proliferation in numerous different cancer cell lines and human tumors, as described in U.S. Patent No. 7,494,982. Imetelstat or sodium imetelstat has been used in clinical trials in patients with myelodysplastic syndrome and hematological malignancies, including myeloproliferative neoplasms and solid tumors. In clinical trials in patients with low-risk myelodysplastic syndrome, intravenously administered imetelstat or sodium imetelstat was shown to achieve sustained transfusion independence in certain patients. Furthermore, clinical trials in patients with myelofibrosis have shown that intravenously administered imetelstat or imetelstat sodium can achieve improvements in overall survival and myelofibrosis, which are correlated with a reduction in the mutational burden of malignant clones and cytogenetic abnormalities.

[0006] In the case of subcutaneous administration, a large volume of the liquid composition of a telomerase inhibitor is administered to achieve an effective dose. The term "subcutaneous" refers to the subcutaneous layer of the skin, such as the subcutaneous lining. The subcutaneous stromal matrix consists of fibrous proteins in a viscoelastic gel of glycosaminoglycans. The glycosaminoglycans in the subcutaneous tissue contain glycohyaluronan (HA), which are unsulfated repeating linear disaccharides. Delivering large volumes of liquid into the subcutaneous tissue can be restrictive or painful. Hyaluronidase enzymes (e.g., soluble hyaluronidase glycoproteins) facilitate the rapid depolymerization of hyaluronan in the extracellular space of the subcutaneous tissue and reduce the viscosity of the stromal tissue, thereby increasing hydrodynamic conductivity and allowing a larger volume of liquid to be administered into the subcutaneous tissue.

[0007] Aspects of the present disclosure include telomerase inhibitor compositions formulated for subcutaneous administration. A composition according to a specific embodiment comprises a hyaluronidase enzyme and a telomerase inhibitor having an oligonucleotide and a lipid moiety linked to the 5' and / or 3' ends of the oligonucleotide. A method of subcutaneously administering a telomerase inhibitor composition for the treatment of cancer and the alleviation of cancer-related symptoms is described. A method of subcutaneously administering a telomerase inhibitor composition in the treatment of hematological malignancies is also described. A method of subcutaneously administering a telomerase inhibitor composition in the treatment of refractory anemia, refractory anemia with hyperblastoma, refractory cytopenia with multiple lineage dysplasia, refractory cytopenia with single lineage dysplasia, and myelodysplastic syndrome (MDS), including diseases such as chronic myelomonocytic leukemia (CMML), is also described. A method of subcutaneously administering a telomerase inhibitor composition in the treatment of myeloproliferative neoplasms such as essential thrombocytosis (ET), polycythemia vera (PV), chronic myeloid leukemia (CML), myelofibrosis (MF), chronic neutrophilic leukemia, chronic eosinophilic leukemia, and acute myeloid leukemia (AML) is also described. A method of subcutaneously administering a telomerase inhibitor composition in the treatment of lymphoid neoplasms is also described. A method of subcutaneously administering a telomerase inhibitor composition in the treatment of solid tumors is also described. Liquid or lyophilized unit dosage forms comprising a hyaluronidase enzyme and a telomerase inhibitor are described. A kit comprising a combination of a telomerase inhibitor and a hyaluronidase enzyme is also provided. A kit having a combination of a telomerase inhibitor and a hyaluronidase enzyme and a subcutaneous injection device is also provided.

[0008] In some embodiments, the hyaluronidase enzyme is recombinant human hyaluronidase. In certain embodiments, the hyaluronidase enzyme is rHuPH20. In some embodiments, the telomerase inhibitor subcutaneous composition comprises one or more soluble hyaluronidase glycoproteins. In some embodiments, the hyaluronidase enzyme is present in the composition in an amount of 100 U to 50,000 U. In some cases, the hyaluronidase enzyme is present in the composition in an amount of 100 U / mL to 50,000 U / mL. In some cases, the hyaluronidase enzyme is present in the composition in an amount of 100 U / mL to 2,000 U / mL. In some embodiments, the hyaluronidase enzyme (e.g., soluble hyaluronidase glycoprotein) is present in an amount that facilitates subcutaneous administration of the target telomerase inhibitor composition. In some cases, the hyaluronidase enzyme is present in an amount that facilitates the rapid depolymerization of hyaluronan in the extracellular space of subcutaneous tissue, thereby reducing the viscosity of the interstitial tissue, increasing hydrodynamic conductivity, and allowing a larger amount of telomerase inhibitor composition to be administered into the subcutaneous tissue.

[0009] In some embodiments, the hyaluronidase enzyme is a variant of recombinant human hyaluronidase. In certain embodiments, the hyaluronidase enzyme is a variant of hyaluronidase PH20 having one or more amino acid residue additions, deletions, or substitutions to the amino acid sequence of wild-type PH20, e.g., a variant hyaluronidase PH20 having one or more amino acid residue additions, deletions, or substitutions to the amino acid sequence of mature wild-type PH20. In certain embodiments, the hyaluronidase enzyme is a fragment of wild-type hyaluronidase PH20. In certain embodiments, the hyaluronidase enzyme is a fragment of wild-type hyaluronidase PH20 having one or more amino acid residue additions, deletions, or substitutions to the amino acid sequence of wild-type hyaluronidase PH20.

[0010] The composition may include one or more pharmaceutically acceptable excipients. In some embodiments, the composition includes one or more sugars. In some cases, the sugars include one or more monosaccharides. The monosaccharides may be present in the composition in an amount of 1 mM to 1000 mM, e.g., 10 mM to 500 mM. In some cases, the sugars include one or more polysaccharides. In some cases, the polysaccharides include sucrose. In other cases, the polysaccharides include trehalose. The polysaccharides may be present in the composition in an amount of 1 mM to 1000 mM, e.g., 10 mM to 500 mM. In certain cases, the polysaccharides are present in the composition in an amount of 100 mM to 300 mM.

[0011] In some embodiments, the composition comprises one or more amino acids. In certain embodiments, the composition comprises a certain amount of methionine. In other embodiments, the composition comprises a certain amount of histidine. In these embodiments, the amino acids may be present in the composition in an amount of 1 mM to 200 mM, e.g., 1 mM to 100 mM. In certain embodiments, the amino acids are present in the composition in an amount of 1 mM to 50 mM.

[0012] The composition may also include one or more buffers. In some cases, the buffer is present in the composition in an amount to maintain the composition at a predetermined pH. For example, one or more buffers may be present in the composition to maintain the composition at a pH of 3.0 to 9.0, e.g., pH 5.5 to 7.5. In some embodiments, the buffer is present in the composition in an amount comprising 1 mM to 250 mM, e.g., 1 to 100 mM and 1 mM to 50 mM. In certain cases, the buffer is a histidine buffer. In certain cases, the composition comprises methionine and polysorbate 80 or polysorbate 20.

[0013] In any embodiment of the present invention, the telomerase inhibitor comprises an oligonucleotide. In some embodiments, the oligonucleotide is complementary to the RNA component of telomerase. In some embodiments, the oligonucleotide is 10 to 20 base pairs long. In some embodiments, the oligonucleotide comprises the sequence TAGGGTTAGACAA. In some embodiments of any embodiment of the present invention, the oligonucleotide comprises at least one N3' → P5' thiophosphoramidate nucleoside linkage. In some embodiments of any embodiment of the present invention, the oligonucleotide comprises all N3' → P5' thiophosphoramidate nucleoside linkages. A telomerase inhibitor according to a specific embodiment is an oligonucleotide comprising a lipid moiety linked to the 5' and / or 3' ends of the oligonucleotide. In certain embodiments, the lipid moiety of the telomerase inhibitor is bound to the 5' and / or 3' ends of the oligonucleotide via a linker. In some cases, the linker is a glycerol or aminoglycerol linker. In certain cases, the lipid portion of the telomerase inhibitor is a palmitoyl (C16) moiety. In some embodiments, the target composition comprises imetelstat or a pharmaceutically acceptable salt thereof. In certain embodiments, the composition comprises imetelstat sodium.

[0014] In some embodiments, the amount of telomerase inhibitor in the subcutaneous composition is about 0.5 to about 5 mg, about 5 to about 10 mg, about 10 to about 15 mg, about 15 to about 20 mg, about 20 to about 25 mg, about 20 to about 50 mg, about 25 to about 50 mg, about 50 to about 75 mg, about 50 to about 100 mg, about 75 to about 100 mg, about 100 to about 125 mg, about 125 to about 150 mg, about 150 to about 175 mg, about 175 to about 200 mg, about 200 to about 225 mg, about 225 to about 250 mg, about 250 to about 300 mg, about 300 to about 350 mg, about 350 to about 400 mg, about 400 to about 450 mg, about 450 to about 500 mg, about 500 mg to about 600 mg, about 600 mg to about 700 mg, about 700 mg to about 800 mg, about 800 mg to about 900 mg, about 900 mg to about 1000 mg, about 1100 mg to about 1200 mg, about 1200 mg to about 1300 mg, about 1300 mg to about 1400 mg, about 1400 mg to about 1500 mg, about 1500 mg to about 1600 mg, about 1600 mg to about 1700 mg, about 1700 mg to about 1800 mg, about 1800 mg to about 1900 mg, about 1900 mg to about 2000 mg, about 2000 It is mg to about 2100 mg, about 2100 mg to about 2200 mg, about 2200 mg to about 2300 mg, about 2300 mg to about 2400 mg, and about 2400 mg to about 2500 mg.

[0015] In some embodiments, the amount of telomerase inhibitor is in a unit dosage form having an amount ranging from about 5 mg to about 1000 mg, 5 mg to about 500 mg, e.g., about 30 mg to about 300 mg, or about 50 mg to about 200 mg. In some embodiments, the amount of telomerase inhibitor is in a unit dosage form having an amount ranging from about 500 mg to about 3000 mg, 750 mg to about 2500 mg, e.g., about 1000 mg to about 2000 mg, or about 50 mg to about 200 mg. The unit dosage form may be liquid or lyophilized.

[0016] In some embodiments, the concentration of the telomerase inhibitor in the composition is, for example, about 0.1 to about 300 mg / ml, about 0.1 to about 200 mg / ml, about 0.1 to about 180 mg / ml, about 0.1 to about 160 mg / ml, about 0.1 to about 140 mg / ml, about 0.1 to about 120 mg / ml, about 0.1 to about 100 mg / ml, about 0.1 to about 80 mg / ml, about 0.1 to about 60 mg / ml, about 0.1 to about 40 mg / ml, about 0.1 to about 20 mg / ml, about 0.1 to about 10 mg / ml, about 2 to about 40 mg / ml, about 4 to about 35 mg / ml, about 6 to about 30 mg / ml, about 8 to about 25 mg / ml, about 10 to about It is diluted (about 0.1 mg / ml) or concentrated (about 300 mg / ml) to include either 20 mg / ml, about 12 to about 15 mg / ml, or any of about 0.1 mg / ml, 0.2 mg / ml, 0.3 mg / ml, 0.4 mg / ml, 0.5 mg / ml, 0.6 mg / ml, 0.7 mg / ml, 0.8 mg / ml, 0.9 mg / ml, 1 mg / ml, 1.1 mg / ml, 1.2 mg / ml, 1.3 mg / ml, 1.4 mg / ml, 1.5 mg / ml, 1.6 mg / ml, 1.7 mg / ml, 1.8 mg / ml, 1.9 mg / ml, 2 mg / ml, 2.1 mg / ml, 2.2 mg / ml, 2.3 mg / ml, 2.4 mg / ml, or 2.5 mg / ml. mg / ml). In some embodiments, the concentration of the telomerase inhibitor is at least about 0.1 mg / ml, 0.2 mg / ml, 0.3 mg / ml, 0.4 mg / ml, 0.5 mg / ml, 1.3 mg / ml, 1.5 mg / ml, 2 mg / ml, 3 mg / ml, 4 mg / ml, 5 mg / ml, 6 mg / ml, 7 mg / ml, 8 mg / ml, 9 mg / ml, 10 mg / ml, 11 mg / ml, 12 mg / ml, 13 mg / ml, 14 mg / ml, 15 mg / ml, 16 mg / ml, 17 mg / ml, 18 mg / ml, 19 mg / ml, 20 mg / ml, 21 mg / ml, 22 mg / ml, 23 mg / ml, 24 mg / ml, 25 mg / ml, 26 mg / ml, 27 mg / ml, 28 mg / ml, 29 mg / ml, 30 mg / ml, 31 mg / ml, 32 mg / ml, 33 mg / ml, 33.3 mg / ml, 34 mg / ml, 35 mg / ml, 36 mg / ml, 37 mg / ml, 38 mg / ml, 39 mg / ml, 40 mg / ml, 50 mg / ml, 60 mg / ml, 70 mg / ml, 80 mg / ml, 90 mg / ml, 100 mg / ml, 110 mg / ml, 120 mg / ml, 130 mg / ml, 140 mg / ml, 150 mg / ml, 160 mg / ml, 170 mg / ml, 180 mg / ml, 190 mg / ml, 200 mg / ml, 210 mg / ml, 220 mg / ml, 230 mg / ml, 240 mg / ml, 250 mg / ml, 260 mg / ml, 270 mg / ml, 280 mg / ml, 290 mg / ml, or Either 300 mg / ml. In certain embodiments, the composition is formulated to contain a telomerase inhibitor, such as imetelstat or imetelstat sodium, in doses of about 2.0 mg / kg to about 20.0 mg / kg, about 3.0 mg / kg to about 15.0 mg / kg, for example, about 4.0 mg / kg to about 10 mg / kg, about 7.5 mg / kg to 9.4 mg / kg, about 9 mg / kg to about 11 mg / kg, and about 11 mg / kg to about 14 mg / kg. In certain embodiments, the composition contains a telomerase inhibitor, such as imetelstat or imetelstat sodium, at 4.0 mg / kg, 4.1 mg / kg, 4.2 mg / kg, 4.3 mg / kg, 4.4 mg / kg, 4.5 mg / kg, 4.6 mg / kg, 4.7 mg / kg, 4.8 mg / kg, 4.9 mg / kg, 5.0 mg / kg, 5.1 mg / kg, 5.2 mg / kg, 5.3 mg / kg, 5.4 mg / kg, 5.5 mg / kg, 5.6 mg / kg, 5.7 mg / kg, 5.8 mg / kg, 5.9 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, 7.0 mg / kg, 7.1 mg / kg, 7.2 mg / kg, 7.3 mg / kg, 7.4 mg / kg, 7.5 mg / kg, 7.6 mg / kg, 7.7 mg / kg, 7.8 mg / kg, 7.9 mg / kg, 8.0 mg / kg, 8.1 mg / kg, 8.2 mg / kg, 8.3 mg / kg, 8.4 mg / kg, 8.5 mg / kg, 8.6 mg / kg, 8.7 mg / kg, 8.8 mg / kg, 8.9 mg / kg, 9.0 mg / kg, 9.1 mg / kg, 9.2 mg / kg, 9.3 mg / kg, 9.4 mg / kg, 9.5 mg / kg, 9.6 mg / kg, 9.7 mg / kg, 9.8 mg / kg, 9.9 mg / kg, 10.0 mg / kg, 10.5 mg / kg, 11.0 mg / kg, 11.5 mg / kg, 12.0 mg / kg, 12.5 mg / kg, 13.0 mg / kg, 13.5 mg / kg, 14.0 mg / kg, 14.5 mg / kg, 15.0 mg / kg, 15.5 mg / kg, 16.0 mg / kg, 16.5 mg / kg, 17.0 mg / kg, 17.0 mg / kg, 17.5 mg / kg, 18.0 mg / kg, 18.5 mg / kg, 19.0 mg / kg, 19.5 mg / kg, 또는 20.0 mg / kg의 투여량으로 포함하도록 제형화된다.

[0017] Aspects of the present disclosure also include a method for subcutaneously administering a telomerase inhibitor composition to a subject. In carrying out the method according to a specific embodiment, a composition having a telomerase inhibitor and a hyaluronidase enzyme is administered subcutaneously to a subject. In some embodiments, the composition is administered to the subject by subcutaneous injection. In other embodiments, the composition may be administered to the subject from an implanted device, such as a subcutaneous implanted catheter. In certain embodiments, the telomerase inhibitor composition is administered to the subject using a subcutaneous bolus injection device configured to deliver a predetermined amount of the composition subcutaneously to the subject. In certain embodiments, the telomerase inhibitor and the hyaluronidase enzyme are administered simultaneously. For example, the telomerase inhibitor and the hyaluronidase enzyme may be mixed together prior to administration. In some cases, the telomerase inhibitor and the hyaluronidase enzyme are administered sequentially. For example, the hyaluronidase enzyme may be administered prior to the telomerase inhibitor. In some cases, the hyaluronidase enzyme is administered first, followed by the telomerase inhibitor. For example, the hyaluronidase enzyme may be administered immediately after the telomerase inhibitor is administered.

[0018] In certain embodiments, the target method includes a step of treating a neoplasm. In some embodiments, the neoplasm may be a solid tumor cancer. Examples of cancers for treatment according to embodiments of the present disclosure include, for example, adrenocortical carcinoma, anal cancer, appendicitis, astrocytoma, atypical anomaly / rhabdomyotum, basal cell carcinoma, cholangiocarcinoma (extrahepatic), bladder cancer, bone cancer (e.g., Ewing sarcoma, osteosarcoma and malignant fibrous histiocytoma, etc.), brainstem glioma, brain tumor (e.g., astrocytoma, central nervous system embryonic tumor, central nervous system germ cell tumor, craniopharyngioma, ependymoma, etc.), breast cancer (e.g., female breast cancer, male breast cancer, pediatric breast cancer, etc.), bronchial tumor, carcinoid tumor (e.g., pediatric, gastrointestinal, etc.), unknown primary carcinoma, heart tumor, cervical cancer, colon cancer, colorectal cancer, craniopharyngioma, duct (e.g., bile duct, extrahepatic, etc.), intraductal carcinoma in situ (DCIS), embryonic tumor, endometrial cancer, ependymoma, esophageal cancer. Sensory neuroblastoma, Ewing sarcoma, extracranial germ cell tumor, extragonadal germ cell tumor, extrahepatic cholangiocarcinoma, ocular cancer (intraocular melanoma, retinoblastoma, etc.), fibrous histiocytoma of bone (malignant, osteosarcoma, etc.), gallbladder cancer, gastric cancer, gastrointestinal carcinoid tumor, gastrointestinal stromal tumor (GIST), germ cell tumor (e.g., extracranial, extragonadal, ovarian, testicular, etc.), gestational trophoblastic disease, glioma, hairy cell leukemia, head and neck cancer, heart cancer, hepatocellular carcinoma, histiocytosis (e.g., Langerhans cell, etc.), hypopharyngeal cancer, intraocular melanoma, islet cell tumor (e.g., pancreatic neuroendocrine tumor, etc.), renal cancer (e.g., renal cell, Wilms tumor, pediatric renal tumor, etc.), Langerhans cell histiocytosis, laryngeal cancer, lip and oral cancer, liver cancer (primary), lobular carcinoma in situ (LCIS), lung cancer (e.g., Non-small cell, small cell, etc.), malignant fibrous histiocytoma and osteosarcoma of bone, melanoma, Merkel cell carcinoma, mesothelioma, latent primary metastatic squamous cell cervical cancer, oral cancer, multiple endocrine neoplasm syndrome, nasal and paranasal sinus cancer, nasopharyngeal cancer,Neuroblastoma, non-small cell lung cancer, oral cancer, oral cancer (e.g., lips, etc.), oropharyngeal cancer, osteosarcoma and malignant fibrous histiocytoma of bone, ovarian cancer (e.g., epithelial, germ cell tumors, tumors with low likelihood of malignancy, etc.), pancreatic cancer, pancreatic neuroendocrine tumor (islet cell tumor), papillomatosis, paraganglioma, sinus and nasal cavity cancer, parathyroid cancer, penile cancer, pharyngeal cancer, pheochromocytoma, pituitary tumor, pleural-pulmonary blastoma, prostate cancer, rectal cancer, renal cell (kidney) cancer, renal pelvis and ureter, transitional cell carcinoma, retinoblastoma, rhabdomyosarcoma, salivary gland cancer, Sézary syndrome, skin cancer (e.g., pediatric, melanoma, Merkel cell carcinoma, non-melanoma, etc.), small cell lung cancer, small intestine cancer, soft tissue sarcoma, squamous cell carcinoma, squamous cell cervical cancer (e.g., latent primary, metastatic, etc.), gastric Cancer, testicular cancer, throat cancer, thymomas and thymic carcinomas, thyroid cancer, transitional cell carcinoma of the renal pelvis and ureter, ureteral and renal pelvis cancer, urethral cancer, uterine cancer (e.g., endometrial, etc.), uterine sarcoma, vaginal cancer, vulvar cancer, Waldenström macroglobulinemia, Wilms tumor, etc. may be included, but not limited thereto. In certain embodiments, the method comprises the step of treating a subject having a neoplasm as described in U.S. Patent No. 7,494,982.

[0019] In some embodiments, the neoplasm may be a hematological neoplasm. In certain embodiments, the target method comprises the step of treating a subject having a myeloproliferative neoplasm. In some cases, the myeloproliferative neoplasm is a myelofibrosis (MF), such as primary myelofibrosis or myelofibrosis prior to ET or post-PV (post-ET MF or post-PV MF). In other embodiments, the myeloproliferative neoplasm includes essential thrombocytosis (ET), polycythemia vera (PV), chronic myeloid leukemia (CML), chronic neutrophilic leukemia, chronic eosinophilic leukemia, and acute myeloid leukemia (AML). In other cases, the hematological neoplasm is myelodysplastic syndrome (MDS). In yet another embodiment, the hematological neoplasm is myelodysplastic syndrome (MDS) having a monolithic non-deletion (5q). Myelodysplastic syndrome (MDS) includes diseases such as refractory anemia, refractory anemia with excessive blasts, refractory cytopenia with multiple lineage dysplasia, refractory cytopenia with single lineage dysplasia, and chronic myelomonocytic leukemia (CMML). A method according to a specific embodiment also includes the step of diagnosing a subject as having a myeloproliferative neoplasm. In one example, the method includes the step of diagnosing a subject as having a myelofibrosis (MF), such as primary myelofibrosis. In some embodiments, the subject has not previously received a telomerase inhibitor (e.g., the telomerase inhibitor Naivim). In some embodiments, the subject is a subject with low-risk transfusion-dependent MDS that is relapsed or refractory to an erythropoietin-stimulating agent (ESA). In some embodiments, the subject has not received prior treatment with a hypomethylating agent (HMA). In some embodiments, the subject has not been given prior treatment with lenalidomide. In some embodiments, the subject is a non-deleted (5q) subject. In some embodiments, the subject is a relapsed or refractory subject to a Janus kinase (JAK) inhibitor.In some embodiments, the target method comprises the step of treating a subject having a myeloproliferative neoplasm as described in U.S. Patent No. 9,375,485 and International Patent Publications No. WO 2019 / 023667 and WO 2020 / 028261, the disclosures of which are incorporated herein by reference.

[0020] In certain embodiments, the target method includes the step of treating a target having a lymphoid neoplasm. In some embodiments, the lymphoid neoplasm (e.g., lymphoma) is a B-cell neoplasm. In some embodiments, the lymphoid neoplasm (e.g., lymphoma) is a T-cell and / or putative NK-cell neoplasm.

[0021] In certain embodiments, the telomerase inhibitor and the hyaluronidase enzyme are administered subcutaneously to a subject. The telomerase inhibitor composition and the hyaluronidase enzyme may be administered subcutaneously to a subject at least once a day, e.g., at least twice a day, e.g., at least three times a day, e.g., at least four times a day, and e.g., at least five times a day. In some embodiments, the composition is administered subcutaneously to a subject for at least one day, at least two days, at least three days, at least four days, at least five days, at least six days, at least seven days, e.g., at least ten days, e.g., at least fourteen days, e.g., at least twenty-one days. The administration may be carried out in a cycle of administration of the telomerase inhibitor composition and the hyaluronidase enzyme. In some embodiments, the cycle is once a day. In some embodiments, the cycle is every other day (i.e., once every two days). In some embodiments, the cycle is once every three days. In some embodiments, the cycle is once every 4 days. In some embodiments, the cycle is once every 5 days. In some embodiments, the cycle is once every 6 days. In some embodiments, the cycle is once every 7 days. In some embodiments, the cycle is once every 14 days, in some cases the cycle is once every 21 days, and in other cases the cycle is once every 28 days or more. The administration cycle of the telomerase inhibitor composition may be repeated for 1, 2, 3, 4, 5, 6, 7, 8, or more than 8 dose cycles for a total of 6 months or 1 year or 2 years or 3 years or 4 years or 5 years or 6 years or 7 years or 8 years or 9 years or 10 years or more. For example, a subsequent administration cycle may be performed after not administering the composition for at least 1 day following the performance of an administration cycle. The time between administration cycles may be 1 day or more, 2 days or more, 3 days or more, 4 days or more, 5 days or more, 6 days or more, 7 days or more, 10 days or more, 14 days or more, 21 days or more, or 28 days or more.

[0022] Without being bound by theory, the present invention provides compositions, unit dosage forms, and kits comprising both a telomerase inhibitor and a hyaluronidase enzyme as defined herein, having compatibility and stability for use in a method of treating a subject with a neoplasm by subcutaneous administration. The present invention also provides such compositions formulated for subcutaneous administration, wherein the compositions are safe and tolerable to the subject being treated and enable an effective pharmacokinetic profile for the telomerase inhibitor when co-formulated with the hyaluronidase enzyme. The present invention further provides a dosage form having an appropriate liquid volume containing a sufficient dose of the telomerase inhibitor for subcutaneous administration. Brief explanation of the drawing

[0023] Fig. 1 The time profile of plasma concentration of imetelstat sodium after subcutaneous injection and intravenous delivery in rats according to specific embodiments is shown. Fig. 2 It shows the inhibition of telomerase activity (%) relative to the treatment concentration (μM) for various sample formulations according to specific embodiments. Specific details for implementing the invention

[0024] Definition selection

[0025] The term "nucleoside" refers to a moiety having the following general structure, and , B represents a nucleobase, and the 2' carbon can be substituted as described below. When incorporated into oligomers or polymers, the 3' carbon is additionally connected to an oxygen or nitrogen atom. Nucleosides consist of 2'-deoxy and 2'-hydroxyl ( in other words,It may include forms (deoxyribose and ribose) and analogs thereof. In certain cases, the 5'-NH group may substitute for the 5'-oxygen. With respect to nucleosides, "analogs" include synthetic nucleosides having modified nucleobase moiety (see definition of "nucleobase" below) and / or modified sugar moiety, such as 2'-fluorosugars, among other analogs. These analogs are typically designed to affect binding properties, e.g., stability, specificity, etc. The term nucleoside is used, for example, in the literature (Komberg and Baker, DNA Replication Natural nucleosides, including 2'-deoxy and 2'-hydroxyl forms as described in the 2nd Ed. (Freeman, San Francisco, 1992), and analogs. With respect to nucleosides, "analogs" refer to modified nucleobase moiety (see definition of "nucleobase" below) and / or, for example, generally in the literature (Scheit, Nucleotide Analogs Synthetic nucleosides having modified sugar moiety as described in (John Wiley, New York, 1980). Such analogs include synthetic nucleosides designed to improve binding properties, e.g., stability, specificity, etc., as disclosed in the literature (Uhlmann and Peyman, Chemical Reviews 90:543-584, 1990). Oligonucleotides containing such nucleosides and typically containing linkages between synthetic nuclease-resistant nucleosides may themselves be referred to as "analogs."

[0026] "Polynucleotide" or "oligonucleotide" refers to a ribose and / or deoxyribose nucleoside subunit polymer or oligomer having about 2 to about 200 adjacent subunits. Nucleoside subunits may be joined by various inter-subunit linkages including, but not limited to, phosphodiesters, phosphotriesters, methylphosphonates, P3' → N5' phosphoramidates, N3' → P5' phosphoramidates, N3' → P5' thiophosphamidates, and phosphorothioate bonds. The term also includes such polymers or oligomers having modifications such as sugars (e.g., 2' substitutions), bases, and 3' and 5' ends. In an embodiment in which the oligonucleotide moiety comprises multiple inter-subunit linkages, each linkage may be formed using the same chemistry or a mixture of linkage chemistry may be used. When an oligonucleotide is represented by a character sequence such as "ATGUCCTG", the nucleotides will be understood to be in the order 5' → 3' from left to right. Representing the base sequence of an oligonucleotide in this manner does not imply the use of any specific type of nucleoside subunit in the oligonucleotide.

[0027] "Nuclear bases" refer to (i) natural DNA and RNA nuclear bases (uracil, thymine, adenine, guanine, and cytosine), (ii) modified nuclear bases or nuclear base analogs (e.g., (iii) 5-methylcytosine, 5-bromouracil or inosine, and nucleobases. Nucleobases are compounds whose molecular structure mimics the structure of a typical DNA or RNA base.

[0028] The term “lipid” is used extensively herein to encompass substances that are soluble in organic solvents but nearly, if not entirely, insoluble in water. The term lipid includes, but is not limited to, hydrocarbons, oils, fats (e.g., fatty acids and glycerides), sterols, steroids, and derivatives of these compounds. In some embodiments, lipids are fatty acids and their derivatives, hydrocarbons and their derivatives, and sterols such as cholesterol. Fatty acids usually contain an even number of carbon atoms (typically 12–24 carbons) in a straight chain, can be saturated or unsaturated, and can contain or be modified to contain various substituents. For simplicity, the term “fatty acid” also encompasses fats or fatty acid derivatives such as esters. In some embodiments, the term “lipid” also includes amphiphilic compounds containing both lipids and hydrophilic moiety.

[0029] "Individual," "patient," or "subject" may be a mammal, such as any common laboratory model organism. Mammals include, but are not limited to, human and non-human primates, farm animals, sports animals, pets, mice, rats, and other rodents. In some embodiments, the individual, patient, or subject is a human. In certain embodiments, the subject or patient has not previously received telomerase inhibitor therapy prior to certain embodiments, and such patient is a "telomerase inhibitor naive."

[0030] "Effective dose," "therapeutic effective dose," or "clinical effective dose" refers to the amount of telomerase inhibitor administered to a mammalian subject as a single dose or as part of a series of doses that is effective in producing the desired therapeutic effect.

[0031] As used herein, the terms “neoplasm,” “neoplasia,” or “neoplastic” refer to abnormal new cell growth. Unlike hyperplasia, neoplasmic proliferation persists even in the absence of the original stimulus. “Neoplasmic cells” refer to cells exhibiting relatively autonomous growth and thus exhibit an abnormal growth phenotype characterized by a significant loss of control over cell proliferation. Neoplasmic cells include cells that may be actively replicating or in a transient non-replicating quiescent state (G1 or G0); similarly, neoplasmic cells may include cells with a well-differentiated phenotype, a poorly differentiated phenotype, or a mixture of both types. Thus, not all neoplasmic cells are replicating at any given time. “Neoplasmic cells” encompass cells of benign neoplasms and cells of malignant neoplasms. “Neoplasmic progenitor cells” refer to cells of a cellular composition that possess the ability to become a neoplasm.

[0032] "Proliferative disorders" are any cellular disorders in which cells proliferate faster than normal tissue growth. Thus, "proliferative cells" are cells that proliferate faster than normal cells. Proliferative disorders include, but are not limited to, neoplasms. "Neoplasms" are abnormal tissue growths that form distinct masses that grow by cell proliferation at a faster rate than normal tissue growth. Neoplasms indicate a partial or complete absence of structural, organizational, and functional coordination with normal tissues. They can be broadly classified into three main types. Malignant neoplasms arising from epithelial structures are called carcinomas; malignant neoplasms arising from connective tissues such as muscle, cartilage, fat, or bone are called sarcomas; and malignant tumors affecting hematopoietic structures (structures related to blood cell formation) that include components of the immune system are called leukemias and lymphomas. Tumors are neoplasmic growths of the disease cancer. As used herein, neoplasms also referred to as "tumors" are intended to encompass hematopoietic neoplasms as well as solid neoplasms. Other proliferative diseases include, but are not limited to, neurofibromatosis.

[0033] As used herein, “a,” “an,” and “the” include plural references unless otherwise indicated.

[0034] All maximum numerical limits given throughout this specification are intended to include all lower numerical limits as if such lower numerical limits were expressly described herein. All minimum numerical limits given throughout this specification will include all upper numerical limits as if such upper numerical limits were expressly described herein. All numerical ranges given throughout this specification will include all narrower numerical ranges falling within these wider numerical ranges as if such narrower numerical ranges were all expressly described herein.

[0035] Before the present invention is further described, it should be understood that the specific embodiments described may naturally vary and are not limited thereto. Furthermore, since the scope of the invention is limited only by the appended claims, it should be understood that the terms used herein are merely for describing specific embodiments and are not intended to be limiting.

[0036] Where a range of values ​​is provided, unless otherwise clearly specified in the context, each intermediate value is understood to be encompassed within the upper and lower limits of the range and any other mentioned intermediate values ​​up to one-tenth of the lower limit unit. The upper and lower limits of these smaller ranges may independently be included in the smaller ranges and are also encompassed within the invention according to any specifically excluded limits from the mentioned ranges. Where a specified range includes one or all of the limits, the range excluding one or all of the included limits is also included in the invention.

[0037] For clarity, it is understood that specific features of the invention described in the context of separate embodiments may also be provided in combination in a single embodiment. Conversely, for brevity, various features of the invention described in the context of a single embodiment may also be provided individually or in any suitable sub-combination. All combinations of embodiments relating to the invention are specifically included in the invention, and each and every combination is disclosed herein as individually and explicitly disclosed, to the extent that such combinations include the gist of the matter, for example, a stable compound (i.e., a compound that can be prepared, isolated, characterized, and tested for biological activity). Furthermore, all sub-combinations of various embodiments and elements thereof (e.g., elements of chemical groups listed in the embodiments describing such variables) are also specifically included in the invention, and each and every such sub-combination is disclosed herein as individually and explicitly disclosed.

[0038] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as generally understood by those skilled in the art to which the present invention pertains. Any method and material similar or equivalent to those described herein may be used in the practice or testing of the present invention, but the method and material of interest are described hereafter. All publications mentioned herein are incorporated by reference to disclose and describe methods and / or materials related to those cited in the publications.

[0039] It should be noted that the singular forms (“a,” “an,” and “the”) used herein and in the appended claims include plural references unless the context otherwise clearly indicates. It should be further noted that the claims may be drafted to exclude any optional elements. As such, this statement is intended to serve as a preliminary basis for the use of exclusive terms such as “only,” “only,” etc., in connection with the citation of claim elements or the use of “negative” limitations.

[0040] It is understood that for clarity, specific features of the invention described in the context of separate embodiments may also be provided in combination in a single embodiment. Conversely, for brevity, various features of the invention described in the context of a single embodiment may also be provided individually or in any suitable sub-combination.

[0041] The publications discussed herein are provided only for disclosures prior to the filing date of this application. Nothing in this application shall be construed as an acknowledgment that the present invention is not qualified to precede such publications by prior art. Additionally, since the provided publication dates may differ from the actual publication dates, they must be verified independently.

[0042] Compounds as described herein may be purified by any means known in the art, including chromatographic means such as high-performance liquid chromatography (HPLC), preparative thin-layer chromatography, flash column chromatography, and ion exchange chromatography. Any suitable stationary phase, including normal phase and reverse phase as well as ionic resins, may be used. For example, refer to the literature (Introduction to Modern Liquid Chromatography, 2nd Edition, ed. LR Snyder and JJ Kirkland, John Wiley and Sons, 1979; and Thin Layer Chromatography, ed. E. Stahl, Springer-Verlag, New York, 1969).

[0043] The compounds described herein may contain one or more chiral centers and / or double bonds and may exist as stereoisomers, such as double bond isomers (i.e., geometric isomers), enantiomers, or diastereomers. Accordingly, all possible enantiomers and stereoisomers of the compounds, including stereoisomerically pure forms (e.g., geometrically pure, enantiomerically pure, or diastereomerically pure) and mixtures of enantiomers and stereoisomers, are included in the description of the compounds herein. The mixtures of enantiomers and stereoisomers may be decomposed into their constituent enantiomers or stereoisomers using separation techniques or chiral synthesis techniques well known to those skilled in the art. The compounds may also exist in several tautomeric forms, including enol forms, keto forms, and mixtures thereof. Accordingly, the chemical structures illustrated herein encompass all possible tautomeric forms of the exemplified compounds. The described compounds also include isotope-labeled compounds in which one or more atoms have an atomic mass different from the atomic mass typically found in nature. Examples of isotopes that may be incorporated into the compounds disclosed herein are 2 H, 3 H, 11 C, 13 C, 14 C, 15 N, 18 O, 17 Compounds include, but are not limited to, O, etc. Compounds may exist in solvated forms, including hydrated forms as well as nonsolved forms. Generally, compounds may be hydrated or solvated. Specific compounds may exist in various crystalline or amorphous forms. Generally, all physical forms are intended to be equivalent for the uses considered herein and within the scope of this disclosure.

[0044] details

[0045] Subcutaneous telomerase inhibitor composition

[0046] A aspect of the present disclosure comprises a telomerase inhibitor composition formulated for subcutaneous administration. The term “subcutaneous” is used herein in its ordinary sense to refer to the subcutaneous layer of the skin, such as the subcutaneous tissue layer. The subcutaneous stromal matrix consists of fibrous proteins in a viscoelastic gel of glycosaminoglycans. The glycosaminoglycans of the subcutaneous tissue comprise glycohyaluronan (HA), which is an unsulfated repeating linear disaccharide.

[0047] The composition of the present disclosure comprises a hyaluronidase enzyme. In some embodiments, the hyaluronidase enzyme is a mammalian-type hyaluronidase, such as endo-beta-N-acetylhexosaminidase, having tetrasaccharides and hexasaccharides as major end products. In some cases, the mammalian hyaluronidase possesses both hydrolytic and transglycosidase activity and can degrade hyaluronan and chondroitin sulfate. In other embodiments, the hyaluronidase enzyme is a bacterial hyaluronidase, such as endo-beta-N-acetylhexosaminidase, which produces a disaccharide end product by beta removal. In yet another embodiment, the hyaluronidase enzyme is an endo-beta-glucuronidase that produces tetrasaccharide and hexasaccharide end products through the hydrolysis of β-1-3 linkages. In some embodiments, the hyaluronidase enzyme in the target telomerase inhibitor composition comprises a mammalian hyaluronidase having a neutral active site or an acidic active site. In certain embodiments, the composition of interest comprises a recombinant human hyaluronidase enzyme. In certain cases, the recombinant human hyaluronidase enzyme is a PH20 recombinant human hyaluronidase enzyme (rHuPH20). In some embodiments, the telomerase inhibitor subcutaneous composition comprises one or more soluble hyaluronidase glycoproteins (sHASEGP). In some embodiments, the hyaluronidase enzyme (e.g., soluble hyaluronidase glycoprotein) facilitates the subcutaneous administration of the target composition. In some cases, the hyaluronidase enzyme is present in an amount that rapidly depolymerizes hyaluronan in the extracellular space, reduces the viscosity of the interstitial tissue to increase hydrodynamic conductivity, and allows a larger amount to be administered into the subcutaneous tissue. In certain embodiments, increased hydrodynamic conductivity induced by hyaluronidase enzyme through reduced interstitial viscosity allows for greater dispersion, thereby increasing the systemic bioavailability of the subcutaneously administered telomerase inhibitor described herein.

[0048] In certain embodiments, the composition comprises one or more hyaluronidase enzymes (e.g., soluble hyaluronidase glycoproteins), such as those described in International Patent Publication Nos. WO2004 / 078140 and WO 2006 / 091871 and U.S. Patent No. 7,767,429, the disclosures thereof of which are incorporated herein by reference.

[0049] In some embodiments, the hyaluronidase enzyme is a variant or fragment of recombinant human hyaluronidase that is active and capable of degrading hyaluronan. The sequence of the wild-type human PH20 hyaluronidase enzyme (Sequence No. 1) is shown in Table 1 below.

[0050]

[0051] In certain embodiments, the hyaluronidase is a soluble hyaluronidase. Soluble hyaluronidase includes any that exist in a soluble form upon expression and secretion from a cell. Such soluble hyaluronidase includes, but is not limited to, non-human soluble hyaluronidase, bacterial soluble hyaluronidase, bovine PH20, sheep PH20, and variants thereof. Among soluble hyaluronidases, human PH20 polypeptides modified to be soluble are included. For example, hyaluronidase such as human PH20 containing a glycophosphatidylinositol (GPI) anchor may be made soluble by cleavage and removal of all or part of the GPI anchor. In one example, human hyaluronidase PH20, which is typically membrane-bound via a GPI anchor, is made soluble by cleavage and removal of all or part of the GPI anchor at the C-terminus.

[0052] Soluble hyaluronidases also include neutrally active hyaluronidases such as soluble human PH20 polypeptides. In certain examples, the hyaluronidase for use in the compositions, combinations, and methods of the present invention is a soluble neutrally active hyaluronidase. Examples of hyaluronidases include soluble forms of PH20 from any species, such as the soluble form of PH20. Soluble forms of PH20 are known in the art. These include sheep and bovine PH20 polypeptides, and the soluble form of human PH20 of SEQ ID NO. 1. The soluble form of human PH20 of SEQ ID NO. 1. Such soluble forms include truncated forms thereof lacking all or part of the C-terminal GPI anchor, provided that the hyaluronidase is soluble (secreted upon expression) and retains hyaluronidase activity. Such forms are also typically mature forms lacking the signal peptide when expressed in cells. Full-length mature human PH20 (residues 36-509 of Sequence No. 1) occurs as a GPI-fixed polypeptide. As is known in the art, it becomes soluble by cleavage at the C-terminus. Such cleavage may remove all GPI anchor attachment sequences or only a portion of the GPI anchor attachments. However, the resulting polypeptide is soluble. If the soluble hyaluronidase retains a portion of the GPI anchor attachment signal sequence, 1, 2, 3, 4, 5, 6, 7 or more amino acid residues of the GPI anchor attachment signal sequence may be retained as long as the polypeptide is soluble. A polypeptide containing one or more amino acids of the GPI anchor is designated as an extended soluble hyaluronidase. A person skilled in the art can determine whether the polypeptide is GPI-fixed using methods well known in the art.These methods include, but are not limited to, the step of predicting the presence and location of the GPI anchor attachment signal sequence and the ω-site using a known algorithm, and performing solubility analysis before and after digestion with phosphatidylinositol-specific phospholipase C (PI-PLC) or D (PI-PLD).

[0053] An example of a soluble hyaluronidase is soluble human PH20. A soluble form of recombinant human PH20 has been produced and may be used in the compositions, combinations, and methods described herein. Descriptions and production of such soluble forms of PH20 are described in U.S. Patent Nos. 7,767,429, 8,202,517, 8,431,380, 8,431,124, 8,450,470, 8,765,685, 8,772,246, 7,871,607, 7,846,431, 7,829,081, 8,105,586, 8,187,855, 8,257,699, 8,580,252, 9,677,061, and 9,677,062, which are incorporated herein by reference, for example.

[0054] A recombinant soluble form of human PH20 has been produced and can be used in the compositions, combinations, and methods provided herein. For example, referring to SEQ NO. 1, which presents a sequence of a full-length precursor PH20 containing a signal sequence (residues 1-35), the soluble form is a C-terminal cleavage of human PH20 presented in SEQ NO. 1 having C-terminal amino acid residues 467, 468, 469, 470, 471, 472, 473, 474, 475, 476, 477, 478, 479, 480, 481, 482, 483, 484, 485, 486, 487, 488, 489, 490, 491, 492, 493, 494, 495, 496, 497, 498, 499, or 500 of the amino acid sequence presented in SEQ NO. 1. Polypeptide, or polypeptides exhibiting at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or greater sequence identity therewith, but not limited to, having activity at neutral pH and being soluble (secreted into the medium when expressed in mammalian cells). Soluble forms of human pH20 generally include those containing amino acids 36–464 as presented in SEQ No. 1. For example, when expressed in mammalian cells, the 35-amino acid N-terminal signal sequence is cleaved during processing, and the mature form of the protein is secreted. Accordingly, mature soluble polypeptides include those containing amino acids 36 to 467, 468, 469, 470, 471, 472, 473, 474, 475, 476, 477, 478, 479, 480, 481, 482 and 483 of sequence number 1.Examples of soluble hyaluronidase include a soluble PH20 polypeptide having an amino acid sequence presented as amino acid residues 36-482, 36-477, 366-478, 36-479, 36-480, 36-481, and 36-483 of SEQ ID NO. 1, a soluble human PH20 polypeptide having a length of 442, 443, 444, 445, 446, or 447 amino acids, and a soluble human PH20 polypeptide having a length of 442, 443, 444, 445, 446, or 447 amino acids that maintains hyaluronidase activity, for example, an amino acid sequence presented as amino acid residues 36-482, 36-477, 366-478, 36-479, 36-480, 36-481, and 36-483 of SEQ ID NO. 1 and at least 85%, 86%, 87%, 88%, 89%, It is a variant of this having 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity. These available forms of recombinant human PH20 are described, for example, in U.S. Patent Nos. 7,767,429, 8,202,517, 8,431,380, 8,431,124, 8,450,470, 8,765,685, 8,772,246, 7,871,607, 7,846,431, 7,829,081, 8,105,586, 8,187,855, 8,257,699, 8,580,252, 9,677,061, and 9,677,062, which are incorporated herein by reference. Because glycosylation is important for the catalytic activity and stability of hyaluronidase, the soluble form of PH20 is generally produced using a protein expression system that facilitates precise N-glycosylation to maintain polypeptide activity. These cells include, for example, Chinese hamster ovary (CHO) cells (e.g., DG44 CHO cells).

[0055] Soluble recombinant human PH20, including a recombinant form of human PH20, can be produced recombinantly. One such product is designated as rHuPH20; rHuPH20 generally refers to a composition produced upon expression in cells, e.g., CHO cells, of a nucleic acid encoding residues 36–482 of SEQ ID NO. 1, linked to a natural or heterogeneous signal sequence (residues 1–35 of SEQ ID NO. 1). rHuPH20 is produced by the expression of a nucleic acid molecule such as one encoding amino acids 1–482 (presented in SEQ ID NO. 1). Post-translational processing removes the 35-amino acid signal sequence, leaving a polypeptide or a mixture of polypeptides. As produced in culture medium, the product designated as rHuPH20 has heterogeneity at the C-terminus to include a mixture of species terminating at residues 477, 478, 479, 480, 481, and 482 in varying abundances, with reference to SEQ ID NO. 1. Hyaluronidase rHuPH20 is selected from the group consisting of polypeptides referencing Sequence No. 1 corresponding to amino acid residues 36-477; amino acid residues 36-478, c amino acid residues 36-479, amino acid residues 36-480, amino acid residues 36-481 and amino acid residues 36-482. Generally, the most abundant species is the 446-amino acid polypeptide corresponding to residues 36-481 of Sequence No. 1.

[0056] Soluble human PH20 polypeptides include those referred to as extended soluble hyaluronidase. Extended soluble hyaluronidase may be produced by preparing a C-terminal cleavage of any natural GPI-fixed hyaluronidase such that the resulting polypeptide is soluble and contains one or more amino acid residues from the GPI anchor attachment signal sequence (see, e.g., U.S. Patent No. 8,927,249). Extended soluble human PH20 polypeptides include those terminating at any of residues approximately 495–500 of SEQ ID NO. 1. The mature form starts at residue 36. Extended soluble human PH20 polypeptides are neutrally active and soluble. They may contain amino acid substitutions and have at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, and 95% of the extended soluble PH20 polypeptide.

[0057] Hyaluronidase can be produced recombinantly or purified or partially purified from natural sources, such as testicular extracts, for example. Methods for producing recombinant proteins containing recombinant hyaluronidase are well known in the field. Soluble pH20 is produced in cells that facilitate precise N-glycosylation to maintain activity, such as CHO cells (e.g., DG44 CHO cells).

[0058] Glycosylation, including N- and O-linking glycosylation, can be important for the catalytic activity and stability of some hyaluronidases, including soluble pH 20 hyaluronidases. For some hyaluronidases, the removal of N-linking glycosylation can result in the near-complete inactivation of hyaluronidase activity. N-linked oligosaccharides are classified into several basic types (oligomannose, complex, hybrid, sulfated), all of which possess a (Man) 3-GlcNAc-GlcNAc-core attached via the amide nitrogen of an Asn residue belonging to the -Asn-Xaa-Thr / Ser- sequence (where Xaa is not Pro). Glycosylation at the -Asn-Xaa-Cys- site has been reported for coagulation protein C. In some cases, hyaluronidases such as PH20 hyaluronidase may contain N-glycosidic and O-glycosidic linkages. For example, PH20 has N-linked oligosaccharides as well as O-linked oligosaccharides. N82, N166, N235, N254, N368, and N393 of the human PH20 exemplified in Sequence No. 1 have six potential N-linked glycosylation sites.

[0059] In certain embodiments, the hyaluronidase enzyme is a variant or fragment of hyaluronidase PH20 having one or more amino acid residue additions, deletions, or substitutions in the amino acid sequence of mature wild-type PH20, such as hyaluronidase PH20 having one or more amino acid residue additions, deletions, or substitutions in the amino acid sequence of wild-type PH20. In some embodiments, the variant or fragment hyaluronidase PH20 comprises one or more amino acid substitutions, additions, or deletions located in the alpha-helix 8 region of PH20. In some embodiments, the variant or fragment hyaluronidase PH20 comprises one or more amino acid substitutions, additions, or deletions located in the linker region between alpha-helix 7 and alpha-helix 8 of PH20. In certain embodiments, the variant or fragment hyaluronidase PH20 comprises the amino acid sequence from L36 to S490 of wild-type PH20 (i.e., L36 to S490 of sequence number 1).

[0060] In an embodiment, the PH20 variant or a fragment thereof has amino acid sequence homology with the amino acid sequence of the wild-type PH20 enzyme of SEQ NO. 1, comprising at least 80%, e.g., at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, and at least 99%.

[0061] In some embodiments, the variant or fragment hyaluronidase PH20 comprises, but is not limited to, one or more amino acid residue substitutions at one or more positions selected from T341 to N363, such as T341, L342, S343, 1344, M345, S347, M348, K349, L352, L353, L354, D355, N356, E359, 1361 and N363. For example, amino acid residue substitutions at one or more positions selected from T341, L342, S343, 1344, M345, S347, M348, K349, L352, L353, L354, D355, N356, E359, 1361 and N363 may be, but are not limited to, one or more amino acid residue substitutions selected from T341A, T341C, T341D, T341G, T341S, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, D355K, E359D, I361T and N363G. In an embodiment, an amino acid residue substitution is described by a code number and letter such as "T455S", which means that at number position 455 of a given sequence number, the amino acid residue threonine ("T") is substituted with the amino acid residue serine ("S"). In a specific embodiment, the variant or fragment hyaluronidase PH20 comprises one or more amino acid residue substitutions selected from T341S, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D, and I361T.In certain embodiments, the variant or fragment hyaluronidase PH20 comprises one or more amino acid residue substitutions selected from M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D, and I361T, and may further comprise one or more amino acid residue substitutions selected from the group consisting of T341A, T341C, T341D, T341G, T341S, L342W, S343E, I344N, and N363G, but is not limited thereto.

[0062] In some embodiments, variant or fragment hyaluronidase PH20 is (a) T341S, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D and I361T; (b) L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D and I361T; (c) M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D, I361T, and N363G; (d) T341G, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D, and I361T; (e) T341A, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D, and I361T; (f) T341C, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D, and I361T; (g) T341D, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D, and I361T; (h) I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D and I361T; and (i) one or more amino acid residue substitutions selected from S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D and I361T.

[0063] In some embodiments, the variant or fragment hyaluronidase PH20 comprises a cleavage of the amino acid sequence of SEQ ID NO. 1 prior to F38 at the N-terminus. In some cases, the variant or fragment of hyaluronidase PH20 comprises a cleavage of the amino acid sequence of SEQ ID NO. 1 prior to amino acid residues selected from M1 to P42. For example, the variant or fragment of hyaluronidase PH20 comprises a cleavage at the N-terminus prior to amino acid residues L36, N37, F38, R39, A40, P41, or P42 such that one or more amino acid residues are deleted at the N-terminus. In the embodiments, the phrase "cleavage prior to amino acid residues selected from M1 to P42 at the N-terminus" means that the amino acid residue immediately prior to amino acid residues M1 to P42 at the N-terminus is cleaved and deleted.

[0064] In some embodiments, a variant or fragment of hyaluronidase PH20 comprises a cleavage of the amino acid sequence of SEQ ID NO. 1 following an amino acid selected from V455 to S490. For example, a variant or fragment of hyaluronidase PH20 comprises a cleavage of the amino acid sequence of SEQ ID NO. 1 following amino acid residues V455, C458, D461, C464, I465, D466, A467, F468, K470, P471, P472, M473, E474, T475, E476, E477, P478, Q479, I480, F481, Y482, N483, A484, P486, T488, or S490 such that one or more amino acid residues are deleted at the C-terminus. In an embodiment, the phrase "cleavage after an amino acid residue selected from V455 to S490 at the C-terminus" means that an amino acid residue immediately following amino acid residues V455 to S490 at the C-terminus is cleaved and deleted.

[0065] In some embodiments, a variant or fragment of hyaluronidase PH20 may have the amino acid sequence of SEQ ID NO. 1 and may include one or more amino acid residue substitutions selected from T341S, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D, and I361T, and may include a cleavage up to F38 at the N-terminus and a cleavage down to F468 at the C-terminus (HP46, SEQ ID NO. 99, see Table 2 below). In some cases, a variant or fragment of hyaluronidase PH20 may have the amino acid sequence of SEQ ID NO. 1 and may include one or more amino acid residue substitutions selected from T341A, T341C, T341G, S343E, M345T, K349E, L353A, L354I, N356E, and I361T. In certain cases, one or more amino acid residue substitutions are located in the alpha-helix 8 region (S347 to C381) and / or the linker region (A333 to R346) between alpha-helix 7 and alpha-helix 8. For example, amino acid substitution in the linker region between alpha-helix 7 and alpha-helix 8 may include substitution of one or more amino acid residues in the region composed of amino acid residues T341 to N363, T341 to 1361, L342 to I361, L342 to I361, S343 to I361, I344 to I361, M345 to I361, or M345 to N363.

[0066] In some embodiments, a variant or fragment of hyaluronidase PH20 may be substituted with some amino acid residues of the amino acid sequence of the human HYAL1 region as presented in Tables 2 and 3 having SEQ ID NO. 2, having one or more amino acid residue substitutions located in the alpha-helix 8 region (S347 to C381) and / or the linker region (A333 to R346) between alpha-helix 7 and alpha-helix 8 of wild-type PH20 (e.g., mature wild-type PH20).

[0067]

[0068]

[0069] In certain embodiments, a variant or fragment of hyaluronidase PH20 has an amino acid sequence selected from sequence numbers 60 to 115 as presented in Table 4.

[0070]

[0071]

[0072]

[0073]

[0074]

[0075]

[0076]

[0077]

[0078]

[0079]

[0080]

[0081]

[0082]

[0083]

[0084]

[0085]

[0086]

[0087]

[0088]

[0089]

[0090]

[0091]

[0092]

[0093]

[0094]

[0095]

[0096]

[0097] In some embodiments, the hyaluronidase variant or fragment is a hyaluronidase enzyme comprising, instead of a signal peptide of wild-type PH20 consisting of M1 to T35 but not limited thereto, a human growth hormone signal peptide having the amino acid sequence MATGSRTSLLLAFGLLCLPWLQEGSA of SEQ No. 3 at the N-terminus as shown in Table 5, a human serum albumin signal peptide having the amino acid sequence MKWVTFISLLFLFSSAYS of SEQ No. 4, or a human Hyal1 signal peptide having the amino acid sequence MAAHLLPICALFLTLLDMAQG of SEQ No. 5. In the embodiments, the signal peptide of wild-type PH20 (amino acid residues M1 to T35) is partially or completely deleted. In some embodiments, when a portion of the N-terminus is additionally deleted, for example, when cleavage occurs before N37, F38, R39, A40, P41, or P42 residues, additional deletion of the N-terminus occurs along with deletion of the wild-type PH20 signal peptide.

[0098]

[0099] In some embodiments, the variants or fragments of hyaluronidase include a variant having a C-terminus with a 6×His-tag (indicated as HM), a variant without a 6×His-tag (indicated as HP), a mature wild-type PH20 (L36-S490) having a C-terminus with a 6×His-tag (indicated as WT), and a mature wild-type PH20 (L36 to Y482) having a C-terminus without a 6×His-tag and cleavage occurring after Y482 (indicated as HW2).

[0100] In certain embodiments, the composition comprises one or more variants or fragments of a hyaluronidase enzyme, such as those described in European Patent Publication No. EP3636752A1 and International Patent Publication No. WO 2020 / 197230, the disclosures of which are incorporated herein by reference.

[0101] In an embodiment, the amount of hyaluronidase enzyme present in the composition may vary and may be a hyaluronidase enzyme comprising 100 U or more, e.g., 250 U or more, e.g., 500 U or more, e.g., 750 U or more, e.g., 1000 U or more, e.g., 1500 U or more, e.g., 2000 U or more, e.g., 2500 U or more, e.g., 3000 U or more, e.g., 3500 U or more, e.g., 4000 U or more, e.g., 4500 U or more, e.g., 5000 U or more, e.g., 10,000 U or more, e.g., 20,000 U or more, e.g., 30,000 U or more, e.g., 40,000 U or more, and 50,000 U or more. In some embodiments, the composition comprises about 500 U of hyaluronidase enzyme, about 600 U, about 700 U, about 800 U, about 900 U, about 1,000 U, about 1,100 U, about 1,200 U, about 1,300 U, about 1,400 U, about 1,500 U, about 1,600 U, about 1,700 U, about 1,800 U, about 1,900 U, about 2,000 U, about 2,100 U, about 2,200 U, about 2,300 U, about 2,400 U, about 2,500 U, about 2,600 U, about 2,700 U, about 2,800 U, about 2,900 U, about 3,000 U, about 3,100 U, It contains about 3,200 U, about 3,300 U, about 3,400 U, about 3,500 U, about 3,600 U, about 3,700 U, about 3,800 U, about 3,900 U, about 4,000 U, about 4,100 U, about 4,200 U, about 4,300 U, about 4,400 U, about 4,500 U, about 4,600 U, about 4,700 U, about 4,800 U, about 4,900 U, or about 5,000 U of hyaluronidase enzyme.For example, the amount of hyaluronidase enzyme in the composition may be a range of hyaluronidase enzyme including 50 U to 50,000 U, e.g. 100 U to 45,000 U, e.g. 250 U to 40,000 U, e.g. 500 U to 35,000 U, e.g. 750 U to 30,000 U, e.g. 1000 U to 25,000 U, e.g. 1500 U to 20,000 U, e.g. 2000 U to 15,000 U, e.g. 2500 U to 10,000 U, and 3000 U to 5000 U. For example, the amount of hyaluronidase enzyme in the composition may be a range of hyaluronidase enzymes including 50 U to 50,000 U, e.g. 100 U to 40,000 U, e.g. 300 U to 30,000 U, e.g. 500 U to 20,000 U, e.g. 700 U to 10,000 U, e.g. 800 U to 5,000 U, e.g. 900 U to 4,000 U, e.g. 1,000 U to 3,000 U, e.g. 1,500 U to 2,500 U, and 1,700 U to 2,200 U.

[0102] In certain embodiments, the concentration of hyaluronidase enzyme in the composition is 50 U / mL or more, e.g. 100 U / mL or more, e.g. 250 U / mL or more, e.g. 500 U / mL or more, e.g. 750 U / mL or more, e.g. 1000 U / mL or more, e.g. 2000 U / mL or more, e.g. 2500 U / mL or more, e.g. 3000 U / mL or more, e.g. 3500 U / mL or more, e.g. 4000 U / mL or more, e.g. 4500 U / mL or more, e.g. 5000 U / mL or more. In some cases, the concentration of hyaluronidase enzyme is approximately 100 U / mL, 200 U / mL, 300 U / mL, 400 U / mL, 500 U / mL, approximately 600 U / mL, approximately 700 U / mL, approximately 800 U / mL, approximately 900 U / mL, approximately 1,000 U / mL, approximately 1,100 U / mL, approximately 1,200 U / mL, approximately 1,300 U / mL, approximately 1,400 U / mL, approximately 1,500 U / mL, approximately 1,600 U / mL, approximately 1,700 U / mL, approximately 1,800 U / mL, approximately 1,900 U / mL, approximately 2,000 U / mL, approximately 2,100 U / mL, approximately 2,200 U / mL, approximately 2,300 U / mL, approximately 2,400 U / mL, about 2,500 U / mL, about 2,600 U / mL, about 2,700 U / mL, about 2,800 U / mL, about 2,900 U / mL, about 3,000 U / mL, about 3,100 U / mL, about 3,200 U / mL, about 3,300 U / mL, about 3,400 U / mL, about 3,500 U / mL, about 3,600 U / mL, about 3,700 U / mL, about 3,800 U / mL, about 3,900 U / mL, about 4,000 U / mL, about 4,100 U / mL, about 4,200 U / mL, about 4,300 U / mL, about 4,400 U / mL, about 4,500 U / mL, approx. It is 4,600 U / mL, about 4,700 U / mL, about 4,800 U / mL, about 4,900 U / mL, or about 5,000 U / mL.For example, the concentration of hyaluronidase enzyme in the composition may be in a range of 50 U / mL to 5000 U / mL, e.g. 100 U / mL to 4500 U / mL, e.g. 250 U / mL to 4000 U / mL, e.g. 500 U / mL to 3500 U / mL, e.g. 750 U / mL to 3000 U / mL, 1000 U / mL to 2000 U / mL, and 1500 U / mL to 2500 U / mL.

[0103] The composition according to the embodiment also comprises an oligonucleotide and a telomerase inhibitor having a lipid moiety connected to the 5' and / or 3' ends of the oligonucleotide. In the embodiments, the term "telomerase inhibitor" as used herein refers to a compound capable of reducing or inhibiting the enzymatic activity of telomerase reverse transcriptase in mammalian cells. Telomerase inhibitors of interest include, in some cases, hTR template inhibitors comprising an oligonucleotide. An "hTR template inhibitor" is a compound capable of blocking the template region of the RNA component of human telomerase and inhibiting the activity of the enzyme. In some embodiments, the oligonucleotide comprises a sequence effective for hybridizing to a more specific portion of this region having the sequence 5'-CUAACCCUAAC-3'.

[0104] Telomerase inhibitors of interest comprise oligonucleotides and lipid moieties linked to the 5' and / or 3' ends of the oligonucleotides. In some embodiments, telomerase inhibitors comprise oligonucleotides having "nuclease-resistant linkages" having a backbone comprising subunit linkages that are substantially resistant to nuclease cleavage in a non-hybridized or hybridized form by extracellular and intracellular nucleases. In some cases, the oligonucleotides exhibit little to no nuclease cleavage under physiological conditions.

[0105] In some embodiments, the region of the therapeutic oligonucleotide targeted to the hTR sequence is complementary to the corresponding hTR sequence. In certain embodiments, the base sequence of the oligonucleotide comprises a sequence of five or more nucleotides complementary to the hTR target, e.g., eight or more nucleotides complementary to the hTR target, e.g., ten or more nucleotides, e.g., twelve or more nucleotides, e.g., fifteen or more nucleotides. In certain embodiments, the oligonucleotide of the telomerase inhibitor of the present disclosure is fully complementary to the hTR target sequence, such as when the entire length of the oligonucleotide is complementary to the hTR target sequence.

[0106] Telomerase inhibitors comprise internucleoside linkages such as phosphodiesters, phosphotriesters, methyl phosphonates, P3' → N5' phosphoramidate, N3' → P5' phosphoramidate, N3' → P5' thiophosphoramidate, and phosphorothioate linkages. In certain embodiments, the telomerase inhibitor of interest comprises at least one N3' → P5' phosphoramidate (NP) or N3' → P5' thiophosphoramidate (NPS) linkage, which may be represented by the following structure: 3'-(-NH--P(=O)(--XR)--O-)-5' (where X is O or S and R is selected from the group consisting of hydrogen, alkyl, and aryl); and a pharmaceutically acceptable salt thereof (where XR is OH or SH). In other embodiments, the oligonucleotide includes all NPs or, in some embodiments, all NPS links.In one embodiment, the sequence for the hTR template inhibitor oligonucleotide is a sequence complementary to nucleotides 42–54 of SEQ ID NO. 6. In certain embodiments, the oligonucleotide comprises a sequence complementary or nearly complementary to a portion of the 11-nucleotide region having the sequence 5'-CUAACCCUAAC-3'. The oligonucleotide having this sequence (TAGGGTTAGACAA; SEQ ID NO. 17) and an N3' → P5' thiophosphoramidate (NPS) linkage is designated herein as GRN163. For example, the literature (Asai et al., Cancer Research 63:3931-3939 (2003) and Gryaznov et al., Nucleosides Nucleotides Nucleic Acids Refer to 22(5-8):577-81 (2003). Another target region is the region spanning nucleotides 137–179 of hTR (Pruzan et al., Nucl. Acids Research (Refer to , 30:559-568, 2002). Within this region, the sequence spanning 141–153 is a preferred target. PCT Publication WO 98 / 28442 describes the use of an oligonucleotide of at least 7 nucleotides in length to inhibit telomerase, wherein the oligonucleotide is designed to be complementary to an accessible portion of the hTR sequence outside the template region, including nucleotides 137–196, 290–319, and 350–380 of hTR. A preferred hTR targeting sequence is given below and identified as SEQ ID NO. 7–27. In certain embodiments, the oligonucleotide of the telomerase inhibitor has a sequence targeting human telomerase RNA (hTR), comprising, but not limited to, the following sequence:

[0107]

[0108] The telomerase inhibitors of the present disclosure comprise a lipid moiety attached to the 5' and / or 3' ends of an oligonucleotide. In some cases, the structured group provides superior cellular uptake characteristics, allowing equivalent biological effects to be achieved using a smaller amount of conjugated oligonucleotide compared to the unmodified form. The lipid moiety may be an aliphatic hydrocarbon or fatty acid, such as derivatives of hydrocarbons and fatty acids. For example, the lipid moiety may be a saturated straight-chain compound having 14 to 20 carbons, such as myristic acid (tetradecanoic acid), palmitic acid (hexadecanoic acid), and stearic acid (octadecanoic acid), and their corresponding aliphatic hydrocarbon forms, tetradecane, hexadecane, and octadecane. Other examples of lipid moiety include sterols such as cholesterol, substituted fatty acids, and hydrocarbons, particularly polyfluorinated forms of these groups. In certain embodiments, the lipid moiety comprises one or more derivatives, such as amine, amide, ester, and carbamate derivatives of the lipid moiety. In one example, the lipid moiety is a palmitoyl (C16) moiety, such as palmitoyl amide. The lipid moiety may be conjugated to an oligonucleotide through a linker, such as a glycerol or aminoglycerol linker.

[0109] In some embodiments, the telomerase inhibitor is a compound as described in U.S. Patent No. 9,375,485, the disclosure of which is incorporated herein by reference. In certain embodiments, the telomerase inhibitor is imetelstat (a 5' palmitoylated 13-mer thiophosphoramidate oligonucleotide consisting of sequence 5'-TAGGGTTAGACAA-3' sequence No. 17) or a pharmaceutically acceptable salt thereof, such as imetelstat sodium:

[0110]

[0111] In some embodiments, the amount of a telomerase inhibitor, such as imetelstat or imetelstat sodium, in the subcutaneous composition is about 0.5 to about 5 mg, about 5 to about 10 mg, about 10 to about 15 mg, about 15 to about 20 mg, about 20 to about 25 mg, about 20 to about 50 mg, about 25 to about 50 mg, about 50 to about 75 mg, about 50 to about 100 mg, about 75 to about 100 mg, about 100 to about 125 mg, about 125 to about 150 mg, about 150 to about 175 mg, about 175 to about 200 mg, about 200 to about 225 mg, about 225 to about 250 mg, about 250 to about 300 mg, about 300 to about 350 mg, about 350 to about 400 mg, about 400 to about 450 mg, or about 450 to about 500 mg, about 500 mg to about 600 mg, about 600 mg to about 700 mg, about 700 mg to about 800 mg, about 800 mg to about 900 mg, about 900 mg to about 1000 mg, about 1100 mg to about 1200 mg, about 1200 mg to about 1300 mg, about 1300 mg to about 1400 mg, about 1400 mg to about 1500 mg, about 1500 mg to about 1600 mg, about 1600 mg to about 1700 mg, about 1700 mg to about 1800 mg, about 1800 mg to about 1900 mg, about It is 1900 mg to about 2000 mg, about 2000 mg to about 2100 mg, about 2100 mg to about 2200 mg, about 2200 mg to about 2300 mg, about 2300 mg to about 2400 mg, and about 2400 mg to about 2500 mg.

[0112] In some embodiments, the amount of telomerase inhibitor is a unit dosage form having an amount ranging from about 5 mg to about 1000 mg, 5 mg to about 500 mg, e.g., about 30 mg to about 300 mg, or about 50 mg to about 200 mg. In some embodiments, the amount of telomerase inhibitor is a unit dosage form having an amount ranging from about 200 mg to about 3000 mg, 750 mg to about 2500 mg, e.g., about 1000 mg to about 2000 mg, or about 500 mg to about 2000 mg. The unit dosage form may be liquid or lyophilized.

[0113] In some embodiments, the concentration of the telomerase inhibitor in the composition is, for example, about 0.1 to about 300 mg / ml, 0.1 to about 200 mg / ml, about 0.1 to about 180 mg / ml, about 0.1 to about 160 mg / ml, about 0.1 to about 140 mg / ml, about 0.1 to about 120 mg / ml, about 0.1 to about 100 mg / ml, about 0.1 to about 80 mg / ml, about 0.1 to about 60 mg / ml, about 0.1 to about 40 mg / ml, about 0.1 to about 20 mg / ml, about 0.1 to about 10 mg / ml, about 2 to about 40 mg / ml, about 4 to about 35 mg / ml, about 6 to about 30 mg / ml, about 8 to about 25 mg / ml, about 10 to about It is diluted (about 0.1 mg / ml) or concentrated (about 300 mg / ml) to include either 20 mg / ml, about 12 to about 15 mg / ml, or any of about 0.1 mg / ml, 0.2 mg / ml, 0.3 mg / ml, 0.4 mg / ml, 0.5 mg / ml, 0.6 mg / ml, 0.7 mg / ml, 0.8 mg / ml, 0.9 mg / ml, 1 mg / ml, 1.1 mg / ml, 1.2 mg / ml, 1.3 mg / ml, 1.4 mg / ml, 1.5 mg / ml, 1.6 mg / ml, 1.7 mg / ml, 1.8 mg / ml, 1.9 mg / ml, 2 mg / ml, 2.1 mg / ml, 2.2 mg / ml, 2.3 mg / ml, 2.4 mg / ml, or 2.5 mg / ml. mg / ml). In some embodiments, the concentration of the telomerase inhibitor is at least about 0.1 mg / ml, 0.2 mg / ml, 0.3 mg / ml, 0.4 mg / ml, 0.5 mg / ml, 1.3 mg / ml, 1.5 mg / ml, 2 mg / ml, 3 mg / ml, 4 mg / ml, 5 mg / ml, 6 mg / ml, 7 mg / ml, 8 mg / ml, 9 mg / ml, 10 mg / ml, 11 mg / ml, 12 mg / ml, 13 mg / ml, 14 mg / ml, 15 mg / ml, 16 mg / ml, 17 mg / ml, 18 mg / ml, 19 mg / ml, 20 mg / ml, 21 mg / ml, 22 mg / ml, 23 mg / ml, 24 mg / ml, 25 mg / ml, 26 mg / ml, 27 mg / ml, 28 mg / ml, 29 mg / ml, 30 mg / ml, 31 mg / ml, 32 mg / ml, 33 mg / ml, 33.3 mg / ml, 34 mg / ml, 35 mg / ml, 36 mg / ml, 37 mg / ml, 38 mg / ml, 39 mg / ml, 40 mg / ml, 50 mg / ml, 60 mg / ml, 70 mg / ml, 80 mg / ml, 90 mg / ml, 100 mg / ml, 110 mg / ml, 120 mg / ml, 130 mg / ml, 140 mg / ml, 150 mg / ml, 160 mg / ml, 170 mg / ml, 180 mg / ml, 190 mg / ml, 200 mg / ml, 210 mg / ml, 220 mg / ml, 230 mg / ml, 240 mg / ml, or 250 mg / ml, 260 mg / ml, 270 mg / ml, 280 mg / ml, 290 mg / ml, 300 mg / ml.

[0114] In certain embodiments, the composition is formulated to include a telomerase inhibitor such as imetelstat or imetelstat sodium in a dosage range including about 2.0 mg / kg to about 20.0 mg / kg, e.g., about 3.0 mg / kg to about 15.0 mg / kg, e.g., about 4.0 mg / kg to about 10 mg / kg, e.g., about 6 mg / kg to about 14 mg / kg, e.g., about 7 mg / kg to about 13 mg / kg, e.g., about 8 mg / kg to about 12 mg / kg, e.g., about 7.5 mg / kg to 9.4 mg / kg, e.g., about 9 mg / kg to about 11 mg / kg, and about 11 mg / kg to about 14 mg / kg. In some embodiments, the composition is formulated to contain a telomerase inhibitor, such as imetelstat or imetelstat sodium, in a dosage range of about 7.5 mg / kg to about 9.4 mg / kg. For example, the dosage of telomerase inhibitors is 4.0 mg / kg, 4.1 mg / kg, 4.2 mg / kg, 4.3 mg / kg, 4.4 mg / kg, 4.5 mg / kg, 4.6 mg / kg, 4.7 mg / kg, 4.8 mg / kg, 4.9 mg / kg, 5.0 mg / kg, 5.1 mg / kg, 5.2 mg / kg, 5.3 mg / kg, 5.4 mg / kg, 5.5 mg / kg, 5.6 mg / kg, 5.7 mg / kg, 5.8 mg / kg, 5.9 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, 7.1 mg / kg, 7.2 mg / kg, 7.3 mg / kg, 7.4 mg / kg, 7.5 mg / kg, 7.6 mg / kg, 7.7 mg / kg, 7.8 mg / kg, 7.9 mg / kg, 8 mg / kg, 8.1 mg / kg, 8.2 mg / kg, 8.3 mg / kg, 8.4 mg / kg, 8.5 mg / kg, 8.6 mg / kg, 8.7 mg / kg, 8.8 mg / kg, 8.9 mg / kg, 9 mg / kg, 9.1 mg / kg, 9.2 mg / kg, 9.3 mg / kg, 9.4 mg / kg, 9.5 mg / kg, 9.6 mg / kg, 9.7 mg / kg, 9.8 mg / kg, 9.9 mg / kg, 10 mg / kg, 10.1 mg / kg, 10.2 mg / kg, 10.3 mg / kg, 10.4 mg / kg, 10.5 mg / kg, 10.6 mg / kg, 10.7 mg / kg, 10.8 mg / kg, 10.9 mg / kg, 11 mg / kg, 11.1 mg / kg, 11.2 mg / kg, 11.3 mg / kg, 11.4 mg / kg, 11.5 mg / kg, 11.6 mg / kg, 11.7 mg / kg, 11.8 mg / kg, 11.9 mg / kg, 12 mg / kg, 12.1 mg / kg, 12.2 mg / kg, 12.3 mg / kg, 12.4 mg / kg, 12.5 mg / kg, 12.6 mg / kg, 12.7 mg / kg, 12.8 mg / kg, 12.9 mg / kg, 13 mg / kg, 10.5 mg / kg, 11.0 mg / kg, 11.5 mg / kg, 12.0 mg / kg, 12.5 mg / kg, 13.0 mg / kg, 13.5 mg / kg, 14.0 mg / kg, 14.5 mg / kg, 15.0 mg / kg, 15.5 mg / kg, 16.0 mg / kg, 16.5 mg / kg, 17.0 mg / kg, 17.0 mg / kg, 17.5 mg / kg, 18.0 mg / kg, 18.5 mg / kg, 19.0 mg / kg, 19.5 mg / kg, 또는 20.0 mg / kg일 수 있다.

[0115] In some embodiments, the subcutaneous telomerase inhibitor composition also comprises one or more pharmaceutically acceptable carriers. Exemplary pharmaceutically acceptable carriers may include solvents, dispersion media, coatings, antimicrobial and antifungal agents, isotonic and absorption retardants, aqueous or non-aqueous carriers, or combinations thereof. In certain embodiments, the composition may further comprise one or more pharmaceutically acceptable excipients as part of the pharmaceutical composition. Excipients may include, but are not limited to, carbohydrates, inorganic salts, antimicrobial agents, stabilizers, antioxidants, surfactants, amino acids, buffers, acids, bases, and combinations thereof. For example, excipients suitable for a subcutaneous injection composition may include one or more of water, alcohols, polyols, monosaccharides, polysaccharides, stabilizers, buffers, amino acids, and surfactants. The amount of each pharmaceutically acceptable excipient or carrier may vary and may be in a range including 1 mM to 1000 mM, e.g., 2 mM to 900 mM, e.g., 3 mM to 800 mM, e.g., 4 mM to 700 mM, e.g., 5 mM to 600 mM, e.g., 6 mM to 500 mM, e.g., 7 mM to 400 mM, e.g., 8 mM to 300 mM, e.g., 9 mM to 200 mM, and 10 mM to 100 mM.

[0116] In some embodiments, the composition comprises a buffer. Examples of buffers that may be used are acetic acid, citric acid, formic acid, succinic acid, phosphoric acid, carbonic acid, malic acid, aspartic acid, histidine, boric acid, Tris buffer, HEPPSO, and HEPES. In some cases, the buffer is present in the composition in an amount to maintain the composition at a predetermined pH. For example, one or more buffers may be present in the composition to maintain the composition at a pH of 3.0 to 9.0, e.g., pH 3.5 to 8.5, e.g., pH 4.0 to 8.0, e.g., pH 4.5 to 7.5, e.g., pH 5.0 to 7.0, and pH 5.5 to 7.5. For example, the composition has pH 3.0, pH 3.1, pH 3.2, pH 3.3, pH 3.4, pH 3.5, pH 3.6, pH 3.7, pH 3.8, pH 3.9, pH 4.0, pH 4.1, pH 4.2, pH 4.3, pH 4.4, pH 4.5, pH 4.6, pH 4.7, pH 4.8, pH 4.9, pH 5.0, pH 5.1, pH 5.2, pH 5.3, pH 5.4, pH 5.5, pH 5.6, pH 5.7, pH 5.8, pH 5.9, pH 6.0, pH 6.1, pH 6.2, pH 6.3, pH 6.4, pH 6.5, pH 6.6, pH 6.7, pH 6.8, pH 6.9, pH 7.0, pH 7.1, pH 7.2, pH 7.3, pH 7.4, pH 7.5, pH 7.6, pH 7.7, pH 7.8, pH 7.9, pH 8.0, pH 8.1, pH 8.2, pH 8.3, pH 8.4, pH 8.5, pH 8.6, pH 8.7, pH 8.8, pH It may have a pH of 8.9 or 9.0.The buffer may be present in the composition in amounts including 1 mM to 1000 mM, e.g., 2 mM to 900 mM, e.g., 3 mM to 800 mM, e.g., 4 mM to 700 mM, e.g., 5 mM to 600 mM, e.g., 6 mM to 500 mM, e.g., 7 mM to 400 mM, e.g., 8 mM to 300 mM, e.g., 9 mM to 200 mM, and 10 mM to 100 mM. For example, the buffer is approximately 10 mM, approximately 20 mM, approximately 30 mM, approximately 40 mM, approximately 50 mM, approximately 75 mM, 100 mM, approximately 110 mM, approximately 120 mM, approximately 130 mM, approximately 140 mM, approximately 150 mM, approximately 160 mM, approximately 170 mM, approximately 180 mM, approximately 190 mM, approximately 200 mM, approximately 210 mM, approximately 220 mM, approximately 230 mM, approximately 240 mM, approximately 250 mM, approximately 260 mM, approximately 270 mM, approximately 280 mM, approximately 290 mM, approximately 300 mM, approximately 310 mM, approximately 320 mM, approximately 330 mM, approximately 340 mM, approximately 350 mM, approximately 360 mM, approximately 370 mM, approximately 380 mM, approximately It may be present in the composition at a concentration of 390 mM, about 400 mM, about 410 mM, about 420 mM, about 430 mM, about 440 mM, about 450 mM, about 460 mM, about 470 mM, about 480 mM, about 490 mM, or about 500 mM.

[0117] In some embodiments, the composition comprises carbohydrates such as sugars. Exemplary sugars include monosaccharides, disaccharides, trisaccharides, polysaccharides, sugar alcohols, reducing sugars, and non-reducing sugars, such as glucose, sucrose, trehalose, lactose, fructose, maltose, dextran, glycerin, dextran, erythritol, glycerol, arabitol, syllitol, sorbitol, mannitol, melivise, melesitose, raffinose, mannnotriose, stachyose, maltose, lactulose, maltulose, glucitol, maltitol, lactitol, or isomaltulose. In some cases, the composition comprises sucrose. In other cases, the composition comprises trehalose. Carbohydrates (sugars such as sucrose or trehalose) may be present in the composition in amounts including 1 mM to 1000 mM, e.g., 2 mM to 900 mM, e.g., 3 mM to 800 mM, e.g., 4 mM to 700 mM, e.g., 5 mM to 600 mM, e.g., 6 mM to 500 mM, e.g., 7 mM to 400 mM, e.g., 8 mM to 300 mM, e.g., 9 mM to 200 mM, and 10 mM to 100 mM.For example, carbohydrates (sugars such as sucrose or trehalose) are approximately 10 mM, approximately 20 mM, approximately 30 mM, approximately 40 mM, approximately 50 mM, approximately 75 mM, 100 mM, approximately 110 mM, approximately 120 mM, approximately 130 mM, approximately 140 mM, approximately 150 mM, approximately 160 mM, approximately 170 mM, approximately 180 mM, approximately 190 mM, approximately 200 mM, approximately 210 mM, approximately 220 mM, approximately 230 mM, approximately 240 mM, approximately 250 mM, approximately 260 mM, approximately 270 mM, approximately 280 mM, approximately 290 mM, approximately 300 mM, approximately 310 mM, approximately 320 mM, approximately 330 mM, approximately 340 mM, approximately 350 mM, approximately 360 It may be present in the composition at a concentration of mM, about 370 mM, about 380 mM, about 390 mM, about 400 mM, about 410 mM, about 420 mM, about 430 mM, about 440 mM, about 450 mM, about 460 mM, about 470 mM, about 480 mM, about 490 mM, or about 500 mM.

[0118] In some embodiments, the composition comprises one or more amino acids. Exemplary amino acids include histidine, isoleucine, methionine, glycine, arginine, lysine, L-leucine, tri-leucine, alanine, glutamic acid, L-threonine, and 2-phenylamine. In some cases, the composition comprises methionine. In other cases, the composition comprises histidine. The amino acid (e.g., methionine or histidine) may be present in the composition in amounts including 0.1 mg / mL to about 5 mg / mL, e.g. 0.1 mg / mL to about 2.5 mg / mL, e.g. 1 mg / mL to about 2 mg / mL, e.g. 4 mM to 700 mM, e.g. 5 mM to 600 mM, e.g. 6 mM to 500 mM, e.g. 7 mM to 400 mM, e.g. 8 mM to 300 mM, e.g. 9 mM to 200 mM, and 10 mM to 100 mM. For example, amino acids (e.g., methionine or histidine) at concentrations of about 0.5 mg / mL, about 1 mg / mL, about 1.1 mg / mL, about 1.2 mg / mL, about 1.3 mg / mL, about 1.4 mg / mL, about 1.5 mg / mL, about 1.6 mg / mL, about 1.7 mg / mL, about 1.8 mg / mL, about 1.9 mg / mL, about 2.0 mg / mL, about 2.1 mg / mL, about 2.2 mg / mL, about 2.3 mg / mL, about 2.4 mg / mL, about 2.5 mg / mL, about 2.6 mg / mL, about 2.7 mg / mL, about 2.8 mg / mL, about 2.9 mg / mL, about 3 mg / mL, about 3.5 mg / mL, about 4 mg / mL, about 4.5 mg / mL, or about 5 mg / mL It may be present in the composition. In certain embodiments, the composition is about 0.5 mg / mL, about 1 mg / mL, about 1.1 mg / mL, about 1.2 mg / mL, about 1.3 mg / mL, about 1.4 mg / mL, about 1.5 mg / mL, about 1.6 mg / mL, about 1.7 mg / mL, about 1.8 mg / mL, about 1.9 mg / mL, about 2.It contains histidine in amounts of 0 mg / mL, about 2.1 mg / mL, about 2.2 mg / mL, about 2.3 mg / mL, about 2.4 mg / mL, about 2.5 mg / mL, about 2.6 mg / mL, about 2.7 mg / mL, about 2.8 mg / mL, about 2.9 mg / mL, about 3 mg / mL, about 3.5 mg / mL, about 4 mg / mL, about 4.5 mg / mL, or about 5 mg / mL. In another embodiment, the composition contains methionine in an amount of about 0.5 mg / mL, about 1 mg / mL, about 1.1 mg / mL, about 1.2 mg / mL, about 1.3 mg / mL, about 1.4 mg / mL, about 1.5 mg / mL, about 1.6 mg / mL, about 1.7 mg / mL, about 1.8 mg / mL, about 1.9 mg / mL, about 2.0 mg / mL, about 2.1 mg / mL, about 2.2 mg / mL, about 2.3 mg / mL, about 2.4 mg / mL, about 2.5 mg / mL, about 2.6 mg / mL, about 2.7 mg / mL, about 2.8 mg / mL, about 2.9 mg / mL, about 3 mg / mL, about 3.5 mg / mL, about 4 mg / mL, about 4.5 mg / mL, or about 5 mg / mL.

[0119] In some embodiments, the composition comprises one or more surfactants. Exemplary surfactants include polysorbate (e.g., polysorbate-20 or polysorbate-80); polyoxamer (e.g., poloxamer 188); Triton; sodium octyl glycoside; lauryl-, myristyl-, linoleyl-, or stearyl-sulfobetaine; lauryl-, myristyl-, linoleyl-, or stearyl-sarcosine; linoleyl-, myristyl-, or cetyl-betaine; lauroamidopropyl-, cocamidopropyl-, linoleamidopropyl-, myristamidopropyl-, palmidopropyl-, or isostearamidopropyl-betaine (e.g., lauroamidopropyl); myristamidopropyl-, palmidopropyl-, or isostearamidopropyl-dimethylamine; Sodium methyl cocoyl- or disodium methyl oleyl-taurate; and MONAQUA™ series (Mona Industries, Inc., Paterson, NJ), polyethyl glycol, polypropyl glycol, and copolymers of ethylene and propylene glycol (e.g., PLURONICS™, PF68, etc.). In some cases, the composition comprises a polysorbate surfactant. The surfactant may be present in the composition in amounts including 1 mM to 1000 mM, e.g., 2 mM to 900 mM, e.g., 3 mM to 800 mM, e.g., 4 mM to 700 mM, e.g., 5 mM to 600 mM, e.g., 6 mM to 500 mM, e.g., 7 mM to 400 mM, e.g., 8 mM to 300 mM, e.g., 9 mM to 200 mM, and 10 mM to 100 mM.For example, the surfactant is about 10 mM, about 20 mM, about 30 mM, about 40 mM, about 50 mM, about 75 mM, 100 mM, about 110 mM, about 120 mM, about 130 mM, about 140 mM, about 150 mM, about 160 mM, about 170 mM, about 180 mM, about 190 mM, about 200 mM, about 210 mM, about 220 mM, about 230 mM, about 240 mM, about 250 mM, about 260 mM, about 270 mM, about 280 mM, about 290 mM, about 300 mM, about 310 mM, about 320 mM, about 330 mM, about 340 mM, about 350 mM, about 360 mM, about 370 mM, about 380 mM, It may be present in the composition at a concentration of about 390 mM, about 400 mM, about 410 mM, about 420 mM, about 430 mM, about 440 mM, about 450 mM, about 460 mM, about 470 mM, about 480 mM, about 490 mM, or about 500 mM.

[0120] The composition may include one or more pharmaceutically acceptable salts. The pharmaceutically acceptable salt is formed from (1) an inorganic acid such as hydrochloric acid, hydrobromide, sulfuric acid, nitric acid, phosphoric acid, etc.; Acetic acid, propionic acid, hexanoic acid, cyclopentanepropionic acid, glycolic acid, pyruvate, lactic acid, malonic acid, succinic acid, malic acid, maleic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, 3(4-hydroxybenzoyl)benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, 1,2-ethanedisulfonic acid, 2-hydroxyethanesulfonic acid, benzenesulfonic acid, 4-chlorobenzenesulfonic acid, 2-naphthalenesulfonic acid, 4-toluenesulfonic acid, camphorsulfonic acid, 4-methylbicyclo[2.2.2]oct-2-en-1-carboxylic acid, glucoheptonic acid, 3-phenylpropionic acid, trimethylacetic acid, tertiary butylacetic acid, lauryl sulfate, gluconic acid, glutamic acid, Acid addition salt formed with an organic acid such as hydroxynaphthoic acid, salicylic acid, stearic acid, muconic acid, etc.; or (2) when an acidic proton present in the compound is replaced by a metal ion such as an alkali metal ion, alkaline earth ion, or aluminum ion; or may be a salt formed when coordinated with an organic base such as ethanolamine, diethanolamine, triethanolamine, N-methylglucarmine, etc. In certain embodiments, the composition comprises other pharmaceutically acceptable salts such as sodium chloride salt or magnesium sulfate. Pharmaceutically acceptable salts may be present in the composition in amounts including 1 mM to 1000 mM, e.g., 2 mM to 900 mM, e.g., 3 mM to 800 mM, e.g., 4 mM to 700 mM, e.g., 5 mM to 600 mM, e.g., 6 mM to 500 mM, e.g., 7 mM to 400 mM, e.g., 8 mM to 300 mM, e.g., 9 mM to 200 mM, and 10 mM to 100 mM.For example, pharmaceutically acceptable salts are approximately 10 mM, approximately 20 mM, approximately 30 mM, approximately 40 mM, approximately 50 mM, approximately 75 mM, 100 mM, approximately 110 mM, approximately 120 mM, approximately 130 mM, approximately 140 mM, approximately 150 mM, approximately 160 mM, approximately 170 mM, approximately 180 mM, approximately 190 mM, approximately 200 mM, approximately 210 mM, approximately 220 mM, approximately 230 mM, approximately 240 mM, approximately 250 mM, approximately 260 mM, approximately 270 mM, approximately 280 mM, approximately 290 mM, approximately 300 mM, approximately 310 mM, approximately 320 mM, approximately 330 mM, approximately 340 mM, approximately 350 mM, approximately 360 mM, approximately 370 mM, approximately It may be present in the composition at a concentration of 380 mM, approximately 390 mM, approximately 400 mM, approximately 410 mM, approximately 420 mM, approximately 430 mM, approximately 440 mM, approximately 450 mM, approximately 460 mM, approximately 470 mM, approximately 480 mM, approximately 490 mM, or approximately 500 mM.

[0121] Acids or bases may also be present in the target composition. For example, acids may include, but are not limited to, hydrochloric acid, acetic acid, phosphoric acid, citric acid, malic acid, lactic acid, formic acid, trichloroacetic acid, nitric acid, perchloric acid, phosphoric acid, sulfuric acid, fumaric acid, and any combination thereof. Examples of bases include, but are not limited to, sodium hydroxide, sodium acetate, ammonium hydroxide, potassium hydroxide, ammonium acetate, potassium acetate, sodium phosphate, potassium phosphate, sodium carbonate monohydrate, sodium citrate, sodium formate, sodium sulfate, potassium sulfate, potassium fumarate, and any combination thereof. The acid or base may be present in the composition at concentrations including 1 mM to 1000 mM, e.g., 2 mM to 900 mM, e.g., 3 mM to 800 mM, e.g., 4 mM to 700 mM, e.g., 5 mM to 600 mM, e.g., 6 mM to 500 mM, e.g., 7 mM to 400 mM, e.g., 8 mM to 300 mM, e.g., 9 mM to 200 mM, and 10 mM to 100 mM.For example, the acid or base is about 10 mM, about 20 mM, about 30 mM, about 40 mM, about 50 mM, about 75 mM, 100 mM, about 110 mM, about 120 mM, about 130 mM, about 140 mM, about 150 mM, about 160 mM, about 170 mM, about 180 mM, about 190 mM, about 200 mM, about 210 mM, about 220 mM, about 230 mM, about 240 mM, about 250 mM, about 260 mM, about 270 mM, about 280 mM, about 290 mM, about 300 mM, about 310 mM, about 320 mM, about 330 mM, about 340 mM, about 350 mM, about 360 mM, about 370 mM, about 380 mM, It may be present in the composition at a concentration of about 390 mM, about 400 mM, about 410 mM, about 420 mM, about 430 mM, about 440 mM, about 450 mM, about 460 mM, about 470 mM, about 480 mM, about 490 mM, or about 500 mM.

[0122] In some embodiments, the composition comprises one or more antioxidants. Antioxidants capable of preventing degradation of the composition by reducing or preventing oxidation may include, for example, ascorbyl palmitate, butylated hydroxyanisole, butylated hydroxytoluene, hypophosphoric acid, methionine, monothioglycerol, propyl gallate, sodium bisulfite, sodium formaldehyde sulfoxylate, sodium metabisulfite, and any combination thereof. Antioxidants may be present in the composition at concentrations including 1 mM to 1000 mM, e.g., 2 mM to 900 mM, e.g., 3 mM to 800 mM, e.g., 4 mM to 700 mM, e.g., 5 mM to 600 mM, e.g., 6 mM to 500 mM, e.g., 7 mM to 400 mM, e.g., 8 mM to 300 mM, e.g., 9 mM to 200 mM, and 10 mM to 100 mM. For example, the antioxidant is approximately 10 mM, approximately 20 mM, approximately 30 mM, approximately 40 mM, approximately 50 mM, approximately 75 mM, 100 mM, approximately 110 mM, approximately 120 mM, approximately 130 mM, approximately 140 mM, approximately 150 mM, approximately 160 mM, approximately 170 mM, approximately 180 mM, approximately 190 mM, approximately 200 mM, approximately 210 mM, approximately 220 mM, approximately 230 mM, approximately 240 mM, approximately 250 mM, approximately 260 mM, approximately 270 mM, approximately 280 mM, approximately 290 mM, approximately 300 mM, approximately 310 mM, approximately 320 mM, approximately 330 mM, approximately 340 mM, approximately 350 mM, approximately 360 mM, approximately 370 mM, approximately 380 mM, approximately It may be present in the composition at a concentration of 390 mM, about 400 mM, about 410 mM, about 420 mM, about 430 mM, about 440 mM, about 450 mM, about 460 mM, about 470 mM, about 480 mM, about 490 mM, or about 500 mM.

[0123] In some embodiments, the composition comprises one or more preservatives. Preservatives capable of reducing or preventing degradation of the composition, such as that caused by microbial growth, may include, for example, antioxidants, antimicrobial agents and chelating agents, and may include methyl, ethyl, propyl and butyl parabens, aryl and alkyl acids, citric acid, sorbic acid, sodium sorbate, K & Ca, benzoic acid, sodium benzoate, K & Ca, benzyl alcohol, sodium metabisulfite, bronopol, propylene glycol (15-30%), BHT (butylated hydroxytoluene), BHA (butylated hydroxyanisole), propyl gallate, EDTA, chlorobutanol, benzaldehyde, phenol, metacresol, chlorocresol, benzylkonium chloride, benzethonium chloride, and mercury compounds such as thiomersal and phenylmercury nitrate, and any combination thereof. The preservative may be present in the composition at concentrations including 1 mM to 1000 mM, e.g., 2 mM to 900 mM, e.g., 3 mM to 800 mM, e.g., 4 mM to 700 mM, e.g., 5 mM to 600 mM, e.g., 6 mM to 500 mM, e.g., 7 mM to 400 mM, e.g., 8 mM to 300 mM, e.g., 9 mM to 200 mM, and 10 mM to 100 mM.For example, the preservative is approximately 10 mM, approximately 20 mM, approximately 30 mM, approximately 40 mM, approximately 50 mM, approximately 75 mM, 100 mM, approximately 110 mM, approximately 120 mM, approximately 130 mM, approximately 140 mM, approximately 150 mM, approximately 160 mM, approximately 170 mM, approximately 180 mM, approximately 190 mM, approximately 200 mM, approximately 210 mM, approximately 220 mM, approximately 230 mM, approximately 240 mM, approximately 250 mM, approximately 260 mM, approximately 270 mM, approximately 280 mM, approximately 290 mM, approximately 300 mM, approximately 310 mM, approximately 320 mM, approximately 330 mM, approximately 340 mM, approximately 350 mM, approximately 360 mM, approximately 370 mM, approximately 380 mM, approximately It may be present in the composition at a concentration of 390 mM, about 400 mM, about 410 mM, about 420 mM, about 430 mM, about 440 mM, about 450 mM, about 460 mM, about 470 mM, about 480 mM, about 490 mM, or about 500 mM.

[0124] Pharmaceutical excipients, along with other excipients that may be used in the target telomerase inhibitor composition, are described in the literature (A. Gennaro (2000) "Remington: The Science and Practice of Pharmacy", 20th edition, Lippincott, Williams, & Wilkins; Pharmaceutical Dosage Forms and Drug Delivery Systems (1999) HC Ansel et al., eds 7th ed., Lippincott, Williams, & Wilkins; and Handbook of Pharmaceutical Excipients (2000) AH Kibbe et al., eds., 3rd ed. Amer. Pharmaceutical Assoc.), the disclosures of which are incorporated herein by reference.

[0125] Method of subcutaneously administering a telomerase inhibitor composition

[0126] Aspects of the present disclosure also include a method for subcutaneously administering a telomerase inhibitor composition to a subject. In carrying out the method according to a specific embodiment, one or more compositions as described herein having a telomerase inhibitor and a hyaluronidase enzyme are subcutaneously administered to a subject. In some embodiments, the composition is administered to the subject by subcutaneous injection or subcutaneous infusion. In other embodiments, the composition may be administered to the subject from an implanted device, such as a subcutaneous implanted catheter. In a specific embodiment, the telomerase inhibitor composition is administered to the subject using a subcutaneous bolus injection device configured to deliver a predetermined amount of the composition subcutaneously to the subject.

[0127] In some embodiments, the method comprises the step of subcutaneously administering to a subject one or more compositions as described herein having a telomerase inhibitor and a hyaluronidase enzyme to treat a neoplasm. In some embodiments, the neoplasm may be a solid tumor cancer. Examples of cancers for treatment according to embodiments of the present disclosure include, for example, adrenocortical carcinoma, anal cancer, appendicitis, astrocytoma, atypical anomaly / rhabdomyotum, basal cell carcinoma, cholangiocarcinoma (extrahepatic), bladder cancer, bone cancer (e.g., Ewing sarcoma, osteosarcoma and malignant fibrous histiocytoma, etc.), brainstem glioma, brain tumor (e.g., astrocytoma, central nervous system embryonic tumor, central nervous system germ cell tumor, craniopharyngioma, ependymoma, etc.), breast cancer (e.g., female breast cancer, male breast cancer, pediatric breast cancer, etc.), bronchial tumor, carcinoid tumor (e.g., pediatric, gastrointestinal, etc.), unknown primary carcinoma, heart tumor, cervical cancer, colon cancer, colorectal cancer, craniopharyngioma, duct (e.g., bile duct, extrahepatic, etc.), intraductal carcinoma in situ (DCIS), embryonic tumor, endometrial cancer, ependymoma, esophageal cancer. Sensory neuroblastoma, Ewing sarcoma, extracranial germ cell tumor, extragonadal germ cell tumor, extrahepatic cholangiocarcinoma, ocular cancer (e.g., intraocular melanoma, retinoblastoma, etc.), fibrous histiocytoma of bone (malignant, osteosarcoma, etc.), gallbladder cancer, gastric cancer, gastrointestinal carcinoid tumor, gastrointestinal stromal tumor (GIST), germ cell tumor (e.g., extracranial, extragonadal, ovarian, testicular, etc.), gestational trophoblastic disease, glioma, hairy cell leukemia, head and neck cancer, heart cancer, hepatocellular carcinoma, histiocytosis (e.g., Langerhans cell, etc.), hypopharyngeal cancer, intraocular melanoma, islet cell tumor (e.g., pancreatic neuroendocrine tumor, etc.), renal cancer (e.g., renal cell, Wilms tumor, pediatric renal tumor, etc.), Langerhans cell histiocytosis, laryngeal cancer, lip and oral cancer, liver cancer (primary), lobular carcinoma in situ (LCIS), lung cancer (e.g. For example, non-small cell, small cell, etc.), malignant fibrous histiocytoma and osteosarcoma of bone, melanoma,Merkel cell carcinoma, mesothelioma, latent primary metastatic squamous cell cervical cancer, oral cancer, multiple endocrine neoplasm syndrome, nasal and paranasal sinus cancer, nasopharyngeal cancer, neuroblastoma, non-small cell lung cancer, oral cancer, oral cancer (e.g., lips, etc.), oropharyngeal cancer, osteosarcoma and malignant fibrous histiocytoma of bone, ovarian cancer (e.g., epithelial, germ cell tumors, tumors with low likelihood of malignancy, etc.), pancreatic cancer, pancreatic neuroendocrine tumor (island cell tumor), papillomatosis, paraganglioma, paranasal and nasal sinus cancer, parathyroid cancer, penile cancer, pharyngeal cancer, pheochromocytoma, pituitary tumor, pleural-pulmonary blastoma, prostate cancer, rectal cancer, renal cell (kidney) cancer, renal pelvis and ureter, transitional cell carcinoma, retinoblastoma, rhabdomyosarcoma, salivary gland cancer, Sézary syndrome, skin cancer (e.g., pediatric, melanoma, Merkel cell carcinoma, may include, but is not limited to, non-melanoma, etc.), small cell lung cancer, small intestine cancer, soft tissue sarcoma, squamous cell carcinoma, squamous cell cervical cancer (e.g., latent primary, metastatic, etc.), gastric (stomach) cancer, testicular cancer, throat cancer, thymomas and thymic carcinomas, thyroid cancer, transitional cell carcinoma of the renal pelvis and ureter, ureteral and renal pelvis cancer, urethral cancer, uterine cancer (e.g., endometrial, etc.), uterine sarcoma, vaginal cancer, vulvar cancer, Waldenström macroglobulinemia, Wilms tumor, etc. In certain embodiments, the method comprises the step of treating a subject having a neoplasm as described in U.S. Patent No. 7,494,982.

[0128] In some embodiments, the method comprises the step of subcutaneously administering to a subject one or more compositions as described herein, comprising a telomerase inhibitor and a hyaluronidase enzyme, to treat a hematological neoplasm. In some cases, treating the hematological neoplasm involves inducing apoptosis of hematological neoplasm cells, such as inducing apoptosis of hematological neoplasm cells in vitro. In other cases, treating the hematological neoplasm involves inducing apoptosis of hematological neoplasm cells in a subject. In some embodiments, the hematological neoplasm cells are malignant hematopoietic stem cells (HSCs). In other embodiments, the hematological neoplasm cells are malignant hematopoietic progenitor cells (HPCs).

[0129] In some embodiments, the method comprises the step of subcutaneously administering to a subject one or more compositions as described herein, comprising a telomerase inhibitor and a hyaluronidase enzyme, to treat a myeloproliferative neoplasm. In some cases, treating the myeloproliferative neoplasm involves inducing apoptosis of myeloproliferative neoplasm cells, such as inducing apoptosis of myeloproliferative neoplasm cells in vitro. In other cases, treating the myeloproliferative neoplasm involves inducing apoptosis of myeloproliferative neoplasm cells in a subject. In some embodiments, the myeloproliferative neoplasm cells are malignant hematopoietic stem cells (HSCs). In other embodiments, the myeloproliferative neoplasm cells are malignant hematopoietic progenitor cells (HPCs). The myeloproliferative neoplasm treated according to the target method may include myelofibrosis (MF), such as primary myelofibrosis, for example, or myelofibrosis after prior ET or PV (ET-post-MF or PV-post-MF). In other embodiments, myeloproliferative neoplasms include essential thrombocytosis (ET), polycythemia vera (PV), chronic myeloid leukemia (CML), chronic neutrophilic leukemia, chronic eosinophilic leukemia, and acute myeloid leukemia (AML).

[0130] In another embodiment, the hematological neoplasm is myelodysplastic syndrome (MDS). In yet another embodiment, the hematological neoplasm is myelodysplastic syndrome (MDS) having an isolated non-deleted (5q). Myelodysplastic syndrome (MDS) includes diseases such as refractory anemia, refractory anemia with excessive blast cells, refractory cytopenia with multiple lineage dysplasia, refractory cytopenia with single lineage dysplasia, and chronic myelomonocytic leukemia (CMML). In yet another embodiment, the hematological neoplasm is a lymphoid neoplasm.

[0131] A method according to a specific embodiment also includes a step of diagnosing a neoplasm. In some embodiments, the method includes a step of diagnosing that the subject has a solid tumor. In some embodiments, the method includes a step of diagnosing that the subject has a hematological neoplasm. In some embodiments, the method includes a step of diagnosing that the subject has a myeloproliferative neoplasm. In one example, the method includes a step of diagnosing that the subject has a myelofibrosis, such as primary myelofibrosis. In some embodiments, the subject has not previously received a telomerase inhibitor (e.g., the telomerase inhibitor Naivim). In some embodiments, the subject is a subject with low-risk transfusion-dependent MDS that is relapsed or refractory to erythropoietin-stimulating agents (ESAs). In some embodiments, the subject has not been given prior treatment with a hypomethylating agent (HMA). In some embodiments, the subject has not been given prior treatment with lenalidomide. In some embodiments, the subject is a non-deleted (5q) subject. In some embodiments, the subject is a subject who has relapsed or is refractory to a Janus kinase (JAK) inhibitor. In some embodiments, the subject method comprises the step of treating a subject having a myeloproliferative neoplasm as described in U.S. Patent No. 9,375,485 and International Patent Publication Nos. WO 2019 / 023667 and WO 2020 / 028261, the disclosures of which are incorporated herein by reference.

[0132] In some embodiments, lymphoid neoplasms (e.g., lymphomas) are B-cell neoplasms. Examples of B-cell neoplasms include precursor B-cell neoplasms (e.g., precursor B-lymphoblastic leukemia / lymphoma) and peripheral B-cell neoplasms (e.g., B-cell chronic lymphoblastic leukemia, pre-lymphoblastic leukemia, small lymphoblastic lymphoma (small lymphoblastic (SL) NHL), lymphoplasmocytic lymphoma / immune cell lymphoma, epithelial lymphoma, follicular lymphoma, follicular lymphoma (e.g., cytological grades: I (small cell), II (mixed small and large cells), III (large cell) and / or subtypes: diffuse and predominantly small cell types), non-Hodgkin lymphoma (NHL), low-grade / follicular non-Hodgkin lymphoma (NHL), intermediate-grade / follicular NHL), marginal zone B-cell lymphoma (e.g., extranodal (e.g., MALT type + / - mononuclear B-cell)) and / or nodal (e.g., + / - mononuclear B-cell)), and splenic marginal zone lymphoma (e.g., + / - chorionic lymphocytes), hairy cell leukemia, plasmacytoma / plasmacytic myeloma (e.g., myeloma and multiple myeloma), diffuse large B-cell lymphoma (e.g., primary mediastinal (thymic) B-cell lymphoma), intermediate-grade diffuse NHL, Burkitt lymphoma, high-grade B-cell lymphoma, Burkitt-like, high-grade immunoblastic NHL, high-grade lymphoblastic NHL, high-grade small non-truncation cell NHL, giant disease NHL, AIDS-associated lymphoma and Waldenström macroglobulinemia, including but not limited to these.

[0133] In some embodiments, lymphoid neoplasms (e.g., lymphomas) are T-cell and / or putative NK-cell neoplasms. Examples of T-cell and / or putative NK-cell neoplasms include progenitor T-cell neoplasms (progenitor T-lymphoblastic lymphoma / leukemia) and peripheral T-cell and NK-cell neoplasms (e.g., T-cell chronic lymphoblastic leukemia / pro-lymphoblastic leukemia and giant granulocyte leukemia (LGL)) (e.g., T-cell type and / or NK-cell type), cutaneous T-cell lymphoma (e.g., mycosis fungoides / Sézary syndrome), primary T-cell lymphoma unspecified (e.g., cytological categories (e.g., medium-sized cell, mixed medium and large cell), large cell, lymphoepithelial cell, subtype hepatosplenic γδ T-cell lymphoma and subcutaneous panniculitis T-cell lymphoma), angioimmunoblastic T-cell lymphoma (AILD), centrovascular lymphoma, intestinal T-cell lymphoma (e.g., + / - related enteropathy), adult T-cell lymphoma / leukemia (ATL), and anaplastic large cell Lymphoma (ALCL) (e.g., CD30+, T- and null-cell types), anaplastic large cell lymphoma and Hodgkin lymphoma) is included but not limited thereto.

[0134] In some embodiments, a lymphoid neoplasm (e.g., lymphoma) is Hodgkin's disease. For example, Hodgkin's disease may be lymphocyte-predominant, tuberous sclerosis, mixed cellular, lymphocyte-depleted, and / or lymphocyte-rich.

[0135] In some embodiments, cancer is leukemia. In some embodiments, leukemia is chronic leukemia. Examples of chronic leukemia include, but are not limited to, chronic myeloid I (granuloid) leukemia, chronic myeloid and chronic lymphocytic leukemia (CLL). In some embodiments, leukemia is acute leukemia. Examples of acute leukemia include, but are not limited to, acute lymphoblastic leukemia (ALL), acute myeloid leukemia, acute lymphoblastic leukemia, and acute myeloid leukemia (e.g., myeloid, promyeloid, myelomonocytic, mononucleic, and erythroleukemia).

[0136] In some embodiments, the cancer is a liquid tumor or a plasmacytoma. Plasmacytoma includes, but is not limited to, myeloma. Myeloma includes, but is not limited to, extramedullary plasmacytoma, solitary myeloma, and multiple myeloma. In some embodiments, the plasmacytoma is multiple myeloma.

[0137] In some embodiments, the cancer is multiple myeloma. Examples of multiple myeloma include, but are not limited to, IgG multiple myeloma, IgA multiple myeloma, IgD multiple myeloma, IgE multiple myeloma, and nonsecretory multiple myeloma. In some embodiments, the multiple myeloma is IgG multiple myeloma. In some embodiments, the multiple myeloma is IgA multiple myeloma. In some embodiments, the multiple myeloma is asymptomatic or painless multiple myeloma. In some embodiments, the multiple myeloma is progressive multiple myeloma. In some embodiments, the multiple myeloma may be resistant to drugs such as, but not limited to, bortezomib, dexamethasone (Dex-), doxorubicin (Dox-), and melphalan (LR).

[0138] In describing the method of the present invention, the term “subject” means a person or organism to which a telomerase inhibitor composition is administered subcutaneously. As such, the subject of the present invention may include, but is not limited to, mammals, e.g., humans, and other primates such as chimpanzees and other ape and monkey species, and in certain embodiments, the subject is a human. The subject may be a subject diagnosed with a myeloproliferative neoplasm, and the subject may have been a subject diagnosed by a healthcare professional with such a pathological condition.

[0139] The dosage of a telomerase inhibitor, such as imetelstat or imetelstat sodium, administered subcutaneously to a subject may vary in a range including about 2.0 mg / kg to 20.0 mg / kg, e.g., about 3.0 mg / kg to about 15.0 mg / kg, e.g., about 4.0 mg / kg to about 10 mg / kg, e.g., about 6 mg / kg to about 14 mg / kg, e.g., about 7 mg / kg to about 13 mg / kg, e.g., about 8 mg / kg to about 12 mg / kg, e.g., about 7.5 mg / kg to about 9.4 mg / kg, and about 9 mg / kg to about 11 mg / kg, and about 11 mg / kg to about 14 mg / kg. In some embodiments, the dosage of the telomerase inhibitor administered to a subject is about 7.5 mg / kg to about 9.4 mg / kg. In some embodiments, the dose of telomerase inhibitor administered to the subject is about 9 mg / kg to about 11 mg / kg. In some embodiments, the dose of telomerase inhibitor administered to the subject is about 11 mg / kg to about 14 mg / kg. For example, the dosage of telomerase inhibitors is 4.0 mg / kg, 4.1 mg / kg, 4.2 mg / kg, 4.3 mg / kg, 4.4 mg / kg, 4.5 mg / kg, 4.6 mg / kg, 4.7 mg / kg, 4.8 mg / kg, 4.9 mg / kg, 5.0 mg / kg, 5.1 mg / kg, 5.2 mg / kg, 5.3 mg / kg, 5.4 mg / kg, 5.5 mg / kg, 5.6 mg / kg, 5.7 mg / kg, 5.8 mg / kg, 5.9 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, 7.1 mg / kg, 7.2 mg / kg, 7.3 mg / kg, 7.4 mg / kg, 7.5 mg / kg, 7.6 mg / kg, 7.7 mg / kg, 7.8 mg / kg, 7.9 mg / kg, 8 mg / kg, 8.1 mg / kg, 8.2 mg / kg, 8.3 mg / kg, 8.4 mg / kg, 8.5 mg / kg, 8.6 mg / kg, 8.7 mg / kg, 8.8 mg / kg, 8.9 mg / kg, 9 mg / kg, 9.1 mg / kg, 9.2 mg / kg, 9.3 mg / kg, 9.4 mg / kg, 9.5 mg / kg, 9.6 mg / kg, 9.7 mg / kg, 9.8 mg / kg, 9.9 mg / kg, 10 mg / kg, 10.1 mg / kg, 10.2 mg / kg, 10.3 mg / kg, 10.4 mg / kg, 10.5 mg / kg, 10.6 mg / kg, 10.7 mg / kg, 10.8 mg / kg, 10.9 mg / kg, 11 mg / kg, 11.1 mg / kg, 11.2 mg / kg, 11.3 mg / kg, 11.4 mg / kg, 11.5 mg / kg, 11.6 mg / kg, 11.7 mg / kg, 11.8 mg / kg, 11.9 mg / kg, 12 mg / kg, 12.1 mg / kg, 12.2 mg / kg, 12.3 mg / kg, 12.4 mg / kg, 12.5 mg / kg, 12.6 mg / kg, 12.7 mg / kg, 12.8 mg / kg, 12.9 mg / kg, 13 mg / kg, 13.5 mg / kg, 14.0 mg / kg, 14.5 mg / kg, 15.0 mg / kg, 15.5 mg / kg, 16.0 mg / kg, 16.5 It may be mg / kg, 17.0 mg / kg, 17.0 mg / kg, 17.5 mg / kg, 18.0 mg / kg, 18.5 mg / kg, 19.0 mg / kg, 19.5 mg / kg, or 20.0 mg / kg. In a specific embodiment, the dose of telomerase inhibitor administered to the subject is about 9.4 mg / kg.

[0140] A dose of a telomerase inhibitor, such as imetelstat or imetelstat sodium, may be administered to a subject at a cycle of once every other day, once a week, once every 2 weeks (14 days), once every 3 weeks (21 days), or once every 4 weeks (28 days), once every 6 weeks, once every 8 weeks, once every 10 weeks, and once every 12 weeks. In a specific embodiment of the method, imetelstat is administered during 1, 2, 3, 4, 5, 6, 7, 8, or more than 8 dosing cycles, each cycle comprising: subcutaneous administration of about 2-11 mg / kg imetelstat once every 3 weeks, subcutaneous administration of about 2-11 mg / kg imetelstat once every 4 weeks, subcutaneous administration of about 2-11 mg / kg imetelstat once every 2 weeks, or subcutaneous administration of about 7.5-9.4 mg / kg imetelstat once every 3 weeks. In certain cases, each dosing cycle comprises subcutaneous administration of about 7.5-9.4 mg / kg imetelstat once every 4 weeks. In some cases, each dosing cycle comprises subcutaneous administration of about 9.4 mg / kg imetelstat once every 3 weeks. In some cases, each dosing cycle comprises subcutaneous administration of about 7.5 mg / kg imetelstat once about 4 weeks. In certain embodiments of the method, imetelstat is administered for 1, 2, 3, 4, 5, 6, 7, 8, or more than 8 dosing cycles, each cycle comprising: subcutaneous administration of about 5-14 mg / kg imetelstat once 3 weeks, subcutaneous administration of about 5-14 mg / kg imetelstat once 4 weeks, subcutaneous administration of about 5-14 mg / kg imetelstat once 2 weeks, or subcutaneous administration of about 7.5-14 mg / kg imetelstat once 3 weeks. In certain cases, each dosing cycle comprises subcutaneous administration of about 7.5-14 mg / kg imetelstat once 4 weeks. In some cases, each dosing cycle involves subcutaneous administration of approximately 14 mg / kg imetelstat once every approximately 3 weeks. In some cases, each dosing cycle involves subcutaneous administration of approximately 12 mg / kg imetelstat once every approximately 4 weeks.

[0141] In some embodiments, the amount of a telomerase inhibitor, such as imetelstat or imetelstat sodium, administered to an individual is about 0.5 to about 5 mg, about 5 to about 10 mg, about 10 to about 15 mg, about 15 to about 20 mg, about 20 to about 25 mg, about 20 to about 50 mg, about 25 to about 50 mg, about 50 to about 75 mg, about 50 to about 100 mg, about 75 to about 100 mg, about 100 to about 125 mg, about 125 to about 150 mg, about 150 to about 175 mg, about 175 to about 200 mg, about 200 to about 225 mg, about 225 to about 250 mg, about 250 to about 300 mg, about 300 to about 350 mg, about 350 to about 400 mg, about 400 to about 450 mg, or about 450 to about 500 mg, about 500 mg to about 600 mg, about 600 mg to about 700 mg, about 700 mg to about 800 mg, about 800 mg to about 900 mg, about 900 mg to about 1000 mg, about 1100 mg to about 1200 mg, about 1200 mg to about 1300 mg, about 1300 mg to about 1400 mg, about 1400 mg to about 1500 mg, about 1500 mg to about 1600 mg, about 1600 mg to about 1700 mg, about 1700 mg to about 1800 mg, about 1800 mg to about 1900 mg, about It is 1900 mg to about 2000 mg, about 2000 mg to about 2100 mg, about 2100 mg to about 2200 mg, about 2200 mg to about 2300 mg, about 2300 mg to about 2400 mg, and about 2400 mg to about 2500 mg.

[0142] In some embodiments, the amount of telomerase inhibitor in an effective dose (e.g., unit dose) administered to an individual is in the range of about 5 mg to about 1000 mg, 5 mg to about 500 mg, e.g., about 30 mg to about 300 mg, or about 50 mg to about 200 mg. In some embodiments, the amount of telomerase inhibitor is in a unit dose having an amount in the range of about 500 mg to about 3000 mg, 750 mg to about 2500 mg, e.g., about 1000 mg to about 2000 mg, or about 50 mg to about 200 mg. The unit dose may be liquid or lyophilized. In a specific embodiment of the method, imetelstat is administered during 1, 2, 3, 4, 5, 6, 7, 8 or more than 8 dosing cycles, each cycle comprising: subcutaneous administration of about 200-3000 mg imetelstat once every 3 weeks, subcutaneous administration of about 200-3000 mg imetelstat once every 4 weeks, subcutaneous administration of about 750-2500 mg imetelstat once every 3 weeks, or subcutaneous administration of about 750-2500 mg imetelstat once every 4 weeks.

[0143] In some embodiments, the concentration of the telomerase inhibitor administered to an individual is, for example, about 0.1 to about 300 mg / ml, about 0.1 to about 200 mg / ml, about 0.1 to about 180 mg / ml, about 0.1 to about 160 mg / ml, about 0.1 to about 140 mg / ml, about 0.1 to about 120 mg / ml, about 0.1 to about 100 mg / ml, about 0.1 to about 80 mg / ml, about 0.1 to about 60 mg / ml, about 0.1 to about 40 mg / ml, about 0.1 to about 20 mg / ml, about 0.1 to about 10 mg / ml, about 2 to about 40 mg / ml, about 4 to about 35 mg / ml, about 6 to about 30 mg / ml, about 8 to about 25 mg / ml, about 10 It is diluted (about 0.1 mg / ml) or concentrated (about 0.1 mg / ml), including either to about 20 mg / ml, about 12 to about 15 mg / ml, or any of about 0.1 mg / ml, 0.2 mg / ml, 0.3 mg / ml, 0.4 mg / ml, 0.5 mg / ml, 0.6 mg / ml, 0.7 mg / ml, 0.8 mg / ml, 0.9 mg / ml, 1 mg / ml, 1.1 mg / ml, 1.2 mg / ml, 1.3 mg / ml, 1.4 mg / ml, 1.5 mg / ml, 1.6 mg / ml, 1.7 mg / ml, 1.8 mg / ml, 1.9 mg / ml, 2 mg / ml, 2.1 mg / ml, 2.2 mg / ml, 2.3 mg / ml, 2.4 mg / ml, or 2.5 mg / ml. 300 mg / ml). In some embodiments, the concentration of the telomerase inhibitor is at least 0.1 mg / ml, 0.2 mg / ml, 0.3 mg / ml, 0.4 mg / ml, 0.5 mg / ml, 1.3 mg / ml, 1.5 mg / ml, 2 mg / ml, 3 mg / ml, 4 mg / ml, 5 mg / ml, 6 mg / ml, 7 mg / ml, 8 mg / ml, 9 mg / ml, 10 mg / ml, 11 mg / ml, 12 mg / ml, 13 mg / ml, 14 mg / ml, 15 mg / ml, 16 mg / ml, 17 mg / ml, 18 mg / ml, 19 mg / ml, 20 mg / ml, 21 mg / ml, 22 mg / ml, 23 mg / ml, 24 mg / ml, 25 mg / ml, 26 mg / ml, 27 mg / ml, 28 mg / ml, 29 mg / ml, 30 mg / ml, 31 mg / ml, 32 mg / ml, 33 mg / ml, 33.3 mg / ml, 34 mg / ml, 35 mg / ml, 36 mg / ml, 37 mg / ml, 38 mg / ml, 39 mg / ml, 40 mg / ml, 50 mg / ml, 60 mg / ml, 70 mg / ml, 80 mg / ml, 90 mg / ml, 100 mg / ml, 110 mg / ml, 120 mg / ml, 130 mg / ml, 140 mg / ml, 150 mg / ml, 160 mg / ml, 170 mg / ml, 180 mg / ml, 190 mg / ml, 200 mg / ml, 210 mg / ml, 220 mg / ml, 230 mg / ml, 240 mg / ml, 250 mg / ml, 260 mg / ml, 270 mg / ml, 280 mg / ml, 290 mg / ml, or 300 mg / ml.

[0144] In an embodiment, each dose of the telomerase inhibitor composition is administered subcutaneously to a subject at least once every 7 days, e.g., at least once every 10 days, e.g., at least once every 14 days, e.g., at least once every 21 days, e.g., at least once every 28 days, and at least once every 35 days. In some embodiments, the telomerase inhibitor composition is administered subcutaneously to a subject every other day. In some embodiments, the telomerase inhibitor composition is administered subcutaneously to a subject once a week. In some embodiments, the telomerase inhibitor composition is administered subcutaneously to a subject once every two weeks. In another embodiment, the telomerase inhibitor composition is administered subcutaneously to a subject once every three weeks. In yet another embodiment, the telomerase inhibitor composition is administered subcutaneously once every four weeks.

[0145] Kit

[0146] Additionally, a kit is provided, and the kit comprises at least one, for example, a plurality of target subcutaneous telomerase inhibitor compositions as described above. In certain embodiments, the target subcutaneous telomerase inhibitor compositions of the kit may be provided in a package. For example, each composition of the kit may be presented in an individual pouch, bottle, or similar container to preserve the composition until use. The kit may further comprise other components for performing the target method, such as a delivery device or fluid for rinsing the skin before administering one or more of the target compositions. In certain embodiments, the kit comprises a subcutaneous injection device configured to deliver a therapeutically effective amount of the composition to a target. In some cases, the injection device comprises a syringe and a needle. In other embodiments, the injection device is a bolus injection device configured to deliver a predetermined amount of the composition subcutaneously. In certain embodiments, the telomerase inhibitor composition is pre-loaded into the subcutaneous injection device. The kit may also include a gauze pad or other device for cleaning the injection site that can be used to perform the target method. In some embodiments, the telomerase inhibitor composition is formulated as a solid or lyophilized material, and the kit may further comprise one or more buffer compositions or solvents for reconstituting the target composition for subcutaneous injection.

[0147] Additionally, the kit may also include instructions on how to use the target telomerase inhibitor composition, and the instructions may include information on how to administer the composition, a dosing schedule, and a record storage device for carrying out the treatment regimen. The instructions are recorded on a suitable recording medium. For example, the instructions may be printed on a substrate such as paper or plastic. As such, the instructions may be presented in the kit as a package insert, on the container of the kit or on the label of its components, etc. (i.e., in connection with the packaging or subpackaging). In another embodiment, the instructions are presented as an electronic storage data file present on a suitable computer-readable storage medium, such as a CD-ROM, diskette, etc. In yet another embodiment, the actual instructions are not present in the kit, but means for obtaining the instructions from a remote source, for example, via the Internet, are provided. An example of this embodiment is a kit that includes a web address where the instructions can be viewed and / or downloaded. As with the instructions, a protocol for obtaining the instructions may be recorded on a suitable substrate.

[0148] Examples

[0149] The following examples are presented to provide those skilled in the art with a complete disclosure and description of the methods for manufacturing and using the present invention, and are not intended to limit the scope of what the inventors consider to be their invention, nor are they intended to indicate that the experiments below are all or only experiments performed. While efforts have been made to ensure accuracy with respect to the figures used (e.g., amounts, temperatures, etc.), some experimental errors and deviations must be taken into account. Unless otherwise indicated, parts are parts by weight, molecular weights are weight-average molecular weights, temperatures are in degrees Celsius, and pressures are atmospheric or near atmospheric pressure. "Average" means the arithmetic mean. Standard abbreviations, e.g., bp, base pair(s); kb, kilobase(s); pl, picoliters(s); s or sec, seconds(s); min, minutes(s); h or hr, hours(s); aa, amino acid(s); kb, kilobase(s); bp, base pair(s); nt, nucleotide(s); im, intramuscular(s); ip, intraperitoneal(s); sc, subcutaneous, etc. can be used.

[0150] Example 1 - Study on the Bioavailability of Imetelstat Sodium in Rats via Subcutaneous and Intravenous Routes

[0151] Materials and Methods

[0152] Sodium imetelstat was administered intravenously to four male Sprague-Dawley rats (approx. 250 g–260 g) (IV volume 37.5 μL; 30 mg / kg) and subcutaneously to four male Sprague-Dawley rats (approx. 250 g–260 g) (SC volume 113 μL; 90 mg / kg). Sodium imetelstat is a 5' palmitoylated 13-mer thiophosphoramidate oligonucleotide consisting of the sequence 5'-TAGGGTTAGACAA-3'. Sodium imetelstat was dissolved in water immediately before administration.

[0153] Blood was collected from each rat at 0.083 hours (approx. 5 min), 0.25 hours (15 min), 1 hour, 2 hours, 4 hours, 8 hours, and 24 hours. 1.5 mL to 2.0 mL of whole blood was collected to obtain approximately 700 μL to 1000 μL of plasma. Blood was collected from IV-administered rats (different from those used for IV administration) via a secondary catheter.

[0154] Blood samples were collected in EDTA tubes and centrifuged immediately after collection. The collected plasma samples were transferred to separate vials. The plasma concentration of sodium imetelstat was analyzed by LC / MS / MS.

[0155] result

[0156] Plasma concentrations of imetelstat sodium analyzed over a 24-hour period are shown in Figure 1. Plasma concentrations of imetelstat sodium were analyzed by non-compartmental PK analysis. Subcutaneous (sc) bioavailability was calculated based on the following formula: F = (AUC(sc) / dose(sc)) / (AUC(iv) / dose(iv)) x 100%. The calculated subcutaneous bioavailability of imetelstat is approximately 95.8%. In a repeated experiment, the calculated subcutaneous bioavailability of imetelstat in rats was approximately 81.5%.

[0157] conclusion

[0158] The results shown in Figure 1 demonstrate that imetelstat sodium was absorbed into systemic circulation after subcutaneous administration and showed a bioavailability of approximately 80-95% compared to IV administration, thereby supporting the use of the imetelstat composition for subcutaneous administration.

[0159] Example 2 - Compatibility and Stability of Imetelstat Sodium Incubated with Hyaluronidase

[0160]

[0161] Preliminary study:

[0162] Confirmation of the compatibility of Imetelstat co-formulated with rHuPH20

[0163] Data generated from an initial feasibility study conducted as a preliminary study for the following key commerciality and stability studies provided support for the commerciality of imetelstat co-formulated with rHuPH20 by demonstrating that the results for the imetelstat + rHuPH20 co-formulations, denoted as formulations D, F, and G (as defined below) manufactured for this preliminary study, were similar to the results for the imetelstat active pharmaceutical ingredient lot without rHuPH20 used to manufacture the co-formulations (see table below).

[0164] The following formulations were prepared.

[0165]

[0166]

[0167]

[0168] Major Research:

[0169] purpose

[0170] The purpose of this study is to demonstrate the compatibility and stability of rHuPH20 co-formulated with imetelstat sodium under various storage conditions.

[0171] substance

[0172] Imetelstat sodium

[0173] rHuPH20-fortified Finished Pharmaceutical Product (EDP): 1 mg / mL, 110,000 U / mL, store at 5℃

[0174] 0.9% sodium chloride

[0175] L-histidine (≥99% (TLC))

[0176] L-methionine (reagent grade, ≥98% (HPLC))

[0177] Polysorbate 80

[0178] NaCl (ACS grade)

[0179] Concentrated hydrochloric acid (ACS reagent, 37%)

[0180] water

[0181] Nylon 47 mm filter membrane 0.2 μm

[0182] Sample formulation

[0183]

[0184] Sample manufacturing

[0185]

[0186] Manufacture of rHuPH20-enhanced finished pharmaceutical product (EDP) buffers

[0187] Add approximately 450 mL of water to a 500 mL volumetric flask. Add approximately 776 mg (±15 mg) of L-histidine, approximately 3.80 g of NaCl (±75 mg), and approximately 746 mg (±15 mg) of L-methionine to the volumetric flask. Dissolve the ingredients completely using a stirring rod. Confirm that the ingredients are dissolved using visual inspection.

[0188] Add 1 mL of the 10.0% polysorbate 80 composition to the solution in a volumetric flask. Mix the composition with a stirring rod. Adjust the pH to a final pH of 6.5 ± 0.3 using concentrated HCl. Prepare a final volume of 500 mL by appropriate amount. Filter the composition through a 47 mm nylon filter membrane with a 0.2 µm filtration system, store at 5°C, and remain stable for at least 2 weeks.

[0189] The composition is stored in a 20 mL borosilicate glass scintillation vial at 5°C in a vertical orientation, 25°C in a vertical orientation, and 37°C in a vertical orientation.

[0190] Stability protocol schedule

[0191] The table below summarizes the number of samples set per interval / condition / combination (initial interval samples are stored at -20℃).

[0192]

[0193] Stability Pull and Test Protocol

[0194] Tests are performed on samples set according to specified time intervals and conditions to evaluate the compatibility and stability of the imetelstat + rHuPH20 co-formulation (see table below). Test methods include the evaluation of appearance, pH, oligo concentration (UV), purity by HPLC, assay by LC / MS, rHuPH20 enzyme activity, and telomerase activity for imetelstat by TRAP assay (Mender and Shay, Bio Protoc. 2015: 5(22)).

[0195]

[0196] To test how rHuPH20 affects the imetelstat telomerase inhibitory activity when co-mixed with imetelstat, three formulations A, D, and F as indicated in the sample formulation table above—namely, imetelstat sodium alone (Formulation A) or imetelstat sodium co-mixed with rHuPH20 (Formulations D and F)—were tested for imetelstat activity using the TRAP (Telomerase Repeat Amplification Protocol) assay. Briefly, cancer cell line Hela cells were treated in vitro with three of each of seven (7) different concentrations (0.1, 1, 5, 7.5, 10, 25, and 50 μM) of each test formulation and one (1) drug-free control for 24 hours. For protein extraction and quantification, cell pellets were lysed, and equal amounts of protein from cells treated with each concentration of formulation were tested using the TRAP assay. Relative telomerase activity (RTA) for each quantitative polymerase chain reaction (qPCR) result was determined from standard curves generated by the TRAP assay of serial dilutions of protein lysates in untreated Hela cells. The mean RTA from the replicates was calculated for each given formulation at each concentration and compared to the mean RTA value of the drug-free control to generate the percentage (%) of telomerase activity inhibition. The drug concentration (IC10) required to achieve 50% inhibition of telomerase activity for each formulation was plotted against the treatment concentration. 50 The value) was determined. The results in Figure 2 are for the IC of all three formulations. 50 It was indicated that the values ​​were very similar and close to the logarithm of 1 (10 μM), indicating that the co-mixing of rHuPH20 and imetelstat sodium did not affect the telomerase inhibitory activity of imetelstat.

[0197] Example 3 - Assessment of rHuPH20 activity in formulations of Imetelstat and rHuPH20

[0198] outline

[0199] In an additional feasibility test performed as a preliminary study for the major compatibility and stability studies described in Example 2 above, a formulation containing imetelstat co-mixed with rHuPH20 was tested for rHuPH20 activity. It provides assay results based on the reaction of rHuPH20 with biotinylated hyaluronic acid (HA) and demonstrates that rHuPH20 activity can be measured in the presence of imetelstat in the co-mixed sample.

[0200] result

[0201] The activity of rHuPH20 was determined in the presence and absence of imetelstat using biotinylated hyaluronic acid. To measure the level of hyaluronic acid degradation by rHuPH20, a composition containing 2000 U / mL rHuPH20 in saline was assayed with biotinylated hyaluronic acid and compared with a composition containing 100 mg / ml imetelstat along with 2000 U / mL rHuPH20. The rHuPH20 activity detected in the samples for each imetelstat + rHuPH20 composition was close to the amount based on the activity observed in the rHuPH20 alone sample (Table below).

[0202]

[0203] Example 4 - Study of single-dose subcutaneous bioavailability and topical tolerance of imetelstat with or without rHuPH20 in rats

[0204] purpose

[0205] The purpose of this study is to evaluate the systemic absorption (in terms of plasma levels) and local (injection site) tolerance of imetelstat following a single subcutaneous (SC) injection in rats, either alone or in combination with rHuPH20 (a recombinant human hyaluronidase enzyme product to be co-formulated with imetelstat). The subcutaneous bioavailability of imetelstat is determined by a group receiving an IV dose of this study product. This study is designed to demonstrate that imetelstat co-formulated with rHuPH20 is tolerable and provides a pharmacokinetic profile for imetelstat suggesting its clinical use in delivering the necessary liquid volume containing a sufficient dose of imetelstat via the subcutaneous administration route.

[0206] animal

[0207] Sprague-Dowley rats, each of standard age at the beginning of the study.

[0208] Test items

[0209] Test imetelstat containing and not containing rHuPH20.

[0210] Study Design / Dosage

[0211] Imetelstat is administered as a single dose on Day 1 by slow-push intravenous injection (Group 1) or SC injection (Group 2). Imetelstat co-formulated with rHuPH20 is administered as a single dose on Day 1 by SC injection (Groups 3a and 3b). The vehicle for imetelstat is simultaneously administered to animals in Group 2 at different SC sites (control injection sites). Likewise, the vehicle for imetelstat co-formulated with rHuPH20 is administered to animals in Groups 3a and 3b at separate control injection sites.

[0212] The allocation by group is shown in the table below.

[0213]

[0214] Observation and sample collection

[0215] Clinical observation Study observations include standard clinical observations, evaluation of the local injection site, food consumption, and body weight.

[0216] PK sample :

[0217] Blood samples are collected from all animals in K2EDTA-containing tubes at various times from 5 minutes to 8 hours after administration. The samples are processed into plasma under refrigerated conditions, and the resulting plasma is stored by rapid freezing.

[0218] PK Sample Bioanalysis and Data Interpretation :

[0219] Rat plasma samples are analyzed for imetelstat concentration using a validated hybridization ELISA method. AUC 0-t , AUC 0-inf , C max , T max , K el , CL, V d and t ½ Calculate PK parameters such as appropriate for imetelstat in plasma.

[0220] Example 5 - Single-dose subcutaneous pharmacokinetics and topical tolerance study in monkeys

[0221] purpose

[0222] The purpose of this study is to evaluate the systemic absorption (in terms of plasma levels) and local (injection site) tolerance of imetelstat following a single subcutaneous injection in crab-eating monkeys, either alone or in combination with rHuPH20 (a recombinant human hyaluronidase enzyme product to be co-formulated with imetelstat). This study is designed to demonstrate that imetelstat co-formulated with rHuPH20 is tolerable and provides a pharmacokinetic profile for imetelstat suggesting its clinical use in delivering the necessary liquid volume containing a sufficient dose of imetelstat via the subcutaneous administration route.

[0223] animal

[0224] Crab-eating monkeys of standard age and weight at the beginning of the study.

[0225] Test items

[0226] Test imetelstat containing and not containing rHuPH20.

[0227] Study Design / Dosage

[0228] Imetelstat is administered as a single dose by subcutaneous injection on Day 1 (Group 1), and imetelstat co-formulated with rHuPH20 is administered as a single dose by subcutaneous injection on Day 1 (Group 2). The vehicle for imetelstat is simultaneously administered to the animals in Group 1 at different subcutaneous sites (control injection sites). Likewise, the vehicle for the co-formulation of imetelstat and rHuPH20 is administered to the animals in Group 2 at separate control sites.

[0229] Group assignments are shown in the table below.

[0230]

[0231] Observation and sample collection

[0232] Clinical observation :

[0233] Study observations including standard clinical observations, evaluation of the local injection site (including histopathological examination), food intake, and body weight.

[0234] PK sample :

[0235] Blood samples are collected from all animals in K2EDTA-containing tubes at various times from 5 minutes to 24 hours after administration. The samples are processed into plasma under refrigerated conditions, and the resulting plasma is stored by rapid freezing.

[0236] PK Sample Bioanalysis and Data Interpretation :

[0237] Monkey plasma samples are analyzed for imetelstat concentration using a validated hybridization ELISA method. AUC 0-t , AUC 0-inf , C max , T max , K el , CL, V d and t ½ Calculate PK parameters such as appropriate for imetelstat in plasma.

[0238] Example 6 - Safety and pharmacokinetics study in healthy volunteers or patients

[0239] purpose

[0240] The purpose of this study is to evaluate the safety, tolerance, and pharmacokinetics of imetelstat after subcutaneous (SC) and intravenous (IV) administration. The study is designed to demonstrate that the co-formulation of rHuPH20 and imetelstat provides the necessary liquid volume of subcutaneous delivery containing a sufficient dose of imetelstat to demonstrate an acceptable safety, tolerance, and pharmacokinetic profile for imetelstat via the subcutaneous administration route.

[0241] Summary of Research Design

[0242] The study is conducted in two parts. Part 1 is the dose escalation phase involving the subcutaneous administration of imetelstat. Up to four cohorts are planned for Part 1. Subjects participate in only one cohort. In each cohort, subjects are administered a single subcutaneous dose of imetelstat on Day 1. Part 2 is an open-label, randomized, two-treatment, crossover, or parallel study design. The dose for Part 2 is selected after reviewing safety and pharmacokinetic data from Part 1.

[0243] If Part 2 is a crossover design, pharmacokinetic samples are collected for 48 hours after administering alternating subcutaneous or intravenous doses of imetelstat according to the randomization schedule on Day 1 of Periods 1 and 2. In Part 2 of the crossover design, a drug-free period is scheduled between imetelstat doses. If Part 2 is a parallel design, pharmacokinetic samples are collected for 48 hours after randomizing patients to receive subcutaneous or intravenous doses of imetelstat on Day 1.

[0244] Dosage, dosage form, route, and dosage therapy

[0245] Subcutaneous imetelstat is formulated as a co-mixture with hyaluronidase (rHuPH20) in a suitable vehicle. IV imetelstat does not contain hyaluronidase and is formulated in 0.9% sodium chloride.

[0246] Part 1:

[0247] In Part 1, a single subcutaneous dose is administered. Cohort doses are selected based on a review of safety and pharmacokinetic data from preclinical pharmacokinetic and topical tolerance studies in rats and monkeys.

[0248] Part 2:

[0249] Administer imetelstat to the subject subcutaneously or intravenously in one of the following sequences:

[0250] Sequence A (crossover design): a single subcutaneous dose of imetelstat on Day 1 followed by a single IV dose of imetelstat after an appropriate rest period; or

[0251] Sequence B (crossover design): A single IV dose of imetelstat on Day 1 followed by a single SC dose of imetelstat after an appropriate rest period; or

[0252] Sequence C (Parallel Design): On day 1, a single subcutaneous dose of imetelstat is administered to one cohort of subjects, and on day 1, a single IV dose of imetelstat is administered to another cohort of subjects.

[0253] The subcutaneous dose is administered as a single injection to one site, and the IV dose is administered as a 2-hour infusion.

[0254] evaluation

[0255] Pharmacokinetics:

[0256] Blood samples for plasma concentrations of imetelstat are collected at various times, before administration and between 0.5 and 48 hours after the start of the subcutaneous dose or infusion.

[0257] Calculate the following PK parameters appropriately for imetelstat in plasma: AUC 0-t , AUC 0-inf , AUC% extrap , C max , T max , K el , C L , C L / F , and t ½ .

[0258] Safety:

[0259] Safety and tolerance assessments may be monitored through standard procedures, which may include electrocardiogram (ECG), physical examination, vital sign measurements, clinical laboratory tests, and AEs. Summary statistics may be calculated as deemed clinically appropriate.

[0260] Although the foregoing invention has been described in some detail by way of examples and illustrations for clear understanding, it is readily apparent to those skilled in the art that certain changes and modifications may be made without departing from the spirit or scope of the appended claims in light of the teachings of the present invention.

[0261] Accordingly, the foregoing merely illustrates the principles of the present invention. It will be understood by those skilled in the art that various configurations embodying the principles of the present invention and falling within the spirit and scope thereof, even if not explicitly stated or indicated herein, may be devised. Furthermore, all examples and conditional language cited herein are intended primarily to help the reader understand the concepts of the inventors who contributed to the principles and technical development of the present invention, and should not be considered limited to such specifically cited examples and conditions. Moreover, all statements in this invention citing specific examples, as well as the principles, aspects, and embodiments of the present invention, are intended to encompass all structural and functional equivalents thereof. Furthermore, such equivalents are intended to include all currently known equivalents and equivalents to be developed in the future, that is, all elements developed to perform the same function regardless of structure. Moreover, nothing disclosed herein is intended to be dedicated to the public, regardless of whether such disclosures are explicitly cited in the claims.

[0262] Accordingly, the scope of the invention is not intended to be limited to the exemplary embodiments shown and described herein. Rather, the scope and spirit of the invention are embodied by the appended claims. In the claims, 35 USC §112(f) or 35 USC §112(6) is explicitly defined to apply to the feature of the claim only if the precise phrase “means for” or the precise phrase “step for” is cited at the beginning of such feature in the claim; if such precise phrase is not used in the feature of the claim, 35 USC §112(f) or 35 USC §112(6) does not apply.

[0263] Other embodiments of the present invention may include the following:

[0264] 1. A composition formulated for subcutaneous administration comprising the following:

[0265] A telomerase inhibitor comprising an oligonucleotide and a lipid moiety linked to the 5' and / or 3' ends of the oligonucleotide; and

[0266] Hyaluronidase enzyme.

[0267] 2. In paragraph 1, the hyaluronidase enzyme is recombinant human hyaluronidase, composition.

[0268] 3. A composition according to paragraph 2, wherein the hyaluronidase enzyme is rHuPH20.

[0269] 4. A composition comprising a variant or fragment of the pH 20 hyaluronidase enzyme in paragraph 1.

[0270] 5. A composition according to paragraph 4, wherein a variant or fragment of PH20 comprises one or more amino acid residue substitutions selected from the group consisting of T341A, T341C, T341G, S343E, M345T, K349E, L353A, L354I, N356E, and I361T in wild-type PH20 having the amino acid sequence of SEQ ID NO. 1.

[0271] 6. A composition according to paragraph 5, wherein a variant or fragment of PH20 comprises one or more amino acid residue substitutions selected from the group consisting of T341A, T341C, L354I, and N356E.

[0272] 7. A composition according to paragraph 4, wherein a variant or fragment of PH20 comprises one or more amino acid residue substitutions in a region corresponding to the alpha-helical region or linker region of wild-type PH20 having the amino acid sequence of SEQ ID NO. 1.

[0273] 8. A composition according to paragraph 7, wherein the alpha-helical region is an alpha-helical 8 region comprising amino acid residues S347 to C381 and the linker region is a linker region comprising amino acid residues A333 to R346 between alpha-helical 7 and alpha-helical 8.

[0274] 9. A composition according to paragraph 7, wherein the alpha-helical region and the linker region comprise amino acid residues T341 to N363, T341 to I361, L342 to I361, S343 to I361, I344 to I361, M345 to I361, or M345 to N363.

[0275] 10. A composition according to paragraph 7, wherein the alpha-helical 8 region and the linker region between alpha-helical 7 and alpha-helical 8 are substituted with one or more amino acid residues of the corresponding region of Hyal1.

[0276] 11. A composition according to paragraph 4, wherein a variant or fragment of PH20 comprises one or more amino acid residue substitutions at one or more positions selected from the group consisting of T341, L342, S343, I344, M345, S347, M348, K349, L352, L353, D355, E359, I361 and N363.

[0277] 12. A composition according to paragraph 11, wherein a variant or fragment of PH20 comprises the following amino acid residue substitution:

[0278] One or more of L354I and N356E; and

[0279] One or more amino acid residue substitutions selected from the group consisting of T341A, T341C, T341D, T341G, T341S, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, D355K, E359D, I361T, and N363G.

[0280] 13. A composition according to paragraph 12, wherein a variant or fragment of PH20 comprises the following:

[0281] (i) Substitutions M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D and I361T;

[0282] (ii) Substitutions T341A, T341C, T341D, T341G, T341S, L342W, S343E, I344N and N363G; or

[0283] (iii) Any one amino acid residue substitution selected from the following amino acid residue substitution groups:

[0284] (a) T341S, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D, and I361T;

[0285] (b) L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D, and I361T;

[0286] (c) M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D, I361T, and N363G;

[0287] (d) T341G, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D, and I361T;

[0288] (e) T341A, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D, and I361T;

[0289] (f) T341C, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D, and I361T;

[0290] (g) T341D, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D, and I361T;

[0291] (h) I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D and I361T; and

[0292] (i) S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D and I361T.

[0293] 14. A composition in which one or more of the N-terminal or C-terminal amino acid residues of a variant or fragment of PH20 are deleted, in any one of paragraphs 4 to 13.

[0294] 15. In paragraph 14, the composition, wherein the cutting is as follows:

[0295] (i) located before an amino acid residue selected from the group consisting of M1 to P42 at the N-terminus such that one or more residues at the N-terminus are deleted;

[0296] (ii) located before amino acid residues L36, N37, F38, R39, A40, P41, or P42 at the N-terminus so as to delete one or more residues at the N-terminus;

[0297] (iii) located after an amino acid residue selected from the group consisting of V455 to L509 at the C-terminus such that one or more amino acid residues are deleted at the C-terminus;

[0298] (iv) located after an amino acid residue selected from V455, C458, D461, C464, I465, D466, A467, F468, K470, P471, P472, M473, E474, T475, E476, E477, P478, Q479, I480, F481, Y482, N483, A484, P486, T488, or S490 at the C-terminus, such that one or more amino acid residues are deleted at the C-terminus.

[0299] 16. A composition comprising, in any one of paragraphs 4 to 15, a human growth hormone-derived signal peptide having the amino acid sequence MATGSRTSLLLAFGLLCLPWLQEGSA of SEQ No. 3 at the N-terminus, a human serum albumin-derived signal peptide having the amino acid sequence MKWVTFISLLFLFSSAYS of SEQ No. 4, or a human Hyal1-derived signal peptide having the amino acid sequence MAAHLLPICALFLTLLDMAQG of SEQ No. 5.

[0300] 22. A composition in any one of paragraphs 4 to 21, wherein the variant or fragment of PH20 is as follows:

[0301] (i) a peptide having at least 90% sequence identity with the amino acid sequence presented as SEQ No. 1 or amino acid residues 36-482, 36-477, 366-478, 36-479, 36-480, 36-481, and 36-483 of SEQ No. 1;

[0302] (ii) a peptide having at least 95% sequence identity with the amino acid sequence presented as SEQ No. 1 or amino acid residues 36-482, 36-477, 366-478, 36-479, 36-480, 36-481, and 36-483 of SEQ No. 1;

[0303] (iii) composed of an amino acid sequence selected from sequence numbers 60 to 115;

[0304] (iv) Has the amino acid sequence of sequence number 99.

[0305] 23. A composition in which, in any one of paragraphs 1-22, hyaluronidase is present in the composition in an amount of 100 U to 50,000 U.

[0306] 24. A composition in which hyaluronidase is present in an amount of 100 U / mL to 50,000 U / mL in any one of paragraphs 1 to 23.

[0307] 25. A composition comprising one or more pharmaceutically acceptable excipients in any one of paragraphs 1 to 24.

[0308] 26. A composition comprising one or more sugars in addition to any one of paragraphs 1 to 25.

[0309] 27. A composition according to paragraph 26, wherein one or more sugars comprise a monosaccharide.

[0310] 28. A composition in any one of paragraphs 26 to 27, wherein one or more sugars comprise a polysaccharide.

[0311] 29. A composition according to paragraph 28, wherein one or more polysaccharides are selected from the group consisting of trehalose and sucrose.

[0312] 30. A composition in which, in any one of paragraphs 26 to 29, one or more sugars are present in the composition in an amount of 10 mM to 500 mM.

[0313] 31. A composition comprising one or more amino acids in addition to any one of paragraphs 1 to 30.

[0314] 32. A composition according to paragraph 31, wherein the amino acid is selected from methionine and histidine.

[0315] 33. A composition in which, in any one of paragraphs 31 to 32, one or more amino acids are present in the composition in an amount of 1 mM to 100 mM, optionally in an amount of 1 mM to 50 mM.

[0316] 34. A composition comprising a buffer additionally comprising, in any one of paragraphs 1 to 33.

[0317] 35. The composition of paragraph 34, wherein the buffer is present in the composition in an amount sufficient to maintain the composition at a pH of 3.0 to 9.0, optionally in an amount sufficient to maintain the composition at a pH of 5.5 to 7.5.

[0318] 36. A composition in which, in any one of paragraphs 34 to 35, a buffer is present in the composition in an amount of 1 to 100 mM, optionally in an amount of 1 mM to 50 mM.

[0319] 37. A composition in any one of paragraphs 1 to 36, wherein the oligonucleotide of the telomerase inhibitor comprises at least one N3' → P5' thiophosphoramidate nucleoside linkage.

[0320] 38. A composition in any one of paragraphs 1 to 37, wherein the lipid moiety of the telomerase inhibitor is connected to the 5' and / or 3' end of an oligonucleotide through a linker, optionally the linker is a glycerol or aminoglycerol linker.

[0321] 39. A composition in any one of paragraphs 1 to 38, wherein the lipid moiety of the telomerase inhibitor is a palmitoyl (C16) moiety.

[0322] 40. A composition in any one of paragraphs 1 to 39, wherein the telomerase inhibitor is imetelstat or a pharmaceutically acceptable salt thereof, and optionally the telomerase inhibitor is imetelstat sodium.

[0323] 41. In any one of paragraphs 1 to 40, a composition in which a telomerase inhibitor is present in the composition at the following dosage:

[0324] (i) about 2.0 mg / kg to 20.0 mg / kg;

[0325] (ii) about 3 mg / kg to about 15 mg / kg;

[0326] (iii) about 9 mg / kg to about 11 mg / kg; or

[0327] (iv) About 11 mg / kg to about 14 mg / kg.

[0328] 42. A composition that is freeze-dried in any one of paragraphs 1 to 41.

[0329] 43. A method for treating a subject having a neoplasm, comprising the step of subcutaneously administering a composition comprising the following to the subject:

[0330] A telomerase inhibitor comprising an oligonucleotide and a lipid moiety linked to the 5' and / or 3' ends of the oligonucleotide; and

[0331] Hyaluronidase enzyme.

[0332] 44. Method in paragraph 43, wherein the neoplasm is a hematological neoplasm selected from myelofibrosis (MF), myelodysplastic syndrome (MDS), essential thrombocytosis (ET), polycythemia vera (PV), chronic myeloid leukemia (CML), chronic neutrophilic leukemia, chronic eosinophilic leukemia, and acute myeloid leukemia (AML).

[0333] 45. A method comprising, in any one of paragraphs 43 to 44, additionally diagnosing the subject as having a hematological neoplasm.

[0334] 46. ​​A method in which, in any one of paragraphs 43 to 45, the composition is administered subcutaneously to a subject as follows:

[0335] (i) Once every other day;

[0336] (ii) Once every 7 days;

[0337] (iii) once every 21 days; or

[0338] (iv) Once every 28 days.

[0339] 47. A method that is repeated at least once, optionally at least five times, in any one of paragraphs 43 to 46.

[0340] 48. A method in any one of paragraphs 43 to 47, wherein a telomerase inhibitor is administered to a subject at the following dose:

[0341] (i) about 2.0 mg / kg to 20.0 mg / kg;

[0342] (ii) about 3 mg / kg to about 15 mg / kg;

[0343] (iii) about 9 mg / kg to about 11 mg / kg; or

[0344] (iv) About 11 mg / kg to about 14 mg / kg.

[0345] 49. A method in any one of paragraphs 43 to 48, wherein the hyaluronidase enzyme is recombinant human hyaluronidase, optionally rHuPH20.

[0346] 50. A method in any one of paragraphs 43 to 48, wherein the composition comprises a variant or fragment of a pH 20 hyaluronidase enzyme.

[0347] 51. The method of paragraph 50, wherein a variant or fragment of PH20 comprises one or more amino acid residue substitutions selected from the group consisting of T341A, T341C, T341G, S343E, M345T, K349E, L353A, L354I, N356E, and I361T of wild-type PH20 having the amino acid sequence of SEQ ID NO. 1, and optionally a variant or fragment of PH20 comprises one or more amino acid residue substitutions selected from the group consisting of T341A, T341C, L354I, and N356E.

[0348] 52. A method according to paragraph 50, wherein a variant or fragment of PH20 comprises one or more amino acid residue substitutions in a region corresponding to the alpha-helical region or linker region of wild-type PH20 having the amino acid sequence of SEQ ID NO. 1.

[0349] 53. In paragraph 52,

[0350] (i) the alpha-helical region is an alpha-helical 8 region containing amino acid residues S347 to C381 and the linker region is a linker region containing amino acid residues A333 to R346 between alpha-helical 7 and alpha-helical 8;

[0351] (ii) the alpha-helical region and the linker region comprise amino acid residues T341 to N363, T341 to I361, L342 to I361, S343 to I361, I344 to I361, M345 to I361, or M345 to N363;

[0352] (iii) A method in which the alpha-helix 8 region and the linker region between alpha-helix 7 and alpha-helix 8 are substituted with one or more amino acid residues of the corresponding region of Hyal1.

[0353] 54. A method according to paragraph 50, wherein a variant or fragment of PH20 comprises one or more amino acid residue substitutions at one or more positions selected from the group consisting of T341, L342, S343, I344, M345, S347, M348, K349, L352, L353, D355, E359, I361 and N363.

[0354] 55. In paragraph 54, a variant or fragment of PH20 is substituted with the following amino acid residue:

[0355] One or more of L354I and N356E; and

[0356] One or more amino acid residue substitutions selected from the group consisting of T341A, T341C, T341D, T341G, T341S, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, D355K, E359D, I361T and N363G,

[0357] A method comprising, optionally comprising variants or fragments of PH20 substituting M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D, and I361T.

[0358] 56. In paragraph 55, a method wherein a variant or fragment of PH20 comprises the following:

[0359] (i) Substitutions T341A, T341C, T341D, T341G, T341S, L342W, S343E, I344N and N363G; or

[0360] (ii) Any one amino acid residue substitution selected from the following amino acid residue substitution groups:

[0361] (a) T341S, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D and I361T;

[0362] (b) L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D and I361T;

[0363] (c) M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D, I361T and N363G;

[0364] (d) T341G, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D and I361T;

[0365] (e) T341A, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D and I361T;

[0366] (f) T341C, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D and I361T;

[0367] (g) T341D, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D and I361T;

[0368] (h) I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D and I361T; and

[0369] (i) S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D and I361T.

[0370] 57. A method in which one or more of the N-terminal or C-terminal amino acid residues of a variant or fragment of PH20 are deleted in any one of paragraphs 50 to 56.

[0371] 58. In paragraph 57, the cut

[0372] (i) an amino acid residue selected from the group consisting of M1 to P42 at the N-terminus such that one or more residues are deleted at the N-terminus;

[0373] (ii) preceding amino acid residues L36, N37, F38, R39, A40, P41, or P42 at the N-terminus so as to delete one or more residues at the N-terminus;

[0374] (iii) located after an amino acid residue selected from the group consisting of V455 to L509 at the C-terminus such that one or more amino acid residues are deleted at the C-terminus;

[0375] (iv) a method having one or more amino acid residues deleted at the C-terminus, having an amino acid residue selected from V455, C458, D461, C464, I465, D466, A467, F468, K470, P471, P472, M473, E474, T475, E476, E477, P478, Q479, I480, F481, Y482, N483, A484, P486, T488, or S490 located after the C-terminus.

[0376] 59. A method comprising, in any one of paragraphs 50 to 58, a human growth hormone-derived signal peptide having the amino acid sequence MATGSRTSLLLAFGLLCLPWLQEGSA of SEQ NO. 3, a human serum albumin-derived signal peptide having the amino acid sequence MKWVTFISLLFLFSSAYS of SEQ NO. 4, or a human Hyal1-derived signal peptide having the amino acid sequence MAAHLLPICALFLTLLDMAQG of SEQ NO. 5.

[0377] 60. In any one of paragraphs 50 to 59, a variant or fragment of PH20

[0378] (i) a peptide having at least 90% sequence identity with the amino acid sequence presented as SEQ NO. 1 or amino acid residues 36-482, 36-477, 366-478, 36-479, 36-480, 36-481, and 36-483 of SEQ NO. 1;

[0379] (ii) a peptide having at least 95% sequence identity with the amino acid sequence presented as SEQ NO. 1 or amino acid residues 36-482, 36-477, 366-478, 36-479, 36-480, 36-481, and 36-483 of SEQ NO. 1;

[0380] (iii) composed of an amino acid sequence selected from sequence numbers 60 to 115;

[0381] (iv) A method having the amino acid sequence of sequence number 99.

[0382] 61. A method in which hyaluronidase is present in the composition in an amount of 100 U to 50,000 U in any one of paragraphs 43 to 60.

[0383] 62. A method in which hyaluronidase is present in the composition in an amount of 100 U / mL to 50,000 U / mL in any one of paragraphs 43 to 61.

[0384] 63. A method in any one of paragraphs 43 to 62, wherein the composition further comprises one or more pharmaceutically acceptable excipients.

[0385] 64. A method in any one of paragraphs 43 to 63, wherein the composition further comprises one or more sugars.

[0386] 65. A method according to paragraph 64, wherein one or more sugars comprise a monosaccharide.

[0387] 66. A method in any one of paragraphs 64 to 65, wherein one or more sugars comprise a polysaccharide.

[0388] 67. A method according to paragraph 66, wherein one or more polysaccharides are selected from the group consisting of trehalose and sucrose.

[0389] 68. A method in which, in any one of paragraphs 64 to 67, one or more sugars are present in the composition in an amount of 10 mM to 500 mM, optionally in an amount of 100 mM to 300 mM.

[0390] 69. A method in any one of paragraphs 43 to 68, wherein the composition further comprises one or more amino acids.

[0391] 70. A method according to paragraph 69 in which the amino acid is selected from methionine and histidine.

[0392] 71. A method in which, in any one of paragraphs 69 to 70, one or more amino acids are present in the composition in an amount of 1 mM to 100 mM, optionally in an amount of 1 mM to 50 mM.

[0393] 72. A method in any one of paragraphs 43 to 71, wherein the composition further comprises a buffer.

[0394] 73. The method of paragraph 72, wherein a buffer is present in the composition in an amount sufficient to maintain the composition at a pH of 3.0 to 9.0, optionally in an amount sufficient to maintain the composition at a pH of 5.5 to 7.5.

[0395] 74. A method in which, in any one of paragraphs 72 to 73, a buffer is present in the composition in an amount of 1 to 100 mM, optionally in an amount of 1 mM to 50 mM.

[0396] 75. A method in any one of paragraphs 72 to 74, wherein the buffer comprises histidine.

[0397] 76. A method in any one of paragraphs 43 to 75, wherein the oligonucleotide of the telomerase inhibitor comprises at least one N3' → P5' thiophosphoramidate nucleoside linkage.

[0398] 77. A method in any one of paragraphs 43 to 76, wherein the lipid moiety of the telomerase inhibitor is connected to the 5' and / or 3' ends of an oligonucleotide through a linker.

[0399] 78. The method of paragraph 77, wherein the linker is a glycerol or aminoglycerol linker.

[0400] 79. A method in which, in any one of paragraphs 43 to 78, the lipid moiety of the telomerase inhibitor is a palmitoyl (C16) moiety.

[0401] 80. A method in any one of paragraphs 43 to 79 and 158, wherein the telomerase inhibitor is imetelstat or a pharmaceutically acceptable salt thereof, optionally imetelstat sodium.

[0402] 81. A unit dosage form comprising a hyaluronidase enzyme and a telomerase inhibitor comprising an oligonucleotide and a lipid moiety linked to the 5' and / or 3' ends of the oligonucleotide.

[0403] 82. In paragraph 81, the hyaluronidase enzyme is recombinant human hyaluronidase, unit dosage form.

[0404] 83. In paragraph 82, a unit dosage form in which the hyaluronidase enzyme is rHuPH20.

[0405] 84. In paragraph 83, a unit dosage form wherein the composition comprises a variant or fragment of a pH 20 hyaluronidase enzyme.

[0406] 85. In paragraph 84, a variant or fragment of PH20

[0407] (i) one or more amino acid residue substitutions selected from the group consisting of T341A, T341C, T341G, S343E, M345T, K349E, L353A, L354I, N356E and I361T in wild-type PH20 having the amino acid sequence of SEQ No. 1;

[0408] (ii) one or more amino acid residue substitutions selected from the group consisting of T341A, T341C, L354I, and N356E; or

[0409] (iii) A unit dosage form comprising one or more amino acid residue substitutions in a region corresponding to the alpha-helical region or linker region in wild-type PH20 having the amino acid sequence of SEQ No. 1.

[0410] 86. A unit dosage type according to paragraph 85, wherein the alpha-helical region is an alpha-helical 8 region containing amino acid residues S347 to C381 and the linker region is a linker region containing amino acid residues A333 to R346 between alpha-helical 7 and alpha-helical 8.

[0411] 87. A unit dosage form according to paragraph 85, wherein the alpha-helical region and the linker region comprise amino acid residues T341 to N363, T341 to I361, L342 to I361, S343 to I361, I344 to I361, M345 to I361, or M345 to N363.

[0412] 88. A unit dosage form according to paragraph 86, wherein the alpha-helical 8 region and the linker region between alpha-helical 7 and alpha-helical 8 are substituted with one or more amino acid residues of the corresponding region of Hyal1.

[0413] 89. In paragraph 84, a unit dosage form in which a variant or fragment of PH20 comprises the following:

[0414] (i) one or more amino acid residue substitutions at one or more positions selected from the group consisting of T341, L342, S343, I344, M345, S347, M348, K349, L352, L353, D355, E359, I361 and N363;

[0415] (ii) Substitution of the following amino acid residues:

[0416] One or more of L354I and N356E; and

[0417] One or more amino acid residue substitutions selected from the group consisting of T341A, T341C, T341D, T341G, T341S, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, D355K, E359D, I361T, and N363G, wherein optionally, a variant or fragment of PH20 comprises the substitutions M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D, and I361T;

[0418] (iii) Substitutions T341A, T341C, T341D, T341G, T341S, L342W, S343E, I344N and N363G; or

[0419] (iv) Any one amino acid residue substitution selected from the following amino acid residue substitution groups:

[0420] (a) T341S, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D, and I361T;

[0421] (b) L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D, and I361T;

[0422] (c) M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D, I361T, and N363G;

[0423] (d) T341G, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D, and I361T;

[0424] (e) T341A, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D, and I361T;

[0425] (f) T341C, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D, and I361T;

[0426] (g) T341D, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D, and I361T;

[0427] (h) I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D and I361T; and

[0428] (i) S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D and I361T.

[0429] 90. A unit dosage form in which one or more of the N-terminal or C-terminal amino acid residues of a variant or fragment of PH20 are deleted, in any one of paragraphs 84 to 89.

[0430] 91. In paragraph 90, the cut

[0431] (i) an amino acid residue selected from the group consisting of M1 to P42 at the N-terminus such that one or more residues are deleted at the N-terminus;

[0432] (ii) preceding amino acid residues L36, N37, F38, R39, A40, P41, or P42 at the N-terminus so as to delete one or more residues at the N-terminus;

[0433] (iii) after an amino acid residue selected from the group consisting of V455 to L509 at the C-terminus such that one or more amino acid residues are deleted at the C-terminus;

[0434] (iv) A unit dosage form located after an amino acid residue selected from V455, C458, D461, C464, I465, D466, A467, F468, K470, P471, P472, M473, E474, T475, E476, E477, P478, Q479, I480, F481, Y482, N483, A484, P486, T488, or S490 at the C-terminus, such that one or more amino acid residues are deleted at the C-terminus.

[0435] 92. A unit dosage form comprising, in any one of paragraphs 84 to 91, a human growth hormone-derived signal peptide having the amino acid sequence MATGSRTSLLLAFGLLCLPWLQEGSA of SEQ No. 3, a human serum albumin-derived signal peptide having the amino acid sequence MKWVTFISLLFLFSSAYS of SEQ No. 4, or a human Hyal1-derived signal peptide having the amino acid sequence MAAHLLPICALFLTLLDMAQG of SEQ No. 5.

[0436] 93. In any one of paragraphs 84 to 92, a variant or fragment of PH20

[0437] (i) a peptide having at least 90% sequence identity with the amino acid sequence presented as SEQ NO. 1 or amino acid residues 36-482, 36-477, 366-478, 36-479, 36-480, 36-481, and 36-483 of SEQ NO. 1;

[0438] (ii) a peptide having at least 95% sequence identity with the amino acid sequence presented as SEQ NO. 1 or amino acid residues 36-482, 36-477, 366-478, 36-479, 36-480, 36-481, and 36-483 of SEQ NO. 1;

[0439] (iii) composed of an amino acid sequence selected from sequence numbers 60 to 115;

[0440] (iv) A unit dosage form having the amino acid sequence of sequence number 99.

[0441] 94. A unit dosage form in which hyaluronidase is present in the composition in an amount of 100 U to 50,000 U in any one of paragraphs 81 to 93.

[0442] 95. A unit dosage form in which hyaluronidase is present in the composition in an amount of 100 U / mL to 50,000 U / mL in any one of paragraphs 81 to 94.

[0443] 96. A unit dosage form in which, in any one of paragraphs 81 to 95, the composition further comprises one or more pharmaceutically acceptable excipients.

[0444] 97. A unit dosage form, wherein the composition further comprises one or more sugars in any one of paragraphs 81 to 96.

[0445] 98. In paragraph 97, a unit dosage form in which one or more sugars comprise a monosaccharide.

[0446] 99. A unit dosage form in which, in any one of paragraphs 97 to 98, one or more sugars comprise a polysaccharide.

[0447] 100. A unit dosage form according to paragraph 99, wherein one or more polysaccharides are selected from the group consisting of trehalose and sucrose.

[0448] 101. A unit dosage form in which, in any one of paragraphs 97 to 100, one or more sugars are present in the composition in an amount of 10 mM to 500 mM.

[0449] 102. A unit dosage form in which, in any one of paragraphs 81 to 100, the composition further comprises one or more amino acids.

[0450] 103. A unit dosage form of paragraph 102, wherein the amino acid is selected from methionine and histidine.

[0451] 104. A unit dosage form in which, in any one of paragraphs 102 to 103, one or more amino acids are present in the composition in an amount of 1 mM to 100 mM, optionally in an amount of 1 mM to 50 mM.

[0452] 105. A unit dosage type in which, in any one of paragraphs 81 to 104, the composition further comprises a buffer.

[0453] 106. A unit dosage form according to paragraph 105, wherein the buffer is present in the composition in an amount sufficient to maintain the composition at a pH of 3.0 to 9.0, optionally in an amount sufficient to maintain the composition at a pH of 5.5 to 7.5.

[0454] 107. A unit dosage form in which, in any one of paragraphs 105 to 106, the buffer is present in the composition in an amount of 1 to 100 mM, optionally in an amount of 1 mM to 50 mM.

[0455] 108. A unit dosage form in which, in any one of paragraphs 81 to 107, the oligonucleotide of the telomerase inhibitor comprises at least one N3' → P5' thiophosphoramidate nucleoside linkage.

[0456] 109. A unit dosage form in any one of paragraphs 81 to 108, wherein the lipid moiety of the telomerase inhibitor is connected to the 5' and / or 3' ends of an oligonucleotide via a linker.

[0457] 110. In paragraph 109, a unit dosage form in which the linker is a glycerol or aminoglycerol linker.

[0458] 111. A unit dosage form in which, in any one of paragraphs 81 to 110, the lipid moiety of the telomerase inhibitor is a palmitoyl (C16) moiety.

[0459] 112. In any one of paragraphs 81 to 111, a unit dosage form in which the telomerase inhibitor is imetelstat or a pharmaceutically acceptable salt thereof, optionally imetelstat sodium.

[0460] 113. A unit dosage form in which, in any one of paragraphs 81 to 112, a telomerase inhibitor is present in the composition at the following dosage:

[0461] (i) about 2.0 mg / kg to 20.0 mg / kg;

[0462] (ii) about 3 mg / kg to about 15 mg / kg;

[0463] (iii) about 9 mg / kg to about 11 mg / kg; or

[0464] (iv) About 11 mg / kg to about 14 mg / kg.

[0465] 114. A unit dosage type in which the composition is a liquid, in any one of paragraphs 81 to 113.

[0466] 115. Kit including the following:

[0467] A composition comprising hyaluronidase enzyme, and

[0468] A composition comprising a telomerase inhibitor comprising an oligonucleotide and a lipid moiety linked to the 5' and / or 3' ends of the oligonucleotide.

[0469] 116. A kit according to paragraph 115, additionally comprising an injection device.

[0470] 117. A kit in which the composition comprising a telomerase inhibitor is lyophilized, in any one of paragraphs 115 to 116.

[0471] 118. A kit further comprising a buffer for producing a reconstituted liquid composition in any one of paragraphs 115 to 117.

[0472] 119. A kit in any one of paragraphs 115 to 118, wherein the subcutaneous injection device comprises a needle and a syringe.

[0473] 120. A kit in any one of paragraphs 115 to 119, wherein the subcutaneous injection device is a bolus injection device configured to deliver a predetermined amount of a composition subcutaneously.

[0474] 121. In any one of paragraphs 115 to 120, the hyaluronidase enzyme is recombinant human hyaluronidase, kit.

[0475] 122. In paragraph 121, the kit in which the hyaluronidase enzyme is rHuPH20.

[0476] 123. A kit in any one of paragraphs 115 to 121, wherein the composition comprises a variant or fragment of a pH 20 hyaluronidase enzyme.

[0477] 124. In paragraph 123, a kit in which a variant or fragment of PH20 comprises the following:

[0478] (i) one or more amino acid residue substitutions selected from the group consisting of T341A, T341C, T341G, S343E, M345T, K349E, L353A, L354I, N356E and I361T in wild-type PH20 having the amino acid sequence of SEQ No. 1;

[0479] (ii) one or more amino acid residue substitutions selected from the group consisting of T341A, T341C, L354I and N356E;

[0480] (iii) Substitution of one or more amino acid residues in the region corresponding to the alpha-helical region or linker region in wild-type PH20 having the amino acid sequence of SEQ No. 1.

[0481] 125. A kit according to paragraph 124, wherein the alpha-helical region is an alpha-helical 8 region containing amino acid residues S347 to C381 and the linker region is a linker region containing amino acid residues A333 to R346 between alpha-helical 7 and alpha-helical 8.

[0482] 126. A kit according to paragraph 124, wherein the alpha-helical region and the linker region comprise amino acid residues T341 to N363, T341 to I361, L342 to I361, S343 to I361, I344 to I361, M345 to I361, or M345 to N363.

[0483] 127. In paragraph 124, the alpha-helix 8 region and the linker region between alpha-helix 7 and alpha-helix 8 are substituted with one or more amino acid residues of the corresponding region of Hyal1.

[0484] 128. A kit according to paragraph 123, wherein a variant or fragment of PH20 comprises one or more amino acid residue substitutions at one or more positions selected from the group consisting of T341, L342, S343, I344, M345, S347, M348, K349, L352, L353, D355, E359, I361 and N363.

[0485] 129. In paragraph 128, a kit in which a variant or fragment of PH20 comprises the following:

[0486] (i) Substitution of the following amino acid residues:

[0487] One or more of L354I and N356E; and

[0488] One or more amino acid residue substitutions selected from the group consisting of T341A, T341C, T341D, T341G, T341S, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, D355K, E359D, I361T, and N363G;

[0489] (ii) Substitutions M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D and I361T;

[0490] (iii) Substitutions T341A, T341C, T341D, T341G, T341S, L342W, S343E, I344N and N363G; or

[0491] (iv) Any one amino acid residue substitution selected from the following amino acid residue substitution groups:

[0492] (a) T341S, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D, and I361T;

[0493] (b) L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D, and I361T;

[0494] (c) M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D, I361T, and N363G;

[0495] (d) T341G, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D, and I361T;

[0496] (e) T341A, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D, and I361T;

[0497] (f) T341C, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D, and I361T;

[0498] (g) T341D, L342W, S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D, and I361T;

[0499] (h) I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D and I361T; and

[0500] (i) S343E, I344N, M345T, S347T, M348K, K349E, L352Q, L353A, L354I, D355K, N356E, E359D and I361T.

[0501] 130. A kit in which one or more of the N-terminal or C-terminal amino acid residues of a variant or fragment of PH20 are deleted, in any one of paragraphs 123 to 129.

[0502] 131. A kit according to paragraph 130, wherein the cleavage is positioned before an amino acid residue selected from the group consisting of M1 to P42 at the N-terminus such that one or more residues at the N-terminus are deleted.

[0503] 132. In paragraph 131, the cut

[0504] (i) preceding amino acid residues L36, N37, F38, R39, A40, P41, or P42 at the N-terminus so as to delete one or more residues at the N-terminus;

[0505] (ii) after an amino acid residue selected from the group consisting of V455 to L509 at the C-terminus such that one or more amino acid residues are deleted at the C-terminus; or

[0506] (iii) Kit located after an amino acid residue selected from V455, C458, D461, C464, I465, D466, A467, F468, K470, P471, P472, M473, E474, T475, E476, E477, P478, Q479, I480, F481, Y482, N483, A484, P486, T488, or S490 at the C-terminus, such that one or more amino acid residues are deleted at the C-terminus.

[0507] 133. A kit comprising, in any one of paragraphs 123 to 132, a human growth hormone-derived signal peptide having the amino acid sequence MATGSRTSLLLAFGLLCLPWLQEGSA of SEQ No. 3, a human serum albumin-derived signal peptide having the amino acid sequence MKWVTFISLLFLFSSAYS of SEQ No. 4, or a human Hyal1-derived signal peptide having the amino acid sequence MAAHLLPICALFLTLLDMAQG of SEQ No. 5.

[0508] 134. In any one of paragraphs 123 to 133, a variant or fragment of PH20

[0509] (i) a peptide having at least 90% sequence identity with the amino acid sequence presented as SEQ NO. 1 or amino acid residues 36-482, 36-477, 366-478, 36-479, 36-480, 36-481, and 36-483 of SEQ NO. 1;

[0510] (ii) a peptide having at least 95% sequence identity with the amino acid sequence presented as SEQ NO. 1 or amino acid residues 36-482, 36-477, 366-478, 36-479, 36-480, 36-481, and 36-483 of SEQ NO. 1;

[0511] (iii) composed of an amino acid sequence selected from sequence numbers 60 to 115;

[0512] (iv) A kit having the amino acid sequence of sequence number 99.

[0513] 135. A kit in which hyaluronidase is present in the composition in an amount of 100 U to 50,000 U, in any one of paragraphs 115 to 134.

[0514] 136. A kit in which hyaluronidase is present in the composition in an amount of 100 U / mL to 50,000 U / mL in any one of paragraphs 115 to 135.

[0515] 137. A kit in any one of paragraphs 115 to 136, wherein the composition further comprises one or more pharmaceutically acceptable excipients.

[0516] 138. A kit in any one of paragraphs 115 to 137, wherein the composition further comprises one or more sugars.

[0517] 139. A kit according to paragraph 138, wherein one or more sugars comprise a monosaccharide.

[0518] 140. A kit in any one of paragraphs 138 to 139, wherein one or more sugars comprise a polysaccharide.

[0519] 141. A kit according to paragraph 140, wherein one or more polysaccharides are selected from the group consisting of trehalose and sucrose.

[0520] 142. A kit in which, in any one of paragraphs 140 to 141, one or more polysaccharides are present in the composition in an amount of 10 mM to 500 mM, optionally in an amount of 100 mM to 300 mM.

[0521] 143. A kit in any one of paragraphs 115 to 142, wherein the composition further comprises one or more amino acids.

[0522] 144. A kit according to paragraph 143, wherein the composition comprises methionine.

[0523] 145. A kit in which, in any one of paragraphs 143 to 144, one or more amino acids are present in the composition in an amount of 1 mM to 100 mM, optionally in an amount of 1 mM to 50 mM.

[0524] 146. A kit in any one of paragraphs 115 to 145, wherein the composition further comprises a buffer.

[0525] 147. The kit according to paragraph 146, wherein the buffer is present in the composition in an amount sufficient to maintain the composition at pH 3.0 to 9.0, optionally in an amount sufficient to maintain the composition at pH 5.5 to 7.5.

[0526] 148. A kit in which, in any one of paragraphs 146 to 147, a buffer is present in the composition in an amount of 1 to 100 mM, optionally in an amount of 1 mM to 50 mM.

[0527] 149. A kit in any one of paragraphs 146 to 148, wherein the buffer comprises histidine.

[0528] 150. A kit in any one of paragraphs 115 to 149, wherein the oligonucleotide of the telomerase inhibitor comprises at least one N3' → P5' thiophosphoramidate nucleoside linkage.

[0529] 151. A kit in any one of paragraphs 115 to 150, wherein the lipid moiety of the telomerase inhibitor is connected to the 5' and / or 3' ends of an oligonucleotide through a linker.

[0530] 152. A kit according to paragraph 151, wherein the linker is a glycerol or aminoglycerol linker.

[0531] 153. In any one of paragraphs 115 to 152, the lipid moiety of the telomerase inhibitor is a palmitoyl (C16) moiety.

[0532] 154. A kit in which, in any one of paragraphs 115 to 153, the telomerase inhibitor is imetelstat or a pharmaceutically acceptable salt thereof, optionally imetelstat sodium.

[0533] 155. In any one of paragraphs 115 to 154, a kit in which a telomerase inhibitor is present in the composition at the following dosage:

[0534] (i) about 2.0 mg / kg to 20.0 mg / kg;

[0535] (ii) about 3 mg / kg to about 15 mg / kg;

[0536] (iii) about 9 mg / kg to about 11 mg / kg; or

[0537] (iv) About 11 mg / kg to about 14 mg / kg.

[0538] 156. In any one of paragraphs 115 to 154, a kit in which a telomerase inhibitor is present in the composition at the following dosage:

[0539] (i) Approximately 200 mg to 3000 mg;

[0540] (ii) about 750 mg to about 2500 mg;

[0541] (iii) about 1,000 mg to about 2,000 mg; or

[0542] (iv) About 500 mg to about 2000 mg.

[0543] 157. In any one of paragraphs 1 to 40, a composition in which a telomerase inhibitor is present in the composition at the following dosage:

[0544] (i) Approximately 200 mg to 3000 mg;

[0545] (ii) about 750 mg to about 2500 mg;

[0546] (iii) about 1,000 mg to about 2,000 mg; or

[0547] (iv) About 500 mg to about 2000 mg.

[0548] 158. Method in any one of paragraphs 43 to 47, wherein a telomerase inhibitor is present in the composition at the following dosage:

[0549] (i) Approximately 200 mg to 3000 mg;

[0550] (ii) about 750 mg to about 2500 mg;

[0551] (iii) about 1,000 mg to about 2,000 mg; or

[0552] (iv) About 500 mg to about 2000 mg.

[0553] 159. Method in any one of paragraphs 81 to 112, wherein a telomerase inhibitor is present in the composition at the following dosage:

[0554] (i) Approximately 200 mg to 3000 mg;

[0555] (ii) about 750 mg to about 2500 mg;

[0556] (iii) about 1,000 mg to about 2,000 mg; or

[0557] (iv) About 500 mg to about 2000 mg.

Claims

Claim 1 1,000 mg to 3,000 mg of imetelstat or a pharmaceutically acceptable salt thereof; and a co-formulation for subcutaneous administration comprising rHuPH20. Claim 2 A co-formulation according to claim 1, characterized in that rHuPH20 is present in the co-formulation in an amount of 100 U to 50,000 U. Claim 3 A co-formulation according to claim 1, wherein the co-formulation further comprises one or more pharmaceutically acceptable excipients; or the co-formulation further comprises one or more sugars; or the co-formulation further comprises one or more amino acids; or the co-formulation further comprises a buffer. Claim 4 A co-formulation according to claim 3, characterized in that one or more sugars are present in the co-formulation in an amount of 10 mM to 500 mM. Claim 5 A co-formulation characterized in that, in claim 3, one or more amino acids are selected from methionine and histidine. Claim 6 A co-formulation according to claim 3, characterized in that one or more amino acids are present in the co-formulation in an amount of 1 mM to 100 mM. Claim 7 A co-formulation according to claim 3, characterized in that the buffer is present in the co-formulation in an amount of 1 mM to 100 mM. Claim 8 A co-formulation according to claim 1, characterized in that imetelstat or a pharmaceutically acceptable salt thereof is imetelstat sodium. Claim 9 A co-formulation according to claim 1, characterized in that imetelstat or a pharmaceutically acceptable salt thereof is administered to a subject in an amount of 13.0 mg / kg to 20.0 mg / kg; or imetelstat or a pharmaceutically acceptable salt thereof is administered to a subject in an amount of 1000 mg to 2500 mg, 1200 mg to 3000 mg, 1200 mg to 2500 mg, 1500 mg to 3000 mg, or 1500 mg to 2500 mg. Claim 10 A co-formulation according to claim 1, characterized in that the co-formulation is freeze-dried. Claim 11 A joint formulation according to claim 1, characterized in that the joint formulation is a liquid formulation. Claim 12 A pharmaceutical composition for treating hematological neoplasms in a subject requiring treatment, comprising a co-formulation of any one of claims 1 to 11. Claim 13 A pharmaceutical composition according to claim 12, characterized in that the hematological neoplasm is selected from myelofibrosis (MF), myelodysplastic syndrome (MDS), essential thrombocytosis (ET), polycythemia vera (PV), chronic myeloid leukemia (CML), chronic neutrophilic leukemia, chronic eosinophilic leukemia, and acute myeloid leukemia (AML). Claim 14 A pharmaceutical composition according to claim 12, characterized in that the subject is diagnosed as having a hematological neoplasm. Claim 15 A pharmaceutical composition according to claim 12, characterized in that the co-formulation is administered subcutaneously to a subject once every 7 days, once every 21 days, or once every 28 days; or repeated one or more times. Claim 16 A pharmaceutical composition according to claim 12, characterized in that imetelstat or a pharmaceutically acceptable salt thereof is administered to a subject in an amount of 13.0 mg / kg to 20.0 mg / kg; or imetelstat or a pharmaceutically acceptable salt thereof is administered to a subject in an amount of 1000 mg to 2500 mg, 1200 mg to 3000 mg, 1200 mg to 2500 mg, 1500 mg to 3000 mg, or 1500 mg to 2500 mg. Claim 17 A kit for treating hematological neoplasms comprising a co-formulation of any one of claims 1 to 11. Claim 18 In claim 17, the kit is characterized by additionally including a subcutaneous injection device. Claim 19 In claim 17, the kit is characterized by further comprising a buffer for producing a reconstituted liquid co-formulation. Claim 20 A kit according to claim 18, characterized in that the subcutaneous injection device comprises a needle and a syringe, or the subcutaneous injection device is a bolus injection device configured to deliver a predetermined amount of a common formulation subcutaneously. Claim 21 delete Claim 22 delete Claim 23 delete Claim 24 delete Claim 25 delete Claim 26 delete Claim 27 delete Claim 28 delete Claim 29 delete Claim 30 delete Claim 31 delete Claim 32 delete Claim 33 delete Claim 34 delete Claim 35 delete Claim 36 delete Claim 37 delete Claim 38 delete Claim 39 delete Claim 40 delete Claim 41 delete Claim 42 delete Claim 43 delete Claim 44 delete Claim 45 delete Claim 46 delete Claim 47 delete Claim 48 delete Claim 49 delete Claim 50 delete Claim 51 delete Claim 52 delete Claim 53 delete Claim 54 delete Claim 55 delete Claim 56 delete Claim 57 delete Claim 58 delete Claim 59 delete Claim 60 delete Claim 61 delete Claim 62 delete Claim 63 delete Claim 64 delete Claim 65 delete Claim 66 delete Claim 67 delete Claim 68 delete Claim 69 delete Claim 70 delete Claim 71 delete Claim 72 delete Claim 73 delete Claim 74 delete Claim 75 delete Claim 76 delete Claim 77 delete Claim 78 delete Claim 79 delete Claim 80 delete Claim 81 delete Claim 82 delete Claim 83 delete Claim 84 delete Claim 85 delete Claim 86 delete Claim 87 delete Claim 88 delete Claim 89 delete Claim 90 delete Claim 91 delete Claim 92 delete Claim 93 delete Claim 94 delete Claim 95 delete Claim 96 delete Claim 97 delete Claim 98 delete Claim 99 delete Claim 100 delete Claim 101 delete Claim 102 delete Claim 103 delete Claim 104 delete Claim 105 delete Claim 106 delete Claim 107 delete Claim 108 delete Claim 109 delete Claim 110 delete Claim 111 delete Claim 112 delete Claim 113 delete Claim 114 delete Claim 115 delete Claim 116 delete Claim 117 delete Claim 118 delete Claim 119 delete Claim 120 delete Claim 121 delete Claim 122 delete Claim 123 delete Claim 124 delete Claim 125 delete Claim 126 delete Claim 127 delete Claim 128 delete Claim 129 delete Claim 130 delete Claim 131 delete Claim 132 delete Claim 133 delete Claim 134 delete Claim 135 delete Claim 136 delete

Images