Methods of administering chimeric antigen receptor immunotherapy

Abstract

To provide a method of treating relapsed or refractory large B-cell lymphoma, such as diffuse large B-cell lymphoma (DLBCL) not otherwise specified, primary mediastinal large B-cell lymphoma, high grade B-cell lymphoma, and DLBCL arising from follicular lymphoma, after two or more lines of systemic therapy.SOLUTION: In one aspect, provided is a method of treating relapsed or refractory large B-cell lymphoma in a patient, after two or more lines of systemic therapy, the method comprising: (a) administering a CD19-directed genetically modified autologous T-cell immunotherapy to a patient in need of treatment; and (b) monitoring the patient following the infusion for signs and symptoms of adverse reactions.SELECTED DRAWING: None

Description

CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the benefit of priority to U.S. Provisional Patent Application No. 62 / 944,903, filed December 6, 2019, U.S. Provisional Patent Application No. 62 / 931,669, filed November 6, 2019, U.S. Provisional Patent Application No. 62 / 868,262, filed June 28, 2019, U.S. Provisional Patent Application No. 62 / 855,828, filed May 31, 2019, and U.S. Provisional Patent Application No. 62 / 843,190, filed May 3, 2019. [Technical field]

[0002] The present disclosure relates generally to T cell therapy, and more specifically to CD19-directed genetically modified autologous T cell immunotherapy, including chimeric antigen receptors (CARs). [Background technology]

[0003] Human cancers, by their very nature, are composed of normal cells that have undergone genetic or epigenetic transformation to become abnormal cancer cells. In doing so, the cancer cells begin to express proteins and other antigens that differ from those expressed by normal cells. These abnormal tumor antigens can be used by the body's natural immune system to specifically target and kill the cancer cells. However, cancer cells use various mechanisms to prevent immune cells, such as T and B lymphocytes, from successfully targeting the cancer cells.

[0004] Chimeric antigen receptors (CARs), which contain a binding domain that can interact with a specific tumor antigen, allow T cells to target and kill cancer cells that express that specific tumor antigen. Summary of the Invention

[0005] As described in detail below, the present disclosure is based in part on the surprising discovery that the administration methods disclosed herein identify and manage adverse side effects and safety of CAR-T cell immunotherapy. Additionally, the present disclosure relates to immunotherapy or T cell therapy, as well as methods for improving the outcome and / or response of the treatment.

[0006] Any aspect or embodiment described herein can be combined with any other aspect or embodiment disclosed herein. While the present disclosure has been described in conjunction with its detailed description, the foregoing description is intended to be illustrative and not limiting of the scope of the present disclosure, which is defined by the appended claims. Other aspects, advantages, and modifications are within the scope of the following embodiments / claims.

[0007] In one aspect, the disclosure provides a method of treating relapsed or refractory diffuse large B-cell lymphoma (DLBCL) not otherwise specified, primary mediastinal large B-cell lymphoma, high-grade B-cell lymphoma, or DLBCL arising from follicular lymphoma after two or more lines of systemic therapy in a patient, comprising administering to a patient in need of treatment about 1×10 per kg of body weight. 6 ~Approx. 2×10 6 1 x 10 CAR-positive viable T cells at a maximum dose 8 The method includes administering a suspension of axicabtagene ciloleucel by intravenous infusion at a dose of up to 1000 CAR-positive viable T cells, where axicabtagene ciloleucel is a CD19-directed genetically modified autologous T cell immunotherapy, comprising harvested patient's own T cells genetically modified ex vivo by retroviral transduction to express a chimeric antigen receptor (CAR) comprising an anti-CD19 single chain variable fragment (scFv) linked to the costimulatory domains of CD28 and CD3 zeta.

[0008] In another aspect, the disclosure provides a method of treating relapsed or refractory diffuse large B-cell lymphoma (DLBCL) and primary mediastinal large B-cell lymphoma (PMBCL) in a patient after two or more lines of systemic therapy, comprising administering to a patient in need of treatment about 0.4×10 8~Approx. 2×10 8 The method includes administering a suspension of axicabtagene ciloleucel by intravenous infusion at a dose of 100 CAR-positive viable T cells, where axicabtagene ciloleucel is a CD19-directed genetically modified autologous T cell immunotherapy comprising harvested patient's own T cells genetically modified ex vivo by retroviral transduction to express a chimeric antigen receptor (CAR) comprising an anti-CD19 single chain variable fragment (scFv) linked to the costimulatory domains of CD28 and CD3 zeta.

[0009] In some embodiments, the intravenous infusion time is 15-120 minutes. In some embodiments, the intravenous infusion time is up to 30 minutes. In some embodiments, the injection volume is 50-100 mL. In some embodiments, the injection volume is about 68 mL. In some embodiments, the immunotherapy is injected from an infusion bag. In some embodiments, the infusion bag is agitated during injection. In some embodiments, the immunotherapy is administered within 3 hours after thawing.

[0010] In some embodiments, the suspension further comprises albumin. In some embodiments, the albumin is present in an amount of about 2-3% by volume. In some embodiments, the albumin is present in an amount of about 2.5% by volume. In some embodiments, the albumin is human albumin. In some embodiments, the suspension further comprises DMSO. In some embodiments, the DMSO is present in an amount of about 4-6% by volume. In some embodiments, the DMSO is present in an amount of about 5% by volume.

[0011] In one aspect, the disclosure provides a method of treating relapsed or refractory large B-cell lymphoma in a patient after two or more lines of systemic therapy, the method comprising: (a) administering a CD19-directed genetically modified autologous T-cell immunotherapy to a patient in need of treatment, and (b) monitoring the patient for signs and symptoms of adverse reactions after the infusion. In some embodiments, the relapsed or refractory large B-cell lymphoma is diffuse large B-cell lymphoma (DLBCL) not otherwise specified, primary mediastinal large B-cell lymphoma, high-grade B-cell lymphoma, or DLBCL arising from follicular lymphoma.

[0012] In some embodiments, the adverse reaction is selected from the group consisting of cytokine release syndrome (CRS), neurotoxicity, hypersensitivity reactions, serious infections, cytopenias, and hypogammaglobulinemia. In some embodiments, the signs and symptoms of the adverse reaction are selected from the group consisting of fever, hypotension, tachycardia, hypoxia, and chills, including cardiac arrhythmias (including atrial fibrillation and ventricular tachycardia), cardiac arrest, cardiac failure, renal insufficiency, capillary leak syndrome, hypotension, hypoxia, organ toxicity, hemophagocytic lymphohistiocytosis / macrophage activation syndrome (HLH / MAS), seizures, encephalopathy, headache, tremor, dizziness, aphasia, delirium, insomnia anxiety, anaphylaxis, febrile neutropenia, thrombocytopenia, neutropenia, and anemia.

[0013] In some embodiments, the method further comprises administering an IL-6 receptor inhibitor. In some embodiments, the method further comprises administering an effective amount of tocilizumab to treat symptoms of the adverse reaction. In some embodiments, tocilizumab is administered intravenously at a dose of about 8 mg / kg. In some embodiments, tocilizumab is administered intravenously over about 1 hour. In some embodiments, tocilizumab is administered about every 8 hours. In some embodiments, tocilizumab is administered within about a 24 hour period.

[0014] In some embodiments, the methods further include administering a steroid (e.g., a corticosteroid to treat symptoms of an adverse reaction) or to improve safety. In some embodiments, the corticosteroid is at least one of methylprednisone or dexamethasone.

[0015] In some embodiments, methylprednisone is administered intravenously at a dose of about 1 mg / kg. In some embodiments, methylprednisone is administered twice daily. In some embodiments, methylprednisone is administered intravenously at a dose of about 1,000 mg / day. In some embodiments, methylprednisone is administered intravenously for about 3 days. In some embodiments, dexamethasone is administered at a dose of about 10 mg. In some embodiments, dexamethasone is administered intravenously about every 6 hours.

[0016] In some embodiments, the adverse reaction is cytokine release syndrome (CRS). In some embodiments, monitoring for signs and symptoms of cytokine release syndrome (CRS) occurs at least daily for about 7 days following the infusion. In some embodiments, monitoring for signs and symptoms of cytokine release syndrome (CRS) occurs at least daily for about 8 days, about 9 days, or about 10 days following the infusion. In some embodiments, monitoring for signs and symptoms of cytokine release syndrome (CRS) occurs at least daily for about 10 days following the infusion. In some embodiments, monitoring for signs and symptoms of cytokine release syndrome (CRS) occurs for about 4 weeks following the infusion.

[0017] In some embodiments, the adverse reaction is neurotoxicity, hi some embodiments, monitoring for signs and symptoms of neurotoxicity is for up to about 8 weeks after infusion.

[0018] In some embodiments, the method further comprises administering a non-sedating anti-seizure drug for seizure prevention, hi some embodiments, the non-sedating anti-seizure drug is levetiracetam.

[0019] In some embodiments, the adverse reaction is a cytopenia. In some embodiments, the cytopenia is thrombocytopenia, neutropenia, and / or anemia.

[0020] In some embodiments, the method further comprises administering at least one of erythropoietin, darbepoetin alfa, platelet transfusion, colony stimulating factor (CSF), granulocyte colony stimulating factor, filgrastim, pegfilgrastim, or granulocyte macrophage colony stimulating factor.

[0021] In some embodiments, the method further comprises measuring cytokine and chemokine levels, hi some embodiments, measuring the levels of at least one of IL-6, IL-8, IL-10, IL-15, TNF-α, IFN-γ, and sIL2Rα.

[0022] In one aspect, the disclosure provides a container comprising a suspension of CD19-directed genetically modified autologous T cells, about 5% dimethylsulfoxide (DMSO), and about 2.5% by volume of human albumin. In another aspect, the container comprises about 0.4×10 8 ~2×10 8 The container comprises a suspension of CD19-directed genetically modified autologous T cells (CAR-positive viable T cells). In some embodiments, the container is a sterile infusion bag. In some embodiments, the volume of the infusion bag is about 100 mL, 250 mL, 500 mL, 750 mL, 1000 mL, 1500 mL, 2000 mL, or 3000 mL.

[0023] In one aspect, the disclosure provides a method of treating relapsed or refractory large B-cell lymphoma in a human after two or more lines of systemic therapy, comprising administering to a human in need of treatment a CD19-directed genetically modified autologous T-cell immunotherapy comprising: (a) administering to the human a CD19-directed chimeric antibody; administering to the patient a composition comprising viable CAR-positive T cells; (b) monitoring the patient for signs and symptoms of adverse reactions following administration; (c) if greater than Grade 2 cytokine release syndrome (CRS) is observed in (b), administer tocilizumab IV at a dose of about 8 mg / kg over 1 hour and repeat tocilizumab every 8 hours as needed if unresponsive to increased IV fluids or supplemental oxygen; (d) if CRS symptoms observed in (b) do not improve 24 hours after (c), administer methylprednisolone IV at about 1 mg / kg twice daily or an equivalent dose of dexamethasone and continue corticosteroids until events are Grade 1 or less, followed by tapering over 3 days; (e) if Grade 3 CRS is observed in (b), administer tocilizumab IV at a dose of about 8 mg / kg over 1 hour and repeat and (f) if CRS Grade 4 is observed in (b), administering tocilizumab at a dose of about 8 mg / kg IV over 1 hour and, if unresponsive to increasing IV fluids or supplemental oxygen, repeating tocilizumab every 8 hours as needed, methylprednisolone 1 mg / kg IV twice daily or an equivalent dexamethasone dose, continuing corticosteroids until events are Grade 1 or less, then tapering over 3 days; and (f) if CRS Grade 4 is observed in (b), administering tocilizumab at a dose of about 8 mg / kg IV over 1 hour and, if unresponsive to increasing IV fluids or supplemental oxygen, repeating tocilizumab every 8 hours as needed, and methylprednisolone at about 1,000 mg IV per day for 3 days.

[0024] In one aspect, the disclosure provides a method of treating relapsed or refractory large B-cell lymphoma in a patient after two or more lines of systemic therapy, comprising administering a CD19-directed genetically modified autologous T-cell immunotherapy to a patient in need of treatment, comprising: (a) administering to the patient a composition comprising CD19-directed chimeric antigen receptor (CAR)-positive viable T cells; (b) monitoring the patient for signs and symptoms of adverse reactions following administration; and (c) managing cytokine release syndrome (CRS) and / or neurotoxicity according to Table 1 and / or Table 2 if CRS and / or neurotoxicity is observed.

[0025] The disclosed methods are further illustrated by the following numbered non-limiting embodiments.

[0026] 1. A method of treating relapsed or refractory diffuse large B-cell lymphoma (DLBCL) not otherwise specified, primary mediastinal large B-cell lymphoma, high-grade B-cell lymphoma, or DLBCL arising from follicular lymphoma in a patient after two or more lines of systemic therapy, comprising: For patients who need treatment, about 1 × 10 6 ~Approx. 2×10 6 Dose of CAR-positive viable T cells / kg body weight, up to approximately 1 × 10 8 administering an axicabtagene ciloleucel suspension by intravenous infusion at a dose of CAR-positive viable T cells; A method in which axicabtagene ciloleucel is a CD19-directed genetically modified autologous T cell immunotherapy comprising a patient's own T cells harvested and genetically modified ex vivo by retroviral transduction to express a chimeric antigen receptor (CAR) comprising an anti-CD19 single chain variable fragment (scFv) linked to the costimulatory domains of CD28 and CD3 zeta.

[0027] 2. The intravenous infusion time is 15 to 120 minutes, or a maximum of 30 minutes; The injection volume is between 50 and 100 mL, or about 68 mL; and / or The immunotherapy is infused from an infusion bag, optionally with the infusion bag being agitated during infusion, and optionally with 2. The method of embodiment 1, wherein the immunotherapy is administered within 3 hours after thawing.

[0028] 3. The suspension is

[0029] 3. The method of embodiment 1 or 2, further comprising albumin, optionally wherein the albumin is present in an amount of about 2-3% by volume, or about 2.5% by volume, optionally wherein the albumin is human albumin, and / or further comprising DMSO.

[0030] 4. A method of treating relapsed or refractory large B-cell lymphoma in a patient after two or more lines of systemic therapy, comprising: (a) administering to a patient in need of treatment a CD19-directed genetically modified autologous T cell immunotherapy, preferably axicabtagene ciloleucel; and (b) monitoring the patient after the infusion for adverse reactions and / or safety signs and symptoms.

[0031] 5. The method of embodiment 4, wherein the relapsed or refractory large B-cell lymphoma is diffuse large B-cell lymphoma (DLBCL) not otherwise specified, primary mediastinal large B-cell lymphoma, high-grade B-cell lymphoma, or DLBCL arising from follicular lymphoma.

[0032] 6. The method of embodiment 4 or 5, wherein the adverse reaction is selected from the group consisting of cytokine release syndrome (CRS), neurotoxicity, hypersensitivity reactions, serious infections, cytopenias, and hypogammaglobulinemia.

[0033] 7. The method of embodiment 6, wherein the signs and symptoms of the adverse reaction are selected from the group consisting of fever, hypotension, tachycardia, hypoxia, and chills, and include cardiac arrhythmias (including atrial fibrillation and ventricular tachycardia), cardiac arrest, cardiac failure, renal insufficiency, capillary leak syndrome, hypotension, hypoxia, organ toxicity, hemophagocytic lymphohistiocytosis / macrophage activation syndrome (HLH / MAS), seizures, encephalopathy, headache, tremor, dizziness, aphasia, delirium, insomnia anxiety, anaphylaxis, febrile neutropenia, thrombocytopenia, neutropenia, and anemia.

[0034] 8. The method of any one of embodiments 1 to 7, further comprising administering an effective amount of an inhibitor or combination of inhibitors of GM-CSF, CSF1, GM-CSFR or CSF1R to treat symptoms of the adverse reaction.

[0035] 9. (i) the GM-CSF inhibitor is selected from lenzilumab, namilumab (AMG203), GSK3196165 / MOR103 / otilimab (GSK / MorphoSys), KB002 and KB003 (KaloBios), MT203 (Micromet and Nycomed), MORAb-022 / dimcirumab (Morphotek), or a biosimilar of any one of them, E21R, and a small molecule; (ii) the CSF1 inhibitor is selected from RG7155, PD-0360324, MCS110 / Lanotuzumab), or a biosimilar version of any one of them, and a small molecule; and / or (iii) GM-CSFR and CSF1R inhibitors include mavrilimumab (formerly CAM-3001; MedImmune, Inc.); cabilalizumab (Five Prime Therapeutics); LY3022855 (IMC-CS4) (Eli Lilly), Emactuzumab, also known as RG7155 or RO5509554; FPA008 (Five Prime / BMS); AMG820 (Amgen); ARRY-382 (Array Biopharma); MCS110 (Novartis); PLX3397 (Plexxikon); ELB041 / AFS98 / TG3003 (ElsaLys Bio, Transgene), SNDX-6352 (Syn dax); a biosimilar version of any one of them, and a small molecule.

[0036] 10. The method of embodiment 8 or 9, wherein the inhibitor or combination of inhibitors is selected from lenzilumab and mavrilimumab, and optionally either lenzilumab or mavrilimumab is administered by IP infusion at 10 mg / kg (maximum 600 mg), 20 mg / kg (maximum 1200 mg), or 30 mg / kg (maximum 1800 mg), respectively, or administered subcutaneously at 100 mg or 150 mg.

[0037] 11. The method of embodiment 8 or 9, wherein at least one of an inhibitor of GM-CSF, CSF1, GM-CSFR, or CSF1R, or a combination of inhibitors, is administered on the same day as axicabtagene ciloleucel.

[0038] 12. The method of any one of embodiments 8 to 11, wherein at least one or a combination of inhibitors of GM-CSF, CSF1, GM-CSFR, or CSF1R is administered 1 to 12 hours prior to, and optionally simultaneously with, administration of axicabtagene ciloleucel.

[0039] 13. The method of any one of embodiments 8-12, wherein at least one or a combination of inhibitors of GM-CSF, CSF1, GM-CSFR, or CSF1R is administered two or more times following administration of axicabtagene ciloleucel.

[0040] 14. Axicabtagene siloleucel is 2×10 6 CAR-positive viable T cells / kg (maximum tolerated dose: 2 × 10 8 The method of any one of embodiments 8-13, wherein the IL-10 antibody is administered as a single IV infusion at a target dose of 10 mg / kg (maximum 600 mg), 20 mg / kg (maximum 1200 mg), or 30 mg / kg (maximum 1800 mg), respectively, and either lenzilumab or mavrilimumab is administered by IP infusion at 10 mg / kg (maximum 600 mg), 20 mg / kg (maximum 1200 mg), or 30 mg / kg (maximum 1800 mg), respectively, or is administered subcutaneously at 100 mg or 150 mg.

[0041] 15. The method of any one of embodiments 8-14, further comprising administering a steroid (e.g., a corticosteroid) to treat symptoms of the adverse reaction.

[0042] 16. The method of any one of embodiments 4 to 15, wherein the adverse reaction is cytokine release syndrome (CRS).

[0043] 17. The method of any one of embodiments 4 to 16, comprising monitoring for signs and symptoms of cytokine release syndrome (CRS) for about 7 days after infusion, optionally at least daily.

[0044] 18. The method of any one of embodiments 4 to 17, wherein the adverse reaction is (1) neurotoxicity, optionally wherein symptoms of neurotoxicity are encephalopathy, headache, tremors, dizziness, aphasia, delirium, insomnia, and / or anxiety, and / or (2) cytopenia.

[0045] 19. The method of any one of embodiments 1-18, further comprising administering a non-sedating anti-seizure drug for seizure prevention, administering at least one of erythropoietin, darbepoetin alfa, platelet transfusions, filgrastim, or pegfilgrastim, and / or administering tocilizumab.

[0046] 20. The method further comprises measuring levels of cytokines and chemokines before and / or after one or more administrations, optionally including measuring levels of IL-2, IL-6, IL-8, IL-10, IL-11, IL-12, IL-13, IL-14, IL-15, IL-16, IL-17, IL-19, IL-20, IL-21, IL-22, IL-23, 20. The method according to any one of the preceding embodiments, wherein the level of at least one of IL-12p40 / p70, IL-15, IL-17a, TNF-α, IFN-γ, GM-CSF and sIL2Rα is measured.

[0047] 21. The method of any one of embodiments 1-20, further comprising administering an effective amount of a corticosteroid and / or tocilizumab to treat symptoms of the adverse reaction.

[0048] 22. Axicabtagene ciloleucel was administered at 2 × 10 6 Target dose of CAR-positive viable T cells / kg (maximum tolerated dose: 2 × 10 8 14. The method of any one of embodiments 8-13, wherein the IL-16 antibody is administered as a single IV infusion at 3 mg / kg / mL, and / or mavrilimumab is administered once subcutaneously, OP / IP, or IV at 3 mg / kg on day 0.

[0049] 23. Axicabtagene ciloleucel was administered at a dose of 2 × 10 6 Target dose of CAR-positive viable T cells / kg (maximum tolerated dose: 2 × 10 8 14. The method of any one of embodiments 8-13, wherein the mavrilimumab is administered as a single IV infusion at 1 mg / kg, 2 mg / kg, 3 mg / kg, 4 mg / kg, 5 mg / kg, 6 mg / kg, 7 mg / kg, 8 mg / kg, 9 mg / kg, 10 mg / kg, 11 mg / kg, 12 mg / kg, 13 mg / kg, 14 mg / kg, 15 mg / kg, 16 mg / kg, 17 mg / kg, 18 mg / kg, 19 mg / kg, 20 mg / kg, 21 mg / kg, 22 mg / kg, 23 mg / kg, 24 mg / kg, 25 mg / kg, 26 mg / kg, 27 mg / kg, 28 mg / kg, 29 mg / kg, or 30 mg / kg once subcutaneously, OP / IP, or IV on day 0.

[0050] 24. 30mg / m 2 / day fludarabine and 500 mg / m 2 24. The method of embodiment 22 or 23, further comprising administering a conditioning chemotherapy comprising cyclophosphamide / day from day -5 to day -3.

[0051] 25. A method of treating relapsed or refractory large B-cell lymphoma in a human after two or more lines of systemic therapy, comprising administering to a human in need of treatment a CD19-directed genetically modified autologous T-cell immunotherapy,

[0052] (a) administering to a patient a composition comprising CD19-directed chimeric antigen receptor (CAR)-positive viable T cells;

[0053] (b) monitoring patients after administration for signs and symptoms of adverse reactions; and

[0054] (c) If greater than Grade 2 cytokine release syndrome (CRS) is observed in (b), administer tocilizumab IV at a dose of approximately 8 mg / kg over 1 hour, repeating tocilizumab every 8 hours as needed if not responsive to increased IV fluids or supplemental oxygen;

[0055] (d) if the CRS symptoms observed in (b) do not improve 24 hours after (c), administer approximately 1 mg / kg IV methylprednisolone twice daily or an equivalent dose of dexamethasone and continue corticosteroids until events are Grade 1 or less, then taper over 3 days;

[0056] (e) If Grade 3 CRS is observed in (b), administer tocilizumab IV at a dose of 8 mg / kg over 1 hour, and if unresponsive to increased IV fluids or supplemental oxygen, repeat tocilizumab every 8 hours as needed and administer methylprednisolone 1 mg / kg twice daily or an equivalent dexamethasone dose until events become Grade 1 or less. Continue corticosteroids until relief, then taper over 3 days; and

[0057] (f) if Grade 4 CRS is observed in (b), administering tocilizumab at a dose of about 8 mg / kg IV over 1 hour, repeating tocilizumab every 8 hours as needed if unresponsive to IV fluids or increased supplemental oxygen, and administering methylprednisolone IV at about 1,000 mg per day for 3 days.

[0058] 26. A method of treating relapsed or refractory large B-cell lymphoma in a patient after two or more lines of systemic therapy, comprising administering to a patient in need of treatment a CD19-directed genetically modified autologous T-cell immunotherapy,

[0059] (a) administering to a patient a composition comprising CD19-directed chimeric antigen receptor (CAR)-positive viable T cells;

[0060] (b) monitoring patients after administration for signs and symptoms of adverse reactions; and

[0061] (c) administering a CD19-directed genetically modified autologous T cell immunotherapy, comprising, if cytokine release syndrome (CRS) and / or neurotoxicity is observed, managing cytokine release syndrome (CRS) and / or neurotoxicity according to Table 1, Table 2, Example 2 and / or Example 3.

[0062] 27. A method of treating relapsed or refractory large cell ALL in a patient, comprising administering to a patient in need of treatment a CD19-directed genetically modified autologous T cell immunotherapy,

[0063] (a) administering to a patient a composition comprising CD19-directed chimeric antigen receptor (CAR)-positive viable T cells;

[0064] (b) monitoring patients after administration for signs and symptoms of adverse reactions; and

[0065] (c) administering a CD19-directed genetically modified autologous T cell immunotherapy, comprising, if cytokine release syndrome (CRS) and / or neurotoxicity is observed, managing cytokine release syndrome (CRS) and / or neurotoxicity according to Table 1, Table 2, Example 2 and / or Example 3.

[0066] 28. A method of treating relapsed or refractory diffuse large B-cell lymphoma (DLBCL) not otherwise specified, primary mediastinal large B-cell lymphoma, high-grade B-cell lymphoma, or DLBCL arising from follicular lymphoma in a patient in need of treatment, comprising:

[0067] For patients who need treatment, about 1 × 10 6 ~Approx. 2×10 6 Dose of CAR-positive viable T cells / kg body weight, up to approximately 1 × 10 8 administering, by intravenous infusion, axicabtagene ciloleucel suspension in combination with rituximab at a dose of CAR-positive viable T cells;

[0068] A method in which axicabtagene ciloleucel is a CD19-directed genetically modified autologous T cell immunotherapy comprising a patient's own T cells harvested and genetically modified ex vivo by retroviral transduction to express a chimeric antigen receptor (CAR) comprising an anti-CD19 single chain variable fragment (scFv) linked to the costimulatory domains of CD28 and CD3 zeta.

[0069] 29. A patient receives rituximab 375 mg / m on day -5. 2 and fludarabine 30 mg / m on days -5, -4, and -3. 2 and cyclophosphamide 500 mg / m 2 Patients received conditioning chemotherapy with 2 × 10 6The method of embodiment 28, wherein axicabtagene ciloleucel is administered at a target dose of anti-CD19 CAR T cells / kg.

[0070] 30. A patient receives lenalidomide 10 mg daily starting 7 days after leukapheresis and continuing until day 3 after axicabtagene ciloleucel infusion, followed by fludarabine 30 mg / m on days -5, -4, and -3. 2 and cyclophosphamide 500 mg / m 2 Patients received conditioning chemotherapy with 2 × 10 6 The method of embodiment 28, wherein axicabtagene ciloleucel is administered at a target dose of anti-CD19 CAR T cells / kg.

[0071] 31. The method of embodiment 28, wherein the patient is refractory to first-line therapy, refractory to second-line or subsequent therapy, and refractory after autologous stem cell transplantation.

[0072] 32. The method of any one of embodiments 28-31, wherein the patient has previously received an anti-CD20 mAb and an anthracycline-containing chemotherapy regimen.

[0073] 33. The method of any one of embodiments 28-32, wherein the patient further receives five additional doses of rituximab at 28 day intervals following the axicabtagene ciloleucel injection.

[0074] 34. The method of any one of embodiments 28-33, wherein the patient further receives lenalidomide 20 mg at 28 day intervals (21 days / 28 days on treatment) for five additional cycles after axicabtagene ciloleucel injection.

[0075] 35. The method of any one of embodiments 28-34, wherein the patient is further administered mesna.

[0076] 36. Axicabtagene ciloleucel was administered at a dose of 2 × 10 6Target dose of CAR-positive viable T cells / kg (maximum tolerated dose: 2 × 10 8 The method of any one of embodiments 8-13, wherein the IL-16 antibody is administered as a single IV infusion at 3 mg / kg body weight on day 0, and mavrilimumab is administered once subcutaneously, OP / IP, or IV at 3 mg / kg body weight on day 0.

[0077] 37. Axicabtagene ciloleucel was administered at 2×10 6 CAR-positive viable T cells / kg (maximum tolerated dose: 2 × 10 8 and / or mavrilimumab is administered as a single IV infusion at a target dose of 1 mg / kg body weight, 2 mg / kg body weight, 3 mg / kg body weight, 4 mg / kg body weight, 5 mg / kg body weight, 6 mg / kg body weight, 7 mg / kg body weight, 8 mg / kg body weight, 9 mg / kg body weight, 10 mg / kg body weight, 11 mg / kg body weight, 12 mg / kg body weight, 13 mg / kg body weight, 14 mg / kg body weight, 15 mg / kg body weight, 16 mg / kg body weight on day 0. 14. The method of any one of embodiments 8-13, wherein the compound is administered once subcutaneously, OP / IP, or IV at 17 mg / kg body weight, 18 mg / kg body weight, 19 mg / kg body weight, 20 mg / kg body weight, 21 mg / kg body weight, 22 mg / kg body weight, 23 mg / kg body weight, 24 mg / kg body weight, 25 mg / kg body weight, 26 mg / kg body weight, 27 mg / kg body weight, 28 mg / kg body weight, 29 mg / kg body weight, or 30 mg / kg.

[0078] 38. A method for the treatment of a patient with advanced disease, further comprising administering conditioning chemotherapy comprising 30 mg / m2 / day fludarabine and 500 mg / m2 / day cyclophosphamide on days -5 to -3. 37. The method of claim 36.

[0079] In one aspect, the disclosure provides a method of treating relapsed or refractory large B-cell lymphoma after two or more lines of systemic therapy in a human, comprising administering a CD19-directed genetically modified autologous T-cell immunotherapy to a human in need of treatment, comprising: (a) administering to the patient a composition comprising viable CD19-directed chimeric antigen receptor (CAR)-positive T cells; (b) monitoring the patient following administration for signs and symptoms of adverse reactions; (c) if cytokine release syndrome (CRS) greater than Grade 2 is observed in (b), administering tocilizumab IV at a dose of about 8 mg / kg over 1 hour, repeating tocilizumab every 8 hours as needed if unresponsive to increased IV fluids or supplemental oxygen; (d) if CRS symptoms observed in (b) do not improve 24 hours after (c), administering methylprednisolone IV at about 1 mg / kg twice daily, or an equivalent dose of dexamethasone, and administering corticosteroids until the event is Grade 1 or less. (e) if CRS Grade 3 is observed in (b), administer tocilizumab at a dose of 8 mg / kg IV over 1 hour and if unresponsive to increasing IV fluids or supplemental oxygen, repeat tocilizumab every 8 hours as needed, administer methylprednisolone 1 mg / kg IV twice daily or an equivalent dexamethasone dose, continue corticosteroids until the event is Grade 1 or less, then taper over 3 days; and (f) if CRS Grade 4 is observed in (b), administer tocilizumab at a dose of about 8 mg / kg IV over 1 hour and if unresponsive to increasing IV fluids or supplemental oxygen, repeat tocilizumab every 8 hours as needed, and administer methylprednisolone at about 1,000 mg IV per day for 3 days.

[0080] In one aspect, the disclosure relates to a method of treating relapsed or refractory large B-cell lymphoma in a patient after two or more lines of systemic therapy, comprising administering a CD19-directed genetically modified autologous T-cell immunotherapy to a patient in need of treatment, comprising: (a) administering to the patient a composition comprising CD19-directed chimeric antigen receptor (CAR)-positive viable T cells; (b) monitoring the patient for signs and symptoms of adverse reactions following administration; and (c) managing cytokine release syndrome (CRS) and / or neurotoxicity according to Table 1, Table 2, Table 3, and / or Table 4 if CRS and / or neurotoxicity is observed.

[0081] In one aspect, the disclosure relates to a method of treating relapsed or refractory large cell ALL in a patient, comprising administering a CD19-directed genetically modified autologous T-cell immunotherapy to a patient in need of treatment, comprising: (a) administering to the patient a composition comprising CD19-directed chimeric antigen receptor (CAR)-positive viable T cells; (b) monitoring the patient for signs and symptoms of adverse reactions following administration; and (c) managing cytokine release syndrome (CRS) and / or neurotoxicity according to Table 1, Table 2, Example 2, and / or Example 3 if CRS and / or neurotoxicity is observed.

[0082] Other features and advantages of the present disclosure will become apparent from the following detailed description, including the examples. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0083] definition In order that this disclosure may be more readily understood, certain terms are first defined below. Additional definitions for the following terms, as well as other terms, are found throughout the specification.

[0084] As used in this specification and the appended claims, the singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise.

[0085] As used herein, unless otherwise stated or clear from the context, the term "or" is understood to be inclusive and includes both "or" and "and."

[0086] The term "and / or" as used herein should be interpreted as a specific disclosure of each of the two specified features or components, whether or not the other is present. Thus, the term "and / or" as used herein, for example, in the phrase "A and / or B," is intended to include A and B, A or B, A (alone), and B (alone). Similarly, the term "and / or" as used, for example, in the phrase "A, B, and / or C," is intended to encompass each of the following embodiments: A, B, and C; A, B, or C; A or C; A or B; B or C; A and C; A and B; B and C; A (alone); B (alone); and C (alone).

[0087] As used herein, the terms "for example" and "i.e." are used merely as examples and are not intended to be limiting, and should not be construed as referring only to the items explicitly listed herein.

[0088] Terms such as "more than," "at least," and "more than," for example, "at least one," are intended to mean, but are not limited to, at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 1 06, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 300, 301, 302, 303, 304, 305, 306, 307, 308, 309, 310, 311, 312, 313, 314, 315, 316, 317, 318, 319, 320, 321, 322, 323, 324, 325, 326, 327, 328, 329, 330, 331, 332, 333, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 344, 345, 346, 347, 348, 349, 350, 351, 352, 353, 354, 355, 356, 357, 3 This is understood to include 9, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, or 150, 200, 300, 400, 500, 600, 700, 800, 900, 1000, 2000, 3000, 4000, 5000, or any value greater than the recited value, as well as any greater number or fraction in between.

[0089] Conversely, the term "less than" includes each value less than the recited value. For example, "100 or fewer nucleotides" includes 100, 99, 98, 97, 96, 95, 94, 93, 92, 91, 90, 89, 88, 87, 86, 85, 84, 83, 82, 81, 80, 79, 78, 77, 76, 75, 74, 73, 72, 71, 70, 69, 68, 67, 66, 65, 64, 63, 62, 61, 60, 59, 58, 57, 56, 55, 54, 55, 56, 57, 58, 59 ... Included are 3, 52, 51, 50, 49, 48, 47, 46, 45, 44, 43, 42, 41, 40, 39, 38, 37, 36, 35, 34, 33, 32, 31, 30, 29, 28, 27, 26, 25, 24, 23, 22, 21, 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2, 1, and 0 nucleotides. Any smaller number or fraction in between is also included.

[0090] Terms such as "plurality," "at least two," "two or more," and "at least a second" are used without limitation. 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63 , 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, 153, 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, 170, 171, 172, 5, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 1 This is understood to include 38, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, or 150, 200, 300, 400, 500, 600, 700, 800, 900, 1000, 2000, 3000, 4000, 5000, or more, as well as any higher numbers or fractions therebetween.

[0091] Throughout this specification the words "comprising" or "comprises" or variations such as "comprising" are understood to mean the inclusion of a stated element, integer or step, or group of elements, integers or steps, but not the exclusion of any other element, integer or step, or group of elements, integers or steps. Whenever an embodiment is described herein with the language "comprising," it is understood that other similar embodiments described with the terms "consisting of" and / or "consisting essentially of" are also presented.

[0092] As used herein, unless specifically stated or clear from the context, the term "about" refers to a value or composition that is within an acceptable error range for a particular value or composition as determined by one of ordinary skill in the art, which depends in part on how the value or composition is measured or determined, i.e., on the limitations of the measurement system. For example, "about" or "approximately" can mean within one or more standard deviations as per the practice in the art. "About" or "approximately" can mean a range of up to 10% (i.e., ±10%). Thus, "about" can be understood to be 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, 0.5%, 0.1%, 0.05%, 0.01%, or 0.001% greater or less than the stated value. For example, about 5 mg can include any amount between 4.5 mg and 5.5 mg. Additionally, particularly with respect to biological systems or processes, the term can mean up to an order of magnitude or up to 5-fold greater. When a particular value or composition is presented in this disclosure, unless otherwise specified, the meaning of "about" or "approximately" should be assumed to be within an acceptable error range for that particular value or composition.

[0093] As described herein, any concentration range, percentage range, ratio range, or integer range should be understood to include any integer value within the recited range, and fractions thereof, as appropriate (such as 1 / 10 and 1 / 100 of an integer), unless otherwise stated.

[0094] Units, prefixes and symbols used herein are presented using the format accepted by the Systeme International de Unites (SI). Numerical ranges are inclusive of the numbers defining the range.

[0095] Unless otherwise defined, all technical and scientific terms used herein are within the meaning of the present disclosure. have the same meaning as commonly understood by those skilled in the relevant art. See, e.g., Juo, "The Concise Dictionary of Biomedicine and Molecular Biology", 2nd Edition, (2001), CRC Press; "The Dictionary of Cell & Molecular Biology", 5th Edition, (2013), Academic Press; and "The Oxford Dictionary Of Biochemistry And Molecular Biology", Cammack et al., eds., 2nd Edition, (2006), Oxford The University Press provides one of skill in the art with a general dictionary for many of the terms used in this disclosure.

[0096] "Administering" refers to the physical introduction of an agent into a subject using any of a variety of methods and delivery systems known to those skilled in the art. Exemplary routes of administration for the formulations disclosed herein include, for example, intravenous, intramuscular, subcutaneous (SQ), intraperitoneal, spinal or other parenteral routes of administration, such as by injection or infusion. As used herein, the phrase "parenteral administration" refers to a mode of administration other than enteral and topical administration, usually by injection, including, but not limited to, intravenous (IV), intramuscular, intraarterial, intrathecal, intralymphatic, intralesional, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal (IP), transtracheal, subcutaneous, subcuticular, intraarticular, subcapsular, subarachnoid, intraspinal, epidural and substernal injection and infusion, and in vivo electroporation. In some embodiments, the formulation is administered via a parenteral route, for example, orally. Other parenteral routes include topical, epidermal or mucosal routes of administration, such as intranasal, intravaginal, rectal, sublingual, or topical. Administration can also be, for example, once, multiple times, and / or over one or more extended periods of time. When more than one therapeutic agent is administered, administration can be simultaneous or sequential. Sequential administration involves administering one agent only after administration of one or more other agents is completed.

[0097] The term "antibody" (Ab) includes, but is not limited to, a glycoprotein immunoglobulin that specifically binds to an antigen. In general, an antibody may include at least two heavy (H) chains and two light (L) chains, or antigen-binding molecules thereof, interconnected by disulfide bonds. Each H chain includes a heavy chain variable region (abbreviated herein as VH) and a heavy chain constant region. The heavy chain constant region includes three constant domains, CH1, CH2, and CH3. Each light chain includes a light chain variable region (abbreviated herein as VL) and a light chain constant region. The light chain constant region includes one constant domain, CL. The VH and VL regions can be further subdivided into hypervariable regions, called complementarity determining regions (CDRs), which are interspersed with more conserved regions, called framework regions (FRs). Each VH and VL includes three CDRs and four FRs, arranged from amino-terminus to carboxy-terminus in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, and FR4. The variable regions of the heavy and light chains contain a binding domain that interacts with an antigen, and the constant regions of the Abs may mediate the binding of the immunoglobulin to host tissues or factors, such as various cells of the immune system (e.g., effector cells) and the first component (C1q) of the classical complement system.

[0098] Antibodies include, for example, monoclonal antibodies, recombinantly produced antibodies, monospecific antibodies, multispecific antibodies (including bispecific antibodies), human antibodies, engineered antibodies, humanized antibodies, chimeric antibodies, immunoglobulins, synthetic antibodies, tetrameric antibodies comprising two heavy chain molecules and two light chain molecules, antibody light chain monomers, antibody heavy chain monomers, antibody light chain dimers, antibody heavy chain dimers, antibody light chain-antibody heavy chain pairs, intrabodies, antibody fusions (sometimes referred to herein as "antibody conjugates"), heteroconjugate antibodies, single domain antibodies, monovalent antibodies, single chain antibodies or single chain Fvs (scFvs), camelized antibodies, affibodies, Fab fragments, F(ab')2 fragments, disulfide-linked Fvs (sdFvs), bispecific (ScFvs), 2 - Fab, anti-idiotypic (anti-Id) antibodies (including, for example, anti-anti-Id antibodies), minibodies, domain antibodies, synthetic antibodies (also referred to herein as "antibody mimetics"); In some embodiments, antibodies described herein include polyclonal antibody populations, bispecific antibodies, IgG fusions (e.g., dual variable domain (DVD)-Ig), Fc fusions (e.g., ScFv / Fc fusions), immunomobilizing mTCRs, and antigen-binding fragments of any of the above. In some embodiments, antibodies described herein refer to polyclonal antibody populations.

[0099] An "antigen-binding molecule," "antigen-binding portion," or "antibody fragment" refers to any molecule that contains an antigen-binding portion (e.g., CDR) of the antibody from which the molecule is derived. An antigen-binding molecule may contain an antigen complementarity determining region (CDR). Examples of antibody fragments include, but are not limited to, Fab, Fab', F(ab')2, and Fv fragments, dAbs, linear antibodies, scFv antibodies, and multispecific antibodies formed from antigen-binding molecules. Peptibodies (i.e., Fc fusion molecules containing a peptide binding domain) are another example of a suitable antigen-binding molecule. In some embodiments, the antigen-binding molecule binds to an antigen on a tumor cell. In some embodiments, the antigen-binding molecule binds to an antigen on a cell involved in a hyperproliferative disease or a viral or bacterial antigen. In some embodiments, the antigen-binding molecule binds to CD19. In further embodiments, the antigen-binding molecule is an antibody fragment that specifically binds to an antigen, including one or more of its complementarity determining regions (CDRs). In further embodiments, the antigen-binding molecule is a single-chain variable fragment (scFv). In some embodiments, the antigen-binding molecule comprises or consists of an avimer.

[0100] "Antigen" refers to any molecule that can elicit an immune response or be bound by an antibody or antigen-binding molecule. The immune response can include either antibody production or activation of specific immunocompetent cells, or both. Those skilled in the art will readily appreciate that any macromolecule, including virtually any protein or peptide, can function as an antigen. Antigens can be expressed endogenously, i.e., by genomic DNA, or recombinantly. Antigens can be specific to a particular tissue, such as cancer cells, or can be broadly expressed. Additionally, fragments of larger molecules can act as antigens. In some embodiments, the antigen is a tumor antigen.

[0101] "CD19-directed genetically modified autologous T cell immunotherapy" refers to a suspension of chimeric antigen receptor (CAR)-positive T cells. An example of such an immunotherapy is axicabtagene ciloleucel (also known as Axi-cel™, YESCARTA®) developed by Kite Pharmaceuticals, Inc. Other non-limiting examples include JCAR017, JCAR015, JCAR014, Kymriah (tisagenlecleucel), Uppsala U. anti-CD19 CAR (NCT02132624), and UCART19 (Celectis).

[0102] The term "neutralizing" refers to an antigen-binding molecule, scFv, antibody, or fragment thereof that binds to a ligand and prevents or reduces the biological action of that ligand. In some embodiments, the antigen-binding molecule, scFv, antibody, or fragment thereof directly blocks a binding site on the ligand or otherwise alters the ability of the ligand to bind through indirect means (such as a structural or energetic change in the ligand). In some embodiments, the antigen-binding molecule, scFv, antibody, or fragment thereof prevents the protein to which it is bound from performing a biological function.

[0103] The term "autologous" refers to any material derived from the same individual that is later reintroduced. For example, the engineered autologous cell therapy (eACT™) method described herein involves the collection of lymphocytes from a patient, which are then engineered to express, for example, a CAR construct, and then administered to the same patient.

[0104] The term "allogeneic" refers to a gene that originates from one individual and is then introduced into another individual of the same species. The term refers to any material that is capable of regenerating cells, such as allogeneic T cell transplantation.

[0105] The terms "transduction" and "transduced" refer to the process by which foreign DNA is introduced into a cell via a viral vector (see Jones et al., Genetics: principles and analysis, Boston: Jones & Bartlett Publ. (1998)). In some embodiments, the vector is a retroviral vector, a DNA vector, an RNA vector, an adenoviral vector, a baculoviral vector, an Epstein-Barr virus vector, a papovavirus vector, a vaccinia virus vector, a herpes simplex virus vector, an adenovirus-associated vector, a lentiviral vector, or any combination thereof.

[0106] "Cancer" refers to a broad group of various diseases characterized by the uncontrolled growth of abnormal cells in the body. Unregulated cell division and growth leads to the formation of malignant tumors that may invade adjacent tissues and metastasize to distant parts of the body via the lymphatic system or bloodstream. "Cancer" or "cancerous tissue" may include tumors. Examples of cancers that may be treated by the methods disclosed herein include, but are not limited to, cancers of the immune system, including lymphomas, leukemias, myelomas, and other white blood cell malignancies. In some embodiments, the methods disclosed herein are directed to treating a variety of cancers, including, for example, bone cancer, pancreatic cancer, skin cancer, head and neck cancer, cutaneous or intraocular malignant melanoma, uterine cancer, ovarian cancer, rectal cancer, cancer of the anal region, stomach cancer, testicular cancer, uterine cancer, cancer of the fallopian tubes, endometrial cancer, cervical cancer, vaginal cancer, vulvar cancer, multiple myeloma, Hodgkin's disease, non-Hodgkin's lymphoma (NHL), primary mediastinal large B-cell lymphoma (PMBC), diffuse large B-cell lymphoma (DLBCL), follicular lymphoma (FL), transformed follicular lymphoma, splenic marginal zone lymphoma (SMZL), esophageal cancer, small intestine cancer, cancer of the endocrine system, thyroid cancer, cancer of the parathyroid gland, adrenal gland cancer, soft tissue cancer, and the like. It may be used to reduce tumor size in tumors derived from sarcoma, urethral cancer, penile cancer, chronic or acute leukemia, acute myeloid leukemia, chronic myelogenous leukemia, acute lymphoblastic leukemia (ALL) (including non-T-cell ALL), chronic lymphocytic leukemia (CLL), childhood solid tumors, lymphocytic lymphoma, bladder cancer, kidney or ureter cancer, renal pelvis cancer, central nervous system (CNS) neoplasms, primary CNS lymphoma, tumor angiogenesis, spinal axis tumor, brain stem glioma, pituitary adenoma, Kaposi's sarcoma, epidermoid carcinoma, squamous cell carcinoma, T-cell lymphoma, environmentally induced cancers including those induced by asbestos, other B-cell malignancies, and combinations of the above cancers. In some embodiments, the cancer is multiple myeloma. Certain cancers may be responsive to chemotherapy or radiation therapy, or certain cancers may be refractory. Refractory cancer refers to cancer that is not amenable to surgical intervention; refractory cancers either do not respond to chemotherapy or radiation therapy in the first place or the cancer becomes unresponsive over time.

[0107] As used herein, "anti-tumor effect" refers to a biological effect that can be presented as a reduction in tumor volume, a reduction in the number of tumor cells, a reduction in tumor cell proliferation, a reduction in the number of metastases, an increase in overall survival or progression-free survival, an increase in life expectancy, or an improvement in various physiological symptoms associated with tumors. Anti-tumor effect can also refer to the prevention of tumor development, e.g., a vaccine.

[0108] As used herein, "cytokine" refers to a non-antibody protein released by one cell in response to contact with a specific antigen, where the cytokine interacts with a second cell to mediate a response in the second cell. As used herein, "cytokine" is meant to refer to a protein released by one cell population that acts on another cell as an intercellular mediator. Cytokines can be expressed endogenously by cells or can be administered to a subject. Cytokines can be released by immune cells, such as macrophages, B cells, T cells, and mast cells, to propagate an immune response. Cytokines can induce a variety of responses in recipient cells. Cytokines include homeostatic cytokines, chemokines, proinflammatory cytokines, effectors, and acute phase proteins. For example, homeostatic cytokines such as interleukin (IL) 7 and IL-15 can promote survival and proliferation of immune cells, and proinflammatory cytokines can promote inflammatory responses. Examples of homeostatic cytokines include, but are not limited to, IL-2, IL-4, IL-5, IL-7, IL-10, IL-12p40, IL-12p70, IL-15, and interferon (IFN) gamma. Examples of proinflammatory cytokines include, but are not limited to, IL-1a, IL-1b, IL-6, IL-13, IL-17a, tumor necrosis factor (TNF)-alpha, TNF-beta, fibroblast growth factor (FGF)2, granulocyte-macrophage colony-stimulating factor (GM-CSF), soluble intercellular adhesion molecule 1 (sICAM-1), soluble vascular adhesion molecule 1 (sVCAM-1), vascular endothelial growth factor (VEGF), VEGF-C, VEGF-D, and placental growth factor (PLGF). Examples of effectors include, but are not limited to, granzyme A, granzyme B, soluble Fas ligand (sFasL), and perforin. Examples of acute phase proteins include, but are not limited to, C-reactive protein (CRP) and serum amyloid A (SAA).

[0109] A "chemokine" is a type of cytokine that mediates chemotaxis or directional movement of cells. Examples of chemokines include, but are not limited to, IL-8, IL-16, eotaxin, eotaxin-3, macrophage-derived chemokine (MDC or CCL22), monocyte chemoattractant protein 1 (MCP-1 or CCL2), MCP-4, macrophage inflammatory protein 1 alpha (MIP-1 alpha, MIP-1a), MIP-1 beta (MIP-1b), gamma-inducible protein 10 (IP-10), and thymus and activation-regulated chemokine (TARC or CCL17).

[0110] A "therapeutically effective amount," "effective dose," "effective amount," or "therapeutically effective dose" of a therapeutic agent, e.g., an engineered CAR T cell, is any amount that, when used alone or in combination with another therapeutic agent, protects a subject from developing a disease or promotes regression of the disease as evidenced by a decrease in the severity of disease symptoms, an increase in the frequency and duration of disease symptom-free periods, or prevention of disability or disability due to disease affliction. The ability of a therapeutic agent to promote regression of a disease can be evaluated using a variety of methods known to those of skill in the art, for example, by assaying the activity of the agent in human subjects during clinical trials, in animal model systems predictive of efficacy in humans, or in in vitro assays. An "effective amount" of an agent (e.g., an inhibitor or combination of inhibitors of GM-CSF, CSF1, GM-CSFR or CSF1R) administered to treat one or more symptoms (e.g., encephalopathy, headache, tremors, dizziness, aphasia, delirium, insomnia, and / or anxiety) of an adverse reaction (e.g., neurotoxicity) is any amount that reduces the severity of one or more of those symptoms and / or increases the frequency and duration of symptom-free periods. The terms "effective amount" and "therapeutically effective amount" may be used interchangeably, where appropriate.

[0111] As used herein, the term "lymphocyte" includes natural killer (NK) cells, T cells, or B cells. NK cells are a type of cytotoxic (cell-toxic) lymphocyte that represents a major component of the genetic immune system. NK cells reject tumor and virus-infected cells. They act through the process of apoptosis, or programmed cell death. They were called "natural killers" because they do not require activation to kill cells. T cells play a major role in cell-mediated immunity (without antibody involvement). Their T cell receptors (TCR) differentiate from other lymphocyte types. The thymus, a specialized organ of the immune system, is primarily responsible for the maturation of T cells. There are six types of T cells: helper T cells (e.g., CD4+ cells), cytotoxic T cells (also known as TCs, cytotoxic T lymphocytes, CTLs, T killer cells, cytolytic T cells, CD8+ T cells or killer T cells), memory T cells ((i) stem memory T cells are CD45RO-, CCR7+, CD45RA+, CD62L+ (L-selectin), CD27+, CD2 8+ and IL-7Rα+, but also express large amounts of CD95, IL-2Rβ, CXCR3, and LFA-1, exhibiting many functional attributes characteristic of memory cells. ), (ii) central memory TCM cells express L-selectin and CCR7 and secrete IL-2, but not IFNγ or IL-4, and (iii) effector memory TEM cells, however, do not express L-selectin or CCR7, but produce effector cytokines such as IFNγ and IL-4), regulatory T cells (Treg, suppressor T cells, or CD4+CD25+ regulatory T cells), natural killer T cells (NKT), and gamma delta T cells. On the other hand, B cells play a major role in humoral immunity (involving antibodies). B cells produce antibodies and antigens, play the role of antigen-presenting cells (APCs), and turn into memory B cells after activation by antigen interaction. In mammals, immature B cells are formed in the bone marrow, from which they get their name.

[0112] The term "genetically engineered" or "engineered" refers to a method of modifying the genome of a cell, including, but not limited to, deleting a coding or non-coding region or portion thereof, or inserting a coding region or portion thereof. In some embodiments, the modified cell is a lymphocyte, e.g., a T cell, and can be obtained from either a patient or a donor. The cell can be modified to express an exogenous construct, e.g., a chimeric antigen receptor (CAR) or a T cell receptor (TCR), which is integrated into the genome of the cell.

[0113] "Immune response" refers to the action of cells of the immune system (e.g., T lymphocytes, B lymphocytes, natural killer (NK) cells, macrophages, eosinophils, mast cells, dendritic cells, and neutrophils) and soluble macromolecules (including Abs, cytokines, and complement) produced either by these cells or the liver, resulting in the selective targeting, binding, damaging, destroying, and / or elimination from the vertebrate body of invading pathogens, pathogen-infected cells or tissues, cancerous or other abnormal cells, or, in the case of autoimmunity or pathological inflammation, normal human cells or tissues.

[0114] The term "immunotherapy" refers to the treatment of a subject suffering from a disease or at risk of suffering from or relapsing from a disease by methods that include inducing, enhancing, suppressing, or otherwise modifying an immune response. Examples of immunotherapy include, but are not limited to, T cell therapy. T cell therapy may include adoptive T cell therapy, tumor infiltrating lymphocyte (TIL) immunotherapy, autologous cell therapy, engineered autologous cell therapy (eACT™), and allogeneic T cell transplantation. However, one of skill in the art will appreciate that the pretreatment methods disclosed herein enhance the efficacy of any transplanted T cell therapy. Examples of T cell therapy are described in U.S. Patent Application Publication Nos. 2014 / 0154228 and 2002 / 0006409, U.S. Patent No. 7,741,465, U.S. Patent No. 6,319,494, U.S. Patent No. 5,728,388, and WO 2008 / 081035.

[0115] T cells for immunotherapy can be derived from any source known in the art. For example, T cells can be differentiated in vitro from a hematopoietic stem cell population, or T cells can be obtained from a subject. T cells can be obtained, for example, from peripheral blood mononuclear cells (PBMCs), bone marrow, lymph node tissue, umbilical cord blood, thymus tissue, tissue from an infection site, ascites, pleural effusion, splenic tissue, and tumors. Additionally, T cells can be derived from one or more T cell lines available in the art. T cells can also be obtained from a unit of blood drawn from a subject using a variety of techniques known to those skilled in the art, such as FICOLL™ separation and / or apheresis. Further methods of isolating T cells for T cell therapy are disclosed in U.S. Patent Application Publication No. 2013 / 0287748, which is incorporated herein by reference in its entirety.

[0116] The term "engineered autologous cell therapy," also known as adoptive cell transfer and which may be abbreviated as "eACT™," refers to the collection of a patient's own T cells and subsequent transfer of one or more specific tumor cells or It is a process of genetic modification to recognize and target one or more antigens expressed on the cell surface of a malignant tumor. T cells can be engineered to express, for example, chimeric antigen receptors (CARs). CAR-positive (+) T cells are engineered to express an extracellular single-chain variable fragment (scFv) specific for a particular tumor antigen linked to an intracellular signaling moiety that includes at least one costimulatory domain and at least one activation domain. CAR scFvs can be designed to target, for example, CD19, a transmembrane protein expressed by cells of the B-cell lineage, including all normal B cells and B-cell malignancies such as, but not limited to, diffuse large B-cell lymphoma (DLBCL), primary mediastinal large B-cell lymphoma, high-grade B-cell lymphoma, and DLBCL arising from follicular lymphoma, NHL, CLL, and non-T-cell ALL. Exemplary CAR T cell therapies and constructs are described in U.S. Patent Application Publication Nos. 2013 / 0287748, 2014 / 0227237, 2014 / 0099309, and 2014 / 0050708, which references are incorporated by reference in their entireties.

[0117] As used herein, a "patient" includes any human suffering from cancer (e.g., lymphoma or leukemia). The terms "subject" and "patient" are used interchangeably herein.

[0118] As used herein, the term "in vitro cells" refers to any cell that is cultured ex vivo. In particular, in vitro cells can include T cells.

[0119] The terms "peptide", "polypeptide" and "protein" are used interchangeably and refer to compounds composed of amino acid residues covalently linked by peptide bonds. A protein or peptide contains at least two amino acids, with no limit on the maximum number of amino acids that may comprise a protein or peptide sequence. A polypeptide includes any peptide or protein that contains two or more amino acids linked together by peptide bonds. As used herein, the term refers to both short chains, also commonly referred to in the art as peptides, oligopeptides and oligomers, for example, and longer chains, generally referred to in the art as proteins, of which there are many varieties. "Polypeptides" include, for example, biologically active fragments, substantially homologous polypeptides, oligopeptides, homodimers, heterodimers, variants of polypeptides, modified polypeptides, derivatives, analogs, fusion proteins, among others. A polypeptide includes natural peptides, recombinant peptides, synthetic peptides, or combinations thereof.

[0120] As used herein, "stimulation" refers to a primary response induced by the binding of a stimulatory molecule to its cognate ligand, which mediates a signal transduction event. A "stimulatory molecule" is a molecule on a T cell, e.g., a T cell receptor (TCR) / CD3 complex, that specifically binds to a cognate stimulatory ligand present on an antigen-presenting cell. A "stimulatory ligand" is a ligand that, when present on an antigen-presenting cell (e.g., an APC, a dendritic cell, a B cell, etc.), specifically binds to a stimulatory molecule on a T cell, thereby mediating a primary response by the T cell, such as, but not limited to, activation, initiation of an immune response, proliferation, etc. Stimulatory ligands include, but are not limited to, anti-CD3 antibodies, peptide-loaded MHC class I molecules, superagonist anti-CD2 antibodies, and superagonist anti-CD28 antibodies.

[0121] As used herein, a "costimulatory signal" refers to a signal that, in combination with a primary signal, such as TCR / CD3 ligation, results in a T cell response, including but not limited to, proliferation and / or upregulation or downregulation of key molecules.

[0122] As used herein, a "costimulatory ligand" refers to a ligand that specifically binds to a cognate costimulatory molecule on a T cell. Costimulatory ligands include molecules on antigen-presenting cells that specifically bind to T cells. Binding of a costimulatory ligand provides a signal that mediates T cell responses, including, but not limited to, proliferation, activation, and differentiation. Costimulatory ligands induce signals, for example, by binding of the T cell receptor (TCR) / CD3 complex to peptide-loaded major histocompatibility complex (MHC) molecules, in addition to the primary signal provided by the stimulatory molecule. Costimulatory ligands can include, but are not limited to, 3 / TR6, 4-1BB ligand, an agonist or antibody that binds to the Toll ligand receptor, B7-1 (CD80), B7-2 (CD86), CD30 ligand, CD40, CD7, CD70, CD83, herpes virus entry mediator (HVEM), human leukocyte antigen G (HLA-G), ILT4, immunoglobulin-like transcript (ILT)3, inducible costimulatory ligand (ICOS-L), intercellular adhesion molecule (ICAM), a ligand that specifically binds to B7-H3, lymphotoxin beta receptor, MHC class I chain-related protein A (MICA), MHC class I chain-related protein B (MICB), OX40 ligand, PD-L2, or programmed death (PD)L1. Costimulatory ligands include, but are not limited to, antibodies that specifically bind to costimulatory molecules present on T cells, such as, but not limited to, 4-1BB, B7-H3, CD2, CD27, CD28, CD30, CD40, CD7, ICOS, a ligand that specifically binds to CD83, lymphocyte function-associated antigen 1 (LFA-1), natural killer cell receptor C (NKG2C), OX40, PD-1, or tumor necrosis factor superfamily member 14 (TNFSF14 or LIGHT).

[0123] A "costimulatory molecule" is a cognate binding partner on a T cell that specifically binds to a costimulatory ligand, thereby mediating a costimulatory response by the T cell, such as, but not limited to, proliferation. Costimulatory molecules include, but are not limited to, a "costimulatory molecule" is a cognate binding partner on a T cell that specifically binds to a costimulatory ligand, thereby mediating a costimulatory response by the T cell, such as, but not limited to, proliferation. Costimulatory molecules include, but are not limited to, 4-1BB / CD137, B7-H3, BAFFR, BLAME (SLAMF8), BTLA, CD33, CD45, CD100 (SEMA4D), CD103, CD134, CD137, CD154, CD16, CD160 (BY55), CD18, CD19, CD19a, CD2, CD22, CD247, CD27, CD276 (B7-H3), CD28, CD29, CD3 (alpha , beta, delta, epsilon, gamma, zeta), CD30, CD37, CD4, CD4, CD40, CD49a, CD49D, CD49f, CD5, CD64, CD69, CD7, CD80, CD83 ligand, CD84, CD86, CD8 alpha, CD8 beta, CD9, CD96 (Tactile), CDl-la, CDl-lb, CDl-lc, CDl-ld, CDS, CEACAM1, CRT AM, DAP-10, DNAM1 (CD226), Fc gamma receptor, GADS, GITR, HVEM (LIGHTR), IA4, ICAM-1, ICAM-1, ICOS, Ig alpha (CD79a), IL2R beta, IL2R gamma, IL7R alpha, integrin, ITGA4, ITGA4, ITGA6, ITGAD, ITGAE, ITGAL, ITGAM, ITGAX, ITGB2, ITGB7, ITGBl, KIRDS2, LAT, LFA-1, LFA-1, LIGHT, LIGHT (tumor necrosis factor superfamily member 14; TNFSF14), LTBR, ​​Ly9 (CD229), lymphocyte function-associated antigen-1 (LFA-1 (CD11 a / CD18), MHC class I molecules, NKG2C, NKG2D, NKp30, NKp44, NKp46, NKp80 (KLRF1), OX40, PAG / Cbp, PD-1, PSGL1, SELPLG (CD162), signaling lymphocyte activation molecule, SLAM (SLAMF1; CD150; IPO-3), SLAMF4 (CD244; 2B4), SLAMF6 (NTB-A; Lyl08), SLAMF7, SLP-76, TNF, TNFr, TNFR2, Toll ligand receptor, TRANCE / RANKL, VLA1, or VLA-6, or fragments, truncations, or combinations thereof.

[0124] The terms "reduce" and "decrease" are used interchangeably herein and indicate any change that is less than the original. "Reduce" and "decrease" are relative terms, requiring a comparison of before and after measurements. "Reduce" and "decrease" include complete depletion.

[0125] "Treatment" or "treating" of a subject refers to any type of intervention or process performed on a subject, or the administration of an active agent to a subject, for the purpose of reversing, mitigating, ameliorating, inhibiting, slowing or preventing the onset, progression, development, severity or recurrence of a symptom, complication or condition, or biochemical manifestations associated with a disease. In some embodiments, "treatment" or "treating" includes partial remission. In other embodiments, "treatment" or "treating" includes complete remission.

[0126] Various aspects of the disclosure are described in further detail in the following subsections.

[0127] Chimeric antigen receptors (CARs) The chimeric antigen receptors (CAR or CAR-T) and T cell receptors (TCR) of the present disclosure are genetically engineered receptors. These engineered receptors can be easily inserted into and expressed by immune cells, such as T cells, according to techniques known in the art. In a CAR, a single receptor can be programmed to recognize a specific antigen and, when bound to that antigen, activate the immune cell to attack and destroy cells bearing or expressing that antigen. If these antigens are present on tumor cells, immune cells expressing the CAR can target and kill the tumor cells.

[0128] One aspect of the present disclosure is a chimeric antigen receptor (CAR) or T cell receptor comprising (i) an antigen binding molecule, (ii) a costimulatory domain, and (iii) an activation domain. The costimulatory domain may comprise an extracellular domain, a transmembrane domain, and an intracellular domain. In some embodiments, the extracellular domain comprises a hinge or truncated hinge domain.

[0129] In some embodiments, an antigen-binding molecule is a molecule that comprises the antigen-binding portion (e.g., CDR) of the antibody from which it is derived. An antigen-binding molecule may comprise an antigen complementarity determining region (CDR). Examples of antigen-binding molecules include, but are not limited to, Fab, Fab', F(ab')2, and Fv fragments, dAbs, linear antibodies, scFv antibodies, and multispecific antibodies formed from antigen-binding molecules. Peptibodies (i.e., Fc fusion molecules containing a peptide binding domain) are another example of a suitable antigen-binding molecule. In one embodiment, the CD19 CAR construct comprises an anti-CD19 single chain Fv. A "single chain Fv" or "scFv" antibody-binding fragment comprises the variable heavy chain (V) of an antibody. H ) domain and the variable light chain (V L) domains, which are present in a single polypeptide chain. Generally, an Fv polypeptide comprises a V H Domain and V L It further comprises a polypeptide linker between the domains which enables the scFv to form the desired structure for antigen binding. All antibody-related terms used herein have their conventional meaning in the art and are well understood by those of skill in the art.

[0130] In some embodiments, the CAR comprises one or more costimulatory domains. In some embodiments, the costimulatory domains include CD28, OX-40, 4-1BB / CD137, CD2, CD7, CD27, CD30, CD40, programmed death-1 (PD-1), inducible T cell costimulatory factor (ICOS), lymphocyte function associated antigen-1 (LFA-1 (CD11a / CD18), CD3 gamma, CD3 delta, CD3 epsilon, CD247, CD276 (B7-H3), LIGHT (tumor necrosis factor superfamily member 14; TNFSF14), NKG2C, Ig alpha (CD79a), DAP-10, Fc gamma receptor, MHC class I molecule, TNF receptor protein, immunoglobulin-like protein, cytokine receptor protein, and the like. receptor, integrin, signaling lymphocyte activation molecule (SLAM protein), activating NK cell receptor, BTLA, Toll ligand receptor, ICAM-1, B7-H3, CDS, ICAM-1, GITR, BAFFR, LIGHT, HVEM (LIGHTR), KIRDS2, SLAMF7, NKp80 (KLRF1), NKp44, NKp30, NKp46, CD19, CD4, CD8 alpha, CD8 beta, IL2R beta, IL2R gamma, IL7R alpha, ITGA4, VLA1, C D49a, ITGA4, IA4, CD49D, ITGA6, VLA-6, CD49f, ITGAD, CD11d, ITGAE, CD103, ITGAL, CD11a, LFA-1, ITGAM, CD11b, ITGAX, CD11c, ITGB1, C D29, ITGB2, CD18, LFA-1, ITGB7, NKG2D, TNFR2, TRANCE / RANKL, DNAM1(CD226), SLAMF4(CD244, 2B4), CD84, CD96(Tactile), CEACAM1, CRT The signal transduction domains are ligands that specifically bind to AM, Ly9 (CD229), CD160 (BY55), PSGL1, CD100 (SEMA4D), CD69, SLAMF6 (NTB-A, Lyl08), SLAM (SLAMF1, CD150, IPO-3), BLAME (SLAMF8), SELPLG (CD162), LTBR, ​​LAT, GADS, SLP-76, PAG / Cbp, CD19a, CD83, or any combination thereof.

[0131] In some embodiments, the intracellular domain is selected from the group consisting of 4-1BB / CD137, activating NK cell receptor, B7-H3, BAFFR, BLAME (SLAMF8), BTLA, CD100 (SEMA4D), CD103, CD160 (BY55), CD18, CD19, CD19a, CD2, CD247, CD27, CD276 (B7-H3), CD29, CD3 delta, CD3 epsilon, CD3 gamma, CD30, CD4, CD40, CD49a, CD49D, CD49f, CD69, CD7, CD84, CD8 alpha, CD8 beta, CD96 (Tactile), CD11a, CD11b, CD11c, CD11d, CDS, CEACAM1, CRT AM, cytokine receptor, DAP-10, DNAM1 (CD226), Fc gamma receptor, GADS, GITR, HVEM (LIGHTR), IA4, ICAM-1, ICAM-1, Ig alpha (CD79a), IL2R beta, IL2R gamma, IL7R alpha, immunoglobulin-like protein, inducible T cell costimulator (ICOS), integrin, ITGA4, ITGA4, ITGA6, ITGAD, ITGAE, ITGAL, ITGAM, ITGAX, ITGB2, ITGB7, ITGB1, KIRDS2, LAT, LFA-1, LFA-1, ligand that specifically binds to CD83, LIGHT, LIGHT (tumor necrosis factor superfamily member 14, TNFSF14), LTBR, ​​Ly9 (C D229), lymphocyte function associated antigen-1 (LFA-1 (CD11a / CD18), MHC class I molecule, NKG2C, NKG2D, NKp30, NKp44, NKp46, NKp80 (KLRF1), OX-40, PAG / Cbp, programmed death 1 (PD-1), PSGL1, SELPLG (CD162), signaling lymphocyte activation molecule (SLAM protein), SLAM (SLAMF1, CD150, IPO-3), SLAMF4 (CD244, 2B4), SLAMF6 (NTB-A, Lyl08), SLAMF7, SLP-76, TNF receptor protein, TNFR2, Toll ligand receptor, TRANCE / RANKL, VLA1, or VLA-6, or combination signaling regions thereof.

[0132] In some embodiments, the CAR comprises a hinge region between the transmembrane domain and the binding molecule. In some embodiments, the hinge region is of IgG1, IgG2, IgG3, IgG4, IgA, IgD, IgE, IgM, CD28, or CD8 alpha. In some embodiments, the transmembrane domain is of CD28, 4-1BB / CD137, the alpha chain of the T cell receptor, the beta chain of the T cell receptor, CD3 epsilon, CD4, CD5, CD8 alpha, CD9, CD16, CD19, CD22, CD33, CD37, CD45, CD64, CD80, CD86, CD134, CD137, CD154, or the T cell receptor. In some embodiments, the activation domain may be derived from, for example, any form of CD3-zeta. In some embodiments, the activation domain is derived from DAP10, DAP12, or other TCR-type activation signaling molecules.

[0133] In one aspect, the present application relates to CD19 CAR T cell therapy. In one embodiment, the CD19 CAR construct comprises an anti-CD19scFv domain, an intracellular domain, a transmembrane domain, one or more costimulatory domains, and an activation domain. In one embodiment, the transmembrane domain is derived from the transmembrane domain of CD28, 4-1BB / CD137, CD8alpha, or any combination thereof. In one embodiment, the costimulatory domain is derived from CD8, CD28, OX40, 4-1BB / CD137, or any combination thereof. In one embodiment, the activation domain is derived from CD3zeta. In one embodiment, the CD19 CAR construct comprises a 4-1BB costimulatory domain. In one embodiment, the CD19 CAR construct comprises a CD28 costimulatory domain. In one embodiment, the CD19 CAR construct comprises an anti-CD19scFv, hinge / transmembrane domain, and costimulatory domain from CD28, and an activation domain from CD3zeta. In one embodiment, the CAR is expressed in axicabtagene ciloleucel. In one embodiment, the CAR is expressed in Kymriah™. Additional CD19-directed CARs that may be used with the methods of the present disclosure include, but are not limited to, JCAR017, JCAR015, JCAR014, Uppsala U. anti-CD19 CAR (NCT02132624), and UCART19 (Celectis), see Sadelain et al., Nature Rev. Cancer Vol. 3 (2003); Ruella et al., Curr Hematol Malig Rep., Springer, NY (2016); and Sadelain et al., Cancer Discovery (Apr 2013).

[0134] Engineered T Cells and Uses The T cells of the immunotherapy can be engineered to express any of the above or other CARs and are called CAR-T cells. The CAR-T cells can be engineered to express other molecules and can be any one of the following exemplary types: first, second, third, fourth, fifth (etc.) CAR-T cells; arm-type CAR-T cells, motile CAR-T cells, TRUCK T cells, switch receptor CAR-T cells; gene-edited CAR T cells; dual receptor CAR T cells; suicide CAR T cells, drug-inducible CAR-T cells, synNotch-inducible CAR T cells; and inhibitory CAR T cells, or others available in the art. In one embodiment, the T cells are autologous T cells. In one embodiment, the T cells are autologous stem cells (for autologous stem cell therapy or ASCT). In one embodiment, the T cells are non-autologous T cells.

[0135] The T cells of the present disclosure may be derived from any source known in the art. For example, T cells may be differentiated in vitro from a hematopoietic stem cell population, or T cells may be obtained from a subject. T cells may be obtained, for example, from peripheral blood mononuclear cells (PBMCs), bone marrow, lymph node tissue, umbilical cord blood, thymus tissue, tissue from an infection site, ascites, pleural effusion, splenic tissue, and tumors. Additionally, T cells may be derived from one or more T cell lines available in the art. T cells may also be obtained from a unit of blood drawn from a subject using various techniques known to those skilled in the art, such as FICOLL™ separation and / or apheresis. Additional methods of isolating T cells for T cell therapy are disclosed in U.S. Patent Application Publication No. 2013 / 0287748, International Application No. PCT / US2015 / 014520 (published as WO 2015 / 120096), and International Application No. PCT / US2016 / 057983 (published as WO 2017 / 070395), all of which are incorporated by reference in their entireties for the purposes of describing these methods.

[0136] CD19 CAR-T cells may be prepared by any manufacturing method that prepares T cells for immunotherapy, including, but not limited to, those described in International Application No. PCT / US2015 / 014520 (published as WO 2015 / 120096) and International Application No. PCT / US2016 / 057983 (published as WO 2017 / 070395), both of which are incorporated by reference in their entirety for the purposes of describing these methods, i.e., any method used for the preparation of axicabtagene ciloleucel or Yescarta®; any method used for the preparation of Tisagenlecleucel / Kymriah™; any method used for the preparation of "off-the-shelf" T cells for immunotherapy; and any other method of preparing lymphocytes for administration to humans. In some embodiments, the manufacturing process is adapted to specifically remove circulating tumor cells from cells obtained from the patient.

[0137] The cells of the present disclosure can be obtained via T cells obtained from a subject. In one embodiment, the T cells can be obtained, for example, from peripheral blood mononuclear cells, bone marrow, lymph node tissue, umbilical cord blood, thymus tissue, tissue from a site of infection, ascites, pleural effusion, spleen tissue, and tumors. Additionally, the T cells can be derived from one or more T cell lines available in the art. The T cells can also be obtained from a unit of blood collected from a subject using various techniques known to those skilled in the art, such as FICOLL™ separation and / or apheresis. In some embodiments, the cells collected by apheresis are washed to remove the plasma fraction and placed into an appropriate buffer or medium for subsequent processing. In some embodiments, the cells are washed with PBS. As will be appreciated, the washing step may be performed, for example, using a semi-automated flow-through centrifuge, such as the Cobe™ 2991 cell processor, Baxter CytoMate™, and the like. In some embodiments, the washed cells are resuspended in one or more biocompatible buffers or other saline solutions with or without buffers. In some embodiments, undesirable components of the apheresis sample are removed. Further methods of isolating T cells for T cell therapy are disclosed in U.S. Patent Application Publication No. 2013 / 0287748, which is incorporated herein by reference in its entirety.

[0138] In some embodiments, T cells are isolated from PBMCs by lysing red blood cells and depleting monocytes, for example using centrifugation through a PERCOLL™ gradient. In some embodiments, specific subpopulations of T cells, such as CD4+, CD8+, CD28+, CD45RA+, and CD45RO+ T cells, are further isolated by positive or negative selection techniques known in the art. For example, enrichment of T cell populations by negative selection can be achieved using a combination of antibodies against surface markers unique to the negatively selected cells. In some embodiments, cell sorting and / or selection may be performed by negative magnetic immunoadhesion or flow cytometry using a cocktail of monoclonal antibodies against cell surface markers present on the negatively selected cells. For example, to enrich for CD4+ cells by negative selection, the monoclonal antibody cocktail typically includes antibodies against CD8, CD11b, CD14, CD16, CD20, and HLA-DR. In some embodiments, flow cytometry and cell sorting are performed to isolate the cell population of interest for use in the present disclosure.

[0139] In some embodiments, PBMCs are used directly for genetic modification of immune cells (such as CARs) using the methods described herein. In some embodiments, after isolating PBMCs, T lymphocytes are further isolated and both cytotoxic and helper T lymphocytes are sorted into subpopulations of naive, memory, and effector T cells before or after genetic modification and / or expansion. In some embodiments, CD8+ cells are further sorted into naive, central memory, and effector cells by identifying cell surface antigens associated with each of these types of CD8+ cells. In some embodiments, expression of phenotypic markers of central memory T cells includes CCR7, CD3, CD28, CD45RO, CD62L, and CD127, and is negative for granzyme B. In some embodiments, central memory T cells are CD8+, CD45RO+, and CD62L+ T cells. In some embodiments, effector T cells are negative for CCR7, CD28, CD62L, and CD127, and positive for granzyme B and perforin. In some embodiments, CD4+ T cells are further classified into subpopulations. For example, CD4+ T helper cells can be sorted into naive cells, central memory cells, and effector cells by identifying cell populations with cell surface antigens.

[0140] In some embodiments, immune cells, e.g., T cells, are genetically modified after isolation using known methods, or immune cells are activated and expanded (or differentiated in the case of precursor cells) in vitro before being genetically modified. In another embodiment, immune cells, e.g., T cells, are genetically modified with a chimeric antigen receptor (e.g., transduced with a viral vector comprising one or more nucleotide sequences encoding a CAR) as described herein, and then activated and / or expanded in vitro. Methods for activating and expanding T cells are known in the art and described, for example, in U.S. Pat. Nos. 6,905,874, 6,867,041, and 6,797,514, and WO 2012 / 079000, the entire contents of which are incorporated herein by reference. In general, such methods include contacting PBMCs or isolated T cells with stimulatory and co-stimulatory agents, e.g., anti-CD3 and anti-CD28 antibodies, typically bound to beads or other surfaces, in culture medium containing appropriate cytokines, such as IL-2. Anti-CD3 and anti-CD28 antibodies bound to the same bead function as "surrogate" antigen-presenting cells (APCs). One example is the Dynabeads® system, a CD3 / CD28 activator / stimulator system for physiological activation of human T cells. In other embodiments, T cells are activated and stimulated to grow with feeder cells and appropriate antibodies and cytokines, for example, using methods described in U.S. Pat. Nos. 6,040,177 and 5,827,642 and WO 2012 / 129514, the contents of which are incorporated herein by reference in their entirety.

[0141] In some embodiments, the T cells are obtained from a donor subject. In some embodiments, the donor subject is a human patient suffering from cancer or a tumor. In some embodiments, the donor subject is a human patient not suffering from cancer or a tumor. In some embodiments, the composition comprises a pharma- ceutically acceptable carrier, diluent, solubilizer, emulsifier, preservative, and / or adjuvant. In some embodiments, the composition comprises an excipient.

[0142] In some embodiments, the composition is selected for delivery via the digestive tract, such as parenterally, by inhalation, or orally. Preparation of such pharmaceutically acceptable compositions is within the capabilities of one of ordinary skill in the art. In some embodiments, a buffer is used to maintain the composition at physiological pH or slightly lower, typically within a pH range of about 5 to about 8. In some embodiments, when parenteral administration is contemplated, the composition is in the form of a pyrogen-free, parenterally acceptable aqueous solution comprising the composition described herein, with or without additional therapeutic agents, in a pharmaceutically acceptable vehicle. In some embodiments, the parenteral injection vehicle is sterile distilled water in which the composition described herein, with or without at least one additional therapeutic agent, is formulated as a properly preserved sterile isotonic solution. In some embodiments, the preparation involves formulating the desired molecule with beads or liposomes, which are polymeric compounds (such as polylactic acid or polyglycolic acid) that allow for controlled or sustained release of the product, which are then delivered by depot injection. In some embodiments, an implantable drug delivery device is used to introduce the desired molecule.

[0143] In some embodiments, the method of treating cancer in a subject in need of treatment comprises T cell therapy. In some embodiments, the T cell therapy disclosed herein is an engineered autologous cell therapy (eACT™). According to this embodiment, the method may include harvesting blood cells from the patient. The isolated blood cells (e.g., T cells) can then be engineered to express a CAR or TCR disclosed herein. In certain embodiments, the CAR T cells or TCR T cells are administered to the patient. In some embodiments, the CAR T cells or TCR T cells treat the tumor or cancer in the patient. In some embodiments, the CAR T cells or TCR T cells reduce the size of the tumor or cancer.

[0144] In some embodiments, donor T cells for use in T cell therapy are obtained from a patient (e.g., for autologous T cell therapy). In other embodiments, donor T cells for use in T cell therapy are obtained from a subject that is not a patient. The T cells may be administered in a therapeutically effective amount. For example, a therapeutically effective amount of T cells is at least about 10 4 Cells, at least about 10 5 Cells, at least about 10 6 Cells, at least about 10 7 Cells, at least about 10 8 Cells, at least about 10 9 , or at least about 10 10 In another embodiment, the therapeutically effective amount of T cells can be about 10 4 cells, about 10 5 cells, about 10 6 cells, about 10 7 cells, or about 10 8 In some embodiments, the therapeutically effective amount of CAR T cells is about 2×10 6 cells / kg, approximately 3×10 6 cells / kg, approximately 4×10 6 cells / kg, approximately 5×10 6 cells / kg, approximately 6×10 6 cells / kg, approximately 7×10 6 cells / kg, approximately 8×10 6 cells / kg, approximately 9×10 6 cells / kg, approximately 1×10 7 cells / kg, approximately 2×10 7 cells / kg, approximately 3×10 7 cells / kg, approximately 4×10 7 cells / kg, approximately 5×10 7 cells / kg, approximately 6×10 7 cells / kg, approximately 7×10 7 cells / kg, approximately 8×10 7 cells / kg, or approximately 9 x 10 7 In some embodiments, the therapeutically effective amount of CAR-positive viable T cells is about 1 x 10 cells / kg of body weight. 6 ~Approx. 2×10 6 CAR-positive viable T cells, the maximum dose is approximately 1 × 10 8In some embodiments, the therapeutically effective amount of CAR-positive viable T cells is about 0.4×10 8 ~Approx. 2×10 8 In some embodiments, the therapeutically effective amount of CAR-positive viable T cells is about 0.4×10 8 , about 0.5×10 8 , about 0.6×10 8 , about 0.7×10 8 , about 0.8×10 8 , about 0.9×10 8 , about 1.0×10 8 , about 1.1×10 8 , about 1.2×10 8 , about 1.3×10 8 , about 1.4×10 8 , about 1.5×10 8 , about 1.6×10 8 , about 1.7×10 8 , about 1.8×10 8 , about 1.9×10 8 , or about 2.0 × 10 8 These are CAR-positive viable T cells.

[0145] Treatment method The methods disclosed herein may be used to treat cancer in a subject, to reduce the size of a tumor, to kill tumor cells, to prevent proliferation of tumor cells, to prevent tumor growth, to eliminate a tumor from a patient, to prevent tumor recurrence, to prevent tumor metastasis, to induce remission in a patient, or any combination thereof. In some embodiments, the methods induce a complete response. In other embodiments, the methods induce a partial response.

[0146] Cancers that may be treated include tumors that are not vascularized, tumors that are not yet substantially vascularized, or tumors that are vascularized. Cancers may also include solid or non-solid tumors. In some embodiments, the cancer is a hematological cancer. In some embodiments, the cancer is a cancer of the white blood cells. In other embodiments, the cancer is a cancer of the plasma cells. In some embodiments, the cancer is a leukemia, lymphoma, or myeloma. In some embodiments, the cancer is acute lymphoblastic leukemia (ALL) (including non-T cell ALL), acute lymphocytic leukemia (ALL) and hemophagocytic lymphohistiocytosis (HLH), B-cell prolymphocytic leukemia, B-cell acute lymphocytic leukemia ("BALL"), blastic plasmacytoid dendritic cell neoplasm, Burkitt's lymphoma, chronic lymphocytic leukemia (CLL), chronic myelogenous leukemia (CML), chronic myelogenous leukemia (CRL), chronic myelogenous leukemia (CRH ... Myeloid leukemia (CML), chronic or acute granulomatous disease, chronic or acute leukemia, diffuse large B-cell lymphoma, diffuse large B-cell lymphoma (DLBCL), follicular lymphoma (FL), hairy cell leukemia, hemophagocytic syndrome (macrophage activation syndrome (MAS), Hodgkin's disease, large cell granuloma, leukocyte adhesion deficiency, malignant lymphoproliferative state, MALT lymphoma, mantle cell lymphoma, marginal lymphoma, monoclonal gammopathy of undetermined significance (MGUS), multiple myeloma, myelodysplasia and myelodysplastic syndromes (MDS), myeloid disorders (AML) including but not limited to acute myeloid leukemia, non-Hodgkin's lymphoma (NHL), plasma cell proliferative disorders ( For example, indolent myeloma (smoldering multiple myeloma or indolent myeloma), plasmablastic lymphoma, plasmacytoid dendritic cell neoplasm, plasmacytoma (e.g., plasma cell dysplasia, solitary myeloma, solitary plasmacytoma, extramedullary plasmacytoma, and multiple myeloma), POEMS syndrome (Crow-Fukase syndrome, Takatsuki disease, PEP syndrome), primary mediastinal large B-cell lymphoma (PMBC), small cell or large cell follicular lymphoma, splenic marginal zone lymphoma (SMZL), systemic amyloid light chain amyloidosis, T-cell acute lymphoblastic leukemia ("TALL"), T-cell lymphoma, transformed follicular lymphoma, Waldenstrom's macroglobulinemia, or a combination thereof.

[0147] In some embodiments, the cancer is a myeloma. In some embodiments, the cancer is multiple myeloma. In some embodiments, the cancer is a leukemia. In some embodiments, the cancer is acute myeloid leukemia. In some embodiments, the CD19-directed genetically modified autologous T-cell immunotherapy is indicated as a first-line therapy, after one line of therapy, or after two or more lines of systemic therapy in the treatment of patients with relapsed or refractory large B-cell lymphoma, such as diffuse large B-cell lymphoma not otherwise specified (DLBCL), primary mediastinal large B-cell lymphoma, high-grade B-cell lymphoma, and DLBCL arising from follicular lymphoma. In some embodiments, the cancer is CLL. In some embodiments, the CD19-directed genetically modified autologous T-cell immunotherapy is axicabtagene ciloleucel (Axi-cel™, YESCARTA®).

[0148] The selected prior therapy may be any prior anti-cancer therapy, such as, but not limited to, a Bruton's tyrosine kinase inhibitor (BTKi), a checkpoint inhibitor (e.g., anti-PD1 antibody, pembrolizumab (Keytruda), cemiplimab (Libtayo), nivolumab (Opdivo); anti-PD-L1 antibody, atezolizumab (Tecentriq), avelumab (Bavencio), durvalumab (Imfinzi); anti-CTLA-4 antibody, ipilimumab (Yervoy)), an anti-CD19 antibody (e.g., blinatumomab), an anti-CD52 antibody (e.g., arentuzumab); an allogeneic stem cell transplant, an anti-CD20 antibody (e.g., rituximab), systemic chemotherapy, rituximab, anthracyclines, ofatumumab, and combinations thereof. The prior therapy may also be administered in combination with the CD19 CAR T therapy of the present disclosure. In one embodiment, eligible patients may have disease refractory to most recent treatment or may have relapsed within one year after autologous hematopoietic stem cell transplantation (HSCT / ASCT).

[0149] In some embodiments, the method further comprises administering a chemotherapeutic agent. In some embodiments, the chemotherapeutic agent selected is a lymphodepleting (preconditioning) chemotherapeutic agent. Beneficial preconditioning treatment regimens, along with correlating beneficial biomarkers, are described in U.S. Provisional Patent Applications Nos. 62 / 262,143 and 62 / 167,750, which are incorporated by reference in their entireties. These include, for example, methods of pretreating a patient in need of T cell therapy, including administering a designated beneficial dose of cyclophosphamide (200 mg / m 2 / day~2000mg / m 2 / day) and the specified dose of fludarabine (20 mg / m 2 / day~900mg / m 2 / day) to the patient. One such dosing regimen comprises administering to the patient about 500 mg / m2 of engineered T cells prior to administering to the patient a therapeutically effective amount of engineered T cells. 2 Cyclophosphatase / day mide, and about 60 mg / m 2 In one embodiment, the conditioning regimen comprises administering fludarabine at 500 mg / m2 for three days. 2 + Fludarabine 30 mg / m 2 In some embodiments, they are administered on days -4, -3, and -2. In some embodiments, they are administered on days -5, -4, and -3. In one embodiment, the conditioning regimen includes 900 mg / m on day -2. 2 , and fludarabine 25 mg / m on days −4, −3, and −2. 2 (Day 0 is the day the cells are administered. In some embodiments, the conditioning regimen includes cyclophosphamide 500 mg / m daily for 2 days. 2 , and fludarabine 30 mg / m for 4 days. 2In some embodiments, the antigen binding molecules, transduced (or otherwise engineered) cells (such as CARs), and chemotherapeutic agents are each administered in an amount effective to treat the disease or condition of the subject, either alone or in combination with other agents and treatments described herein.

[0150] In some embodiments, compositions comprising the CAR-expressing immune effector cells disclosed herein can be administered in conjunction with a variety of chemotherapeutic agents. Exemplary chemotherapeutic agents include alkylating agents, such as thiotepa and cyclophosphamide (CYTOXAN™); alkylsulfonates, such as busulfan, improsulfan, and piposulfan; aziridines, such as benzodopa, carboquone, metholedopa, and uredopa; altretamine, triethylenemelamine, triethylenephosphoramide, triethylenethiophosphamide, ethylenimines and methylamelamines, including trimethylmelamine regimes; nitrogen mustards, such as chloroquine, cyclophosphamide, and cyclophosphamide; Lambucil, Chlornaphazine, Chlorophosphamide, Estramustine, Ifosfamide, Mechlorethamine, Mechlorethamine Oxide Hydrochloride, Melphalan, Novenbitine, Phenesterine, Prednimustine, Trofosfamide, Uracil Mustard; Nitrosoureas, e.g., Carmustine, Chlorozotocin, Fotemustine, Lomustine, Nimustine, Ranimustine; Antibiotics, e.g., Aclacinomycin, Actinomycin, Ausramycin, Azaserine, Bleomycin, Cactinomycin , calicheamicin, carabicin, carminomycin, carzinophilin, chromomycin, dactinomycin, daunorubicin, detrevicin, 6-diazo-5-oxo-L-norleucine, doxorubicin, epirubicin, esorubicin, idarubicin, marcellomycin, mitomycin, mycophenolic acid, nogalamycin, olivomycin, peplomycin, pofilomycin, puromycin, queramycin, rodorubicin, streptonigrin, streptozocin, tubercidin, ubenime antimetabolites such as methotrexate, and 5-fluorouracil (5-FU); folic acid analogues such as denopterin, methotrexate, pteropterin, trimetrexate; purine analogues such as fludarabine, 6-mercaptopurine, thiamiprine, thioguanine; pyrimidine analogues such as ancitabine, azacitidine, 6-azauridine, carmoful, cytarabine, dideoxyuridine, doxifluridine, enocitabine, floxuridine, 5-FU;Androgens, e.g., calsterone, dromostanolone propionate, epithiostanol, mepitiostane, testolactone;Antiadrenergics, e.g., aminoglutethimide, mitotane, trilostane;Folic acid supplements, e.g., furoic acid;Aceglatone;Aldophosphamide glycosides;Aminolevulinic acid;Amsacrine;Bestravcil;Bisantrene;Edatraxate;Defofamine;Demecolcine;Diazicon;Erformitin;Elliptinium acetate;Toglucid;Gallium nitrate;Hydroxyurea;Lentinan;Lonidamine;Mitogua Zon; Mitoxantrone; Mopidamol; Nitracrine; Pentostatin; Phenamet; Pirarubicin; Podophyllic acid; 2-Ethylhydrazide; Procarbazine; PSK (registered trademark); Razoxane; Sizofiran; Spirogermanium; Tenuazonic acid; Triazicon; 2,2',2"-Trichlorotriethylamine; Urethane; Vindesine; Dacarbazine; Mannomustine; Mitobronitrol; Mitolactol; Pipobroman; Gacytosine; Arabinoside ("Ara-C"); Cyclophosphamide; Thiotepa; Taxoids, e.g., Paclitaxel; cetaxel (TAXOL™, Bristol-Myers Squibb) and doxetaxel (TAXOTERE®, Rhone-Poulenc Rorer); chlorambucil; gemcitabine; 6-thioguanine; mercaptopurine; methotrexate; platinum analogues such as cisplatin and carboplatin; vinblastine; platinum; etoposide (VP-16); ifosfamide; mitomycin C; mitoxantrone; vincristine; vinorelbine; navelbine; novantrone; teniposide; daunomycin; aminopterin; xeloda; ibandronate; CPT-11; topoisomerase inhibitor RFS2000; difluoromethylomitin (DMFO); retinoic acid derivatives such as Targretin™ (bexarotene), Panretin™ (alitretinoin); ONTAK™ (denileukin diftitox); esperamycin; capecitabine; and pharmaceutically acceptable salts, acids, or derivatives of any of the above. In some embodiments, compositions comprising the CAR and / or TCR expressing immune effector cells disclosed herein can be administered in conjunction with anti-hormonal agents that act to regulate or inhibit hormone action on tumors, such as antiestrogens, such as tamoxifen, raloxifene, aromatase-inhibiting 4(5)-imidazole, 4-hydroxytamoxifen, trioxyphene, ketoxifene, LY117018, onapristone and toremifene (Fareston); and anti-androgens, such as flutamide, nilutamide, bicalutamide, leuprolide, and goserelin; as well as pharmacologic acceptable salts, acids, or derivatives of any of the above. Combinations of chemotherapeutic agents, including, but not limited to, CHOP, i.e., cyclophosphamide (Cytoxan®), doxorubicin (hydroxydoxorubicin), vincristine (Oncovin®), and prednisone, are also administered as appropriate.

[0151] In some embodiments, the chemotherapeutic agent is administered simultaneously with or within one week after administration of the engineered cells or nucleic acid. In other embodiments, the chemotherapeutic agent is administered 1-4 weeks, or 1 week-1 month, 1 week-2 months, 1 week-3 months, 1 week-6 months, 1 week-9 months, or 1 week-12 months after administration of the engineered cells or nucleic acid. In some embodiments, the chemotherapeutic agent is administered at least one month prior to administration of the cells or nucleic acid. In some embodiments, the method further comprises administering two or more chemotherapeutic agents.

[0152] A variety of additional therapeutic agents can be used in conjunction with the compositions or agents / treatments described herein. For example, potentially useful additional therapeutic agents include PD-1 inhibitors such as nivolumab (OPDIVO®), pembrolizumab (KEYTRUDA®), pembrolizumab, pidilizumab (CureTech), and atezolizumab (Roche). Additional therapeutic agents suitable for use in combination with the compositions or agents / treatments and methods disclosed herein include ibrutinib (IMBRUVICA®), ofatumumab (ARZERRA®), rituximab (RITUXAN®), bevacizumab (AVASTIN®), trastuzumab (HERCEPTIN®), trastuzumab emtansine (KADCYLA®), imatinib (GLEEVEC®), cetuximab (ERBITUX®), panitumumab (VECTIBIX®), catumaxomab, ibritumomab, ofatumumab, tositumomab, buccal, and cefotaxime. Lentuximab, Alemtuzumab, Gemtuzumab, Erlotinib, Gefitinib, Vandetanib, Afatinib, Lapatinib, Neratinib, Axitinib, Masitinib, Pazopanib, Sunitinib, Sorafenib, Tocilizumab, Toceranib, Lestaurtinib, Axitinib, Cediranib, Lenvatinib, Nintedanib, Pazopanib Nitrobacter, regorafenib, semaxanib, sorafenib, sunitinib, tivozanib, toceranib, vandetanib, entrectinib, carbozantinib, imatinib, dasatinib, nilotinib, ponatinib, radotinib, bosutinib, lestaurtinib, ruxolitinib, pacritinib, cobimetinib, selumetinib, trametinib mTOR inhibitors such as rivaroxaban, binimetinib, alectinib, ceritinib, crizotinib, aflibercept, adipotide, denileukin diftitox, everolimus and temsirolimus, hedgehog inhibitors such as sonidegib and vismodegib, CDK inhibitors such as the CDK inhibitor palbociclib.

[0153] In some embodiments, the composition or drug / treatment comprising CAR immune cells is administered with an anti-inflammatory agent. Anti-inflammatory agents or drugs may include, but are not limited to, steroids and glucocorticoids (including betamethasone, budesonide, dexamethasone, hydrocortisone acetate, hydrocortisone, hydrocortisone, methylprednisolone, prednisolone, prednisone, triamcinolone), nonsteroidal anti-inflammatory drugs (NSAIDS) including aspirin, ibuprofen, naproxen, methotrexate, sulfasalazine, leflunomide, anti-TNF drugs, cyclophosphamide and mycophenolic acid. Exemplary NSAIDs include ibuprofen, naproxen, naproxen sodium, Cox-2 inhibitors, and sialylate. Exemplary analgesics include acetaminophen, oxycodone, tramadol of proporoxifene hydrochloride. Exemplary glucocorticoids include cortisone, dexamethasone, hydrocortisone, methylprednisolone, prednisolone, or prednisone. Exemplary biological response modifiers include molecules against cell surface markers (e.g., CD4, CD5, etc.), cytokine inhibitors, such as TNF antagonists (e.g., etanercept (ENBREL®), adalimumab (HUMIRA®) and infliximab (REMICADE®), chemokine inhibitors, and adhesion molecule inhibitors. Biological response modifiers include monoclonal antibodies and recombinant forms of molecules. Exemplary DMARDs include azathioprine, cyclophosphamide, cyclosporine, methotrexate, penicillamine, leflunomide, sulfasalazine, hydroxychloroquine, Gold (oral (auranofin) and intramuscular), and minocycline.

[0154] In some embodiments, the compositions or agents / treatments described herein are administered with a cytokine. Examples of cytokines are lymphokines, monokines, and traditional polypeptide hormones. Cytokines include growth hormones, such as human growth hormone, N-methionyl human growth hormone, and bovine growth hormone; parathyroid hormone; thyroxine; insulin; proinsulin; relaxin; prorelaxin; glycoprotein hormones, such as follicle-stimulating hormone (FSH), thyroid-stimulating hormone (TSH), luteinizing hormone (LH); hepatic growth factor (HGF); fibroblast growth factor (FGF); prolactin; placental lactogen; murelan inhibitor; mouse gonadotropin-related peptide; inhibin; activin; vascular endothelial growth factor; integrins; thrombopoietin (TPO); nerve growth factor (NGF), such as NGF-beta; platelet growth factor; transforming growth factor (TGF), such as TGF-alpha and TGF-beta; insulin-like growth factor (IGGF), ... Examples of cytokines include growth factor-I and growth factor-II; erythropoietin (EPO, Epogen®, Procrit®); bone morphogenetic factors; interferons, e.g., interferon-alpha, beta, and -gamma; colony stimulating factors (CSF), e.g., macrophage-CSF (M-CSF); granulocyte-macrophage CSF (GM-CSF); and granulocyte CSF (G-CSF); interleukins (IL), e.g., IL-1, IL-1 alpha, IL-2, IL-3, IL-4, IL-5, IL-6, IL-7, IL-8, IL-9, IL-10, IL-11, IL-12; IL-15, tumor necrosis factors, e.g., TNF-alpha or TNF-beta; and other polypeptide factors, including LIF and Kit Ligand (KL). As used herein, the term cytokine includes proteins from natural sources or from recombinant cell culture and biologically active equivalents of the native sequence cytokines. In some embodiments, the compositions described herein are administered with a steroid.

[0155] Administration of CD19-directed genetically modified autologous T cell immunotherapy Indications and Use In some embodiments, the CD19-directed genetically modified autologous T-cell immunotherapy is indicated in the treatment of adult patients with relapsed or refractory large B-cell lymphoma, such as diffuse large B-cell lymphoma (DLBCL) not otherwise specified, primary mediastinal large B-cell lymphoma, high-grade B-cell lymphoma, and DLBCL arising from follicular lymphoma, after two or more lines of systemic therapy. In some embodiments, the CD19-directed genetically modified autologous T-cell immunotherapy is not indicated in the treatment of patients with primary central nervous system lymphoma. In some embodiments, the immunotherapy is indicated in the treatment of ALL or CLL. In some embodiments, the immunotherapy is indicated in the first line of treatment. In some embodiments, the immunotherapy is indicated for administration after one line of treatment. Other indications can be found throughout this disclosure.

[0156] Dosage and Administration In some embodiments, an infusion bag of CD19-directed genetically modified autologous T cell immunotherapy contains about 68 mL of a suspension of chimeric antigen receptor (CAR)-positive T cells. The target dose is about 1 x 10 per kg of body weight. 6 ~Approx. 2×10 6 CAR-positive viable T cells, up to 2 × 10 8 It can be a CAR positive viable T cell. In some embodiments, the CD19 directed genetically modified autologous T cell immunotherapy is Axi-cel™ (YESCARTA®, axi-captagen ciloleucel). The amount of CAR T cells, dosing regimen, method of administration, subject, and cancer within these methods are described elsewhere in this disclosure, alone or in combination with another chemotherapeutic agent, with or without preconditioning, for any of the patients described elsewhere in this disclosure.

[0157] In some embodiments, the CD19-directed genetically modified autologous T-cell immunotherapy is for autologous use. The patient's identity must match the patient identifier on the CD19-directed genetically modified autologous T-cell immunotherapy cassette and infusion bag. If the information on the patient-specific label does not match the intended patient, the CD19-directed genetically modified autologous T-cell immunotherapy cannot be administered. In some embodiments, the availability of CD19-directed genetically modified autologous T-cell immunotherapy must be confirmed before starting the lymphodepletion regimen. In some embodiments, the immunotherapy is performed using allogeneic "off-the-shelf" lymphocytes.

[0158] In some embodiments, patients are pretreated with lymphodepleting chemotherapy prior to infusion of CD19-directed genetically modified autologous T cell immunotherapy. In some embodiments, patients are pretreated with cyclophosphamide 500 mg / m 5, 4, and 3 days prior to infusion of CD19-directed genetically modified autologous T cell immunotherapy. 2 IV, and fludarabine 30 mg / m 2 IV lymphodepleting chemotherapy regimens are administered. Other beneficial preconditioning treatment regimens, along with correlating beneficial biomarkers, include those described in U.S. Provisional Patent Applications Nos. 62 / 262,143 and 62 / 167,750, which are incorporated by reference in their entireties. These include, for example, a method of pretreating a patient in need of T cell therapy, comprising administering a designated beneficial dose of cyclophosphamide (200 mg / m 2 / day~2000mg / m 2 / day) and the specified dose of fludarabine (20 mg / m 2 / day~900mg / m 2 / day) to the patient. One such dosing regimen comprises administering to the patient about 500 mg / m2 of engineered T cells prior to administering to the patient a therapeutically effective amount of engineered T cells. 2 / day cyclophosphamide, and approximately 60 mg / m 2The present invention includes treating a patient with fludarabine administered at least once a day for three days at a dose of 100 mg / day of fludarabine. Other examples of such regimens can be found, for example, in U.S. Patent No. 9,855,298.

[0159] In some embodiments, the patient is premedicated orally with a dose of about 500-1000 mg, about 600-1000 mg, about 700-1000 mg, about 800-1000 mg, about 900-1000 mg, about 500-900 mg, about 500-800 mg, about 500-700 mg, about 500-600 mg, about 600-900 mg, about 600-800 mg, about 600-700 mg, about 700-900 mg, about 700-800 mg, or about 800-900 mg of acetaminophen prior to infusion of the CD19-directed genetically modified autologous T cell immunotherapy. In some embodiments, the patient is premedicated with a dose of about 500 mg, about 525 mg, about 550 mg, about 575 mg, about 600 mg, about 625 mg, about 650 mg, about 675 mg, about 700 mg, about 725 mg, about 750 mg, about 775 mg, about 800 mg, about 825 mg, about 850 mg, about 875 mg, about 900 mg, about 925 mg, about 950 mg, about 975 mg, or about 1000 mg of acetaminophen administered orally prior to the infusion of the CD19-directed genetically modified autologous T cell immunotherapy. In some embodiments, the patient is premedicated with 650 mg of acetaminophen administered orally prior to the infusion of the CD19-directed genetically modified autologous T cell immunotherapy and with 12.5 mg of diphenhydramine administered intravenously or orally about 1 hour prior to the infusion of the CD19-directed genetically modified autologous T cell immunotherapy. In some embodiments, the prophylactic use of systemic steroids is avoided as they may interfere with the activity of CD19-directed genetically modified autologous T cell immunotherapy.

[0160] Preparation of CD19-directed genetically modified autologous T cell immunotherapy for infusion The timing of thawing and infusion of the CD19-directed genetically modified autologous T-cell immunotherapy is coordinated, in some embodiments, the infusion time is pre-confirmed and the start time of thawing of the CD19-directed genetically modified autologous T-cell immunotherapy is coordinated so that it is available for infusion when the patient is ready.

[0161] In some embodiments, the patient's identity is confirmed before the CD19-directed genetically modified autologous T-cell immunotherapy is thawed. Prior to formulating the CD19-directed genetically modified autologous T-cell immunotherapy, the patient's identity is matched with the patient identifier on the CD19-directed genetically modified autologous T-cell immunotherapy cassette. In some embodiments, if the information on the patient-specific label does not match the intended patient, the CD19-directed genetically modified autologous T-cell immunotherapy product bag is not removed from the cassette.

[0162] In some embodiments, the patient's identity is confirmed before the CD19-directed genetically modified autologous T cell immunotherapy product bag is removed from the cassette and the patient information on the cassette label is confirmed to match the bag label. In some embodiments, the method includes inspecting the product bag for any defects related to the integrity of the container, e.g., breaks or cracks, prior to thawing. In some embodiments, the infusion bag is placed into a second sterile bag in accordance with local guidelines.

[0163] In some embodiments, the method includes thawing the CD19-directed genetically modified autologous T-cell immunotherapy at about 37° C. using either a water bath method or a dry thawing method until there is no visible ice in the infusion bag. In some embodiments, the method includes mixing or agitating the contents of the bag to disperse clumps of cellular material. In some embodiments, the contents of the bag are gently mixed or agitated. In some embodiments, the method includes inspecting the bag for remaining visible clumps of cells and continuing to mix or agitate. Small clumps of cellular material may require gentle manual mixing to disperse. In some embodiments, the method does not include washing, spinning down and / or resuspending the CD19-directed genetically modified autologous T-cell immunotherapy with fresh media prior to infusion. In some embodiments, once thawed, the CD19-directed genetically modified autologous T-cell immunotherapy may be stored at room temperature (20° C.-25° C.) for up to 3 hours. In some embodiments, the immunotherapy is administered immediately.

[0164] Administration In some embodiments, the methods of administration of a CD19-directed genetically modified autologous T cell immunotherapy of the present disclosure include one or more of the following as steps or considerations: Ensure that tocilizumab and emergency equipment are available before the infusion and during the recovery period. Do not use a leukocyte removal filter. · Central venous access is recommended for infusion of CD19-directed genetically modified autologous T-cell immunotherapy. · Verify that the patient's identity matches the patient identifier on the CD19-directed genetically modified autologous T cell immunotherapy product bag. Prime the tubing with saline before injection. Infuse the entire contents of the CD19-directed genetically modified autologous T-cell immunotherapy bag within 30 minutes, either by gravity or a peristaltic pump. The CD19-directed genetically modified autologous T-cell immunotherapy is stable at room temperature for up to 3 hours after thawing. Gently agitate the product bag during infusion of the CD19-directed genetically modified autologous T cell immunotherapy to prevent cell clumping. After injecting all of the contents of the product bag, rinse the tubing with saline at the same injection rate to ensure all product is delivered. CD19-directed genetically modified autologous T cell immunotherapy comprises human blood cells genetically modified with a replication-incompetent retroviral vector. Universal precautions and local biosafety guidelines should be followed for handling and disposal to avoid potential transmission of infectious diseases.

[0165] monitoring In some embodiments, administration of the CD19-directed genetically modified autologous T cell immunotherapy is performed in a certified medical facility. In some embodiments, the methods disclosed herein include monitoring the patient daily for at least 7 days at a certified medical facility after the infusion for signs and symptoms of CRS and neurotoxicity. In some embodiments, the methods disclosed herein include monitoring the patient daily for at least 10 days at a certified medical facility after the infusion for signs and symptoms of CRS and neurotoxicity. In some embodiments, the patient is instructed to remain near the certified medical facility for at least 4 weeks after the infusion.

[0166] Management of severe adverse reactions In some embodiments, the method includes managing adverse reactions. In some embodiments, the adverse reactions are selected from the group consisting of cytokine release syndrome (CRS), neurotoxicity, hypersensitivity reactions, serious infections, cytopenias, and hypogammaglobulinemia. In some embodiments, neutralization or reduction of the CSF / CSFR1 axis is used to prevent or treat the adverse reactions, alone or in combination with other treatments. In some embodiments, prevention and / or treatment of adverse reactions and their syndromes is performed with agents such as tocilizumab (or another anti-IL6 / IL6R agent / antagonist) and / or steroids (e.g., corticosteroids).

[0167] In some embodiments, the agent is an antagonist or inhibitor of TL-6 or IL-6 receptor (TL-6R). In some embodiments, the agent is an antibody that neutralizes TL-6 activity, e.g., an antibody or antigen-binding fragment that binds to TL-6 or IL-6R. For example, in some embodiments, the agent is or includes an anti-IL-6R antibody that is tocilizumab (atlizumab) or sarilumab. In some embodiments, the agent is an anti-IL-6R antibody described in U.S. Pat. No. 8,562,991. In some cases, the agent targeting IL-6 is an anti-TL-6 antibody, e.g., siltuximab, elcilimomab, ALD518 / BMS-945429, sirukumab (CNTO 136), C-cell activator ... PSI-2634, ARGX 109, FE301, FM101 or olokizumab (CDP6038). In some embodiments, the agent may neutralize IL-6 activity by inhibiting ligand-receptor interaction. In some embodiments, the IL-6 / IL-6R antagonist or inhibitor is an IL-6 mutein, such as those described in U.S. Pat. No. 5,591,827. In some embodiments, the agent that is an antagonist or inhibitor of IL-6 / IL-6R is a small molecule, a protein or peptide, or a nucleic acid.

[0168] In some embodiments, other agents that can be used to manage adverse reactions and their symptoms include cytokine receptor or cytokine antagonists or inhibitors. In some embodiments, the cytokine or receptor is IL-10, TL-6, TL-6 receptor, IFNy, IFNGR, IL-2, IL-2R / CD25, MCP-1, CCR2, CCR4, MIP13, CCR5, TNFalpha, TNFR1, e.g., TL-6 receptor (IL-6R), IL-2 receptor (IL-2R / CD25), MCP-1 (CCL2) receptor (CCR2 or CCR4), TGF-beta receptor (TGF-βI, II, or III), IFN-gamma receptor (IFNGR), MIP1P receptor (e.g., CCR5), TNFalpha receptor (e.g., TNFR1), IL-1 receptor (IL1-Ra / IL-1RP) or IL-10 receptor (IL-10R), IL-1 and IL-1Ralpha / IL-1beta. In some embodiments, the agent comprises cituximab, sarilumab, olokizumab (CDP6038), elcilimomab, ALD518 / BMS-945429, sirukumab (CNTO 136), CPSI-2634, ARGX 109, FE301, or FM101. In some embodiments, the agent is a cytokine antagonist or inhibitor, such as transforming growth factor beta (TGF-beta), interleukin 6 (TL-6), interleukin 10 (IL-10), IL-2, MIP13 (CCL4), TNF alpha, IL-1, interferon gamma (IFN-gamma), or monocyte chemotactic protein-I (MCP-1). In some embodiments, the compound targets (e.g., inhibits or is an antagonist of) a cytokine receptor, such as IL-6 receptor (IL-6R), IL-2 receptor (IL-2R / CD25), MCP-1 (CCL2) receptor (CCR2 or CCR4), TGF-beta receptor (TGF-beta I, II, or III), IFN-gamma receptor (IFNGR), MIP1P receptor (e.g., CCR5), TNF alpha receptor (e.g., TNFR1), IL-1 receptor (IL1-Ra / IL-1RP), or IL-10 receptor (IL-10R).

[0169] In some embodiments, tocilizumab is administered at a dosage of 1 mg / kg to 10 mg / kg (or about 1 mg / kg to 10 mg / kg), 2 mg / kg to 8 mg / kg (or about 2 mg / kg to 8 mg / kg), 2 mg / kg to 6 mg / kg (or about 2 mg / kg to 6 mg / kg), 2 mg / kg to 4 mg / kg (or about 2 mg / kg to 4 mg / kg), or 6 mg / kg to 8 mg / kg (or about 6 mg / kg to 8 mg / kg), or tocilizumab is administered at a dosage of at least or at least about, 2 mg / kg, 4 mg / kg, 6 mg / kg, or 8 mg / kg. In some embodiments, tocilizumab is administered at a dosage of about 1 mg / kg to 12 mg / kg, e.g., 10 mg / kg, or about 10 mg / kg. In some embodiments, tocilizumab is administered by intravenous infusion.

[0170] In some embodiments, the signs and symptoms of the adverse reaction are selected from the group consisting of fever, hypotension, tachycardia, hypoxia, and chills, and include cardiac arrhythmias (including atrial fibrillation and ventricular tachycardia), cardiac arrest, cardiac failure, renal insufficiency, capillary leak syndrome, hypotension, hypoxia, organ toxicity, hemophagocytic lymphohistiocytosis / macrophage activation syndrome (HLH / MAS), seizures, encephalopathy, headache, tremor, dizziness, aphasia, delirium, insomnia anxiety, anaphylaxis, febrile neutropenia Symptoms include thrombocytopenia, thrombocytopenia, neutropenia, and anemia.

[0171] Cytokine release syndrome In some embodiments, the method includes identifying cytokine release syndrome (CRS) based on clinical symptoms. In some embodiments, the method includes evaluating and treating other causes of fever, hypoxia, and hypotension. If CRS is observed or suspected, it can be managed according to the recommendations in Table 1, which can also be used in combination with other therapies of the present disclosure, such as neutralization or reduction of the CSF / CSFR1 axis. Patients experiencing grade 2 or higher CRS (e.g., hypotension unresponsive to fluids, or hypoxia requiring supplemental oxygenation) should be monitored with continuous cardiac telemetry and pulse oximetry. In some embodiments, for patients experiencing severe CRS, consider performing an echocardiogram to assess cardiac function. In cases of severe or life-threatening CRS, intensive supportive care may be considered. In some embodiments, a biosimilar to tocilizumab or its equivalent may be used in place of tocilizumab in the methods disclosed herein. In other embodiments, another anti-IL6R may be used in place of tocilizumab. [Table 1]

[0172] Neurotoxicity In some embodiments, the method includes monitoring the patient for signs and symptoms of neurotoxicity (Table 2). In some embodiments, the method includes ruling out other causes of neurological symptoms. Patients experiencing grade 2 or higher neurotoxicity should be monitored with continuous cardiac telemetry and pulse oximetry. Severe or life-threatening neurotoxicity If severe, provide intensive supportive care. Consider a non-sedating anti-seizure medication (e.g., levetiracetam) for seizure prophylaxis for any grade 2 or higher neurotoxicity. The following treatments may be used in combination with other treatments of the present disclosure, such as neutralization or reduction of the CSF / CSFR1 axis. [Table 2]

[0173] In one embodiment, the disclosure provides a method of treating relapsed or refractory large B-cell lymphoma after two or more lines of systemic therapy in a human, comprising administering to a human in need of treatment a CD19-directed genetically modified autologous T-cell immunotherapy, (a) administering to a patient a composition comprising CD19-directed chimeric antigen receptor (CAR)-positive viable T cells; (b) monitoring patients after administration for signs and symptoms of adverse reactions; and (c) If greater than Grade 2 cytokine release syndrome (CRS) is observed in (b), administer tocilizumab IV at a dose of approximately 8 mg / kg over 1 hour, repeating tocilizumab every 8 hours as needed if not responsive to increased IV fluids or supplemental oxygen; (d) if the CRS symptoms observed in (b) do not improve 24 hours after (c), administer approximately 1 mg / kg IV methylprednisolone twice daily or an equivalent dose of dexamethasone and continue corticosteroids until events are Grade 1 or less, then taper over 3 days; (e) if Grade 3 CRS is observed in (b), administer tocilizumab at a dose of 8 mg / kg IV over 1 hour, repeat every 8 hours as needed if unresponsive to IV fluids or increased supplemental oxygen, administer methylprednisolone 1 mg / kg twice daily or an equivalent dose of dexamethasone, and continue corticosteroids until events are Grade 1 or less, then taper over 3 days; and (f) if CRS grade 4 is observed in (b), administering tocilizumab at a dose of about 8 mg / kg IV over 1 hour, repeating tocilizumab every 8 hours as needed if unresponsive to IV fluids or increased supplemental oxygen, and administering methylprednisolone IV at about 1,000 mg per day for 3 days.

[0174] In another embodiment, the disclosure provides a method of treating relapsed or refractory large B-cell lymphoma after two or more lines of systemic therapy in a patient, comprising administering to a patient in need of treatment a CD19-directed genetically modified autologous T-cell immunotherapy, (a) administering to a patient a composition comprising CD19-directed chimeric antigen receptor (CAR)-positive viable T cells; (b) monitoring patients after administration for signs and symptoms of adverse reactions; and (c) if cytokine release syndrome (CRS) and / or neurotoxicity is observed, managing cytokine release syndrome (CRS) and / or neurotoxicity according to Table 1 and / or Table 2.

[0175] Further safety management measures with corticosteroids

[0176] Administration of corticosteroids and / or tocilizumab in grade 1 can be considered preventive. Supportive care can be provided in all protocols for all CRS and NE severity grades. In one embodiment of the protocol for managing adverse events related to CRS, tocilizumab and / or corticosteroids are administered as follows: Grade 1 CRS: no tocilizumab; no corticosteroids; Grade 2 CRS: tocilizumab (only if comorbidity or elderly); and / or corticosteroids (only if comorbidity or elderly); Grade 3 CRS: tocilizumab; and / or corticosteroids; Grade 4 CRS: tocilizumab; and / or corticosteroids. In another embodiment of the protocol for managing adverse events associated with CRS, tocilizumab and / or corticosteroids are administered as follows: Grade 1 CRS: tocilizumab (if no improvement after 3 days); and / or corticosteroids (if no improvement after 3 days); Grade 2 CRS: tocilizumab; and / or corticosteroids; Grade 3 CRS: tocilizumab; and / or corticosteroids; Grade 4 CRS: tocilizumab; and / or corticosteroids, high dose. In one embodiment of the protocol for managing adverse events related to NE, tocilizumab and / or corticosteroids are administered as follows: Grade 1 NE: no tocilizumab; no corticosteroids; Grade 2 NE: no tocilizumab; no corticosteroids; Grade 3 NE: tocilizumab; and / or corticosteroids (standard dose only if no improvement to tocilizumab); Grade 4 NE: tocilizumab; and / or corticosteroids. In another embodiment of the protocol for managing adverse events associated with NE, tocilizumab and / or corticosteroids are administered as follows: Grade 1 NE: no tocilizumab; and / or corticosteroids; Grade 2 NE: tocilizumab; and / or corticosteroids; Grade 3 NE: tocilizumab; and / or corticosteroids, high dose; Grade 4 NE: tocilizumab; and / or corticosteroids, high dose.

[0177] In one embodiment, corticosteroid treatment is initiated for CRS of grade 2 or higher and tocilizumab treatment is initiated for CRS of grade 2 or higher. In one embodiment, corticosteroid treatment is initiated for CRS of grade 1 or higher and tocilizumab treatment is initiated for CRS of grade 1 or higher. In one embodiment, corticosteroid treatment is initiated for NE of grade 3 or higher and tocilizumab treatment is initiated for CRS of grade 3 or higher. In one embodiment, corticosteroid treatment is initiated for CRS of grade 1 or higher and tocilizumab treatment is initiated for CRS of grade 2 or higher. In some embodiments, prophylactic use of tocilizumab administered on day 2 may reduce the rate of CRS of grade 3 or higher.

[0178] Any corticosteroid may be suitable for this use. In one embodiment, the corticosteroid is dexamethasone. In some embodiments, the corticosteroid is methylprednisolone. In some embodiments, a combination of the two is administered. In some embodiments, the glucocorticoids include synthetic and non-synthetic glucocorticoids. Exemplary glucocorticoids include alclomethasone, alginate, beclomethasone (e.g., beclomethasone dipropionate), betamethasone (e.g., betamethasone 17-valerate, betamethasone sodium acetate, betamethasone sodium phosphate, betamethasone valerate), budesonide, clobetasol (e.g., clobetasol propionate), clobetasone, clocortolone (e.g., clocortolone pivalate), Cloprednol, corticosterone, cortisone and hydrocortisone (e.g., hydrocortisone acetate), cortivazol, deflazacor, desonide, desoximetasone, dexamethasone (e.g., dexamethasone 21 phosphate, dexamethasone acetate, dexamethasone sodium phosphate), diflorasone (e.g., diflorasone diacetate), diflucortralone, difluprednate, enoxolone, furazacor, fluclonide, fludrocorticoid fludrocortisone (e.g., fludrocortisone acetate), flumethasone (e.g., flumethasone pivalate), flunisolide, fluocinolone (e.g., fluocinolone acetonide), fluocinonide, flucortine, flutrolone, fluorometholone (e.g., fluorometholone acetate), fluperolone (e.g., fluperone acetate), fluprednidene, fluprednisolone, flurandrenolide, fluticasone (e.g., fluticasone propionate), formocortal, Halcinonide, halobetasol, halometasone, halopredone, hydrocortamate, hydrocortisone (e.g., hydrocortisone 21-butyrate, hydrocortisone aceponate, hydrocortisone acetate, hydrocortisone buteprate, hydrocortisone butyrate, hydrocortisone cypionate, hydrocortisone hemisuccinate, hydrocortisone probutate, hydrocortisone sodium phosphate, hydrocortisone sodium succinate,hydrocortisone valerate), loteprednol etabonate, mazipredone, medrysone, meprednisone, methylprednisolone (methylprednisolone aceponate, methylprednisolone acetate, methylprednisolone hemisuccinate, methylprednisolone sodium succinate), mometasone (e.g., mometasone furoate), paramethasone (e.g., paramethasone acetate), prednicarbate, prednisolone (e.g., prednisolone 25-diethylaminoacetate, prednisolone sodium phosphate, prednisolone 21-hemisuccinate, prednisolone acetate; prednisolone furoate These glucocorticoids and their salts include, but are not limited to, prednisolone hemisuccinate, prednisolone-21 (beta-D-glucuronide), prednisolone metasulfobenzoate, prednisolone stearate, prednisolone tebutate, prednisolone tetrahydrophthalate), prednisone, prednival, prednylidene, rimexolone, tixocortol, triamcinolone (e.g., triamcinolone acetonide, triamcinolone benetonide, triamcinolone hexacetonide, triamcinolone acetonide 21 palmitate, triamcinolone diacetate). These glucocorticoids and their salts are described, for example, in Rem, Remington's Pharmaceutical Sciences, A. Osol, ed., Mack Pub. Co., Easton, Pa. (16th ed. 1980), and Remington: The Science and Practice of Pharmacy, 22nd Edition, Lippincott Williams & Wilkins, Philadelphia, Pa. (2013), and any other editions, which are incorporated herein by reference. In some embodiments, the glucocorticoid is selected from among cortisone, dexamethasone, hydrocortisone, methylprednisolone, prednisone, and prednisone. In one embodiment, the glucocorticoid is dexamethasone. In other embodiments, the steroid is a mineralocorticoid. Any other steroid may be used in the methods provided herein.

[0179] The one or more corticosteroids may be administered at any dose and frequency of administration that may be compatible with the severity / grade of the adverse event (e.g., CRS and NE). Tables 1 and 2 provide examples of dosing regimens for managing CRS and NE, respectively. In another embodiment, administration of the corticosteroid comprises dexamethasone 10 mg orally or IV 1-4 times per day. Another embodiment, sometimes referred to as a "high-dose" corticosteroid, comprises methylprednisone 1 g / day IV administered alone or in combination with dexamethasone. In some embodiments, the one or more corticosteroids are administered at a dose of 1-2 mg / kg per day.

[0180] The corticosteroid can be administered in any amount effective to ameliorate one or more symptoms associated with CRS or an adverse event, such as neurotoxicity. Corticosteroids, e.g., glucocorticoids, can be administered in an amount of about 0.1 to 100 mg, 0.1 to 80 mg, 0.1 to 60 mg, 0.1 to 40 mg, 0.1 to 30 mg, 0.1 to 20 mg, 0.1 to 15 mg, 0.1 to 10 mg, 0.1 to 5 mg, 0.2 to 40 mg, 0.2 to 30 mg, 0.2 to 20 mg, 0.2 to 15 mg, 0.2 to 10 mg, 0.2 to 5 mg, 0.4 to 40 mg, 0.4 to 30 mg, 0.4 to 20 mg, 0.4 to 15 mg, 0.4 to 10 mg, 0.4 to 5 mg, 0.4 to 4 mg, 1 to 20 mg, 1 to 15 mg, or 1 to 10 mg per dose to a 70 kg adult subject, for example. Typically, corticosteroids, such as glucocorticoids, are administered to an average adult human subject in an amount of about 0.4 to 20 mg per dose, e.g., about 0.4 mg, 0.5 mg, 0.6 mg, 0.7 mg, 0.75 mg, 0.8 mg, 0.9 mg, 1 mg, 2 mg, 3 mg, 4 mg, 5 mg, 6 mg, 7 mg, 8 mg, 9 mg, 10 mg, 11 mg, 12 mg, 13 mg, 14 mg, 15 mg, 16 mg, 17 mg, 18 mg, 19 mg, or 20 mg.

[0181] In some embodiments, the corticosteroid is administered at a dose of, for example, 0.001 or about 0.001 mg / kg (of subject), 0.002 mg / kg, 0.003 mg / kg, 0.004 mg / kg, 0.005 mg / kg, 0.006 mg / kg, 0.007 mg / kg, 0.008 mg / kg, 0.009 mg / kg, 0.01 mg / kg, 0.015 mg / kg, 0.02 mg / kg, 0.025 mg / kg, 0.03 mg / kg, 0.035 mg / kg, 0.04 mg / kg, 0.045 mg / kg, 0.05 mg / kg, 0.055 mg / kg, 0.05 mg / kg, 0.06 mg / kg, 0.07 mg / kg, 0.08 mg / kg, 0.09 mg / kg, 0.01 mg / kg, 0.015 mg / kg, 0.02 mg / kg, 0.025 mg / kg, 0.03 mg / kg, 0.035 mg / kg, 0.04 mg / kg, 0.045 mg / kg, 0.0 ...7 mg / .06mg / kg, 0.065mg / kg, 0.07mg / kg, 0.075mg / kg, 0.08mg / kg, 0.085mg / kg, 0.09mg / kg, 0.095mg / kg, 0.1mg / kg, 0.15mg / kg, 0.2mg / kg, 0.25mg / kg, 0.30mg / kg, 0.35mg / kg , 0.40mg / kg, 0.45mg / kg, 0.50mg / kg, 0.55mg / kg, 0.60mg / kg, 0.65mg / kg, 0.70mg / kg, 0.75mg / kg, 0.80mg / kg, 0.85mg / kg, 0.90mg / kg, 0.95mg / kg, 1mg / kg, 1.05mg / kg, 1. It may be administered at dosages of 1 mg / kg, 1.15 mg / kg, 1.20 mg / kg, 1.25 mg / kg, 1.3 mg / kg, 1.35 mg / kg, or 1.4 mg / kg.

[0182] Generally, the dose of corticosteroid administered depends on the individual corticosteroid, since there are differences in potency between different corticosteroids. Typically, drugs vary in potency, so it is understood that doses to achieve equivalent effect may differ. Equivalents in terms of potency for various glucocorticoids and routes of administration are well known. Information regarding equivalent steroid dosing (in a non-chronotherapeutic mode) can be found in the British National Formulary (BNF) 37 (March 1999).

[0183] In some embodiments, any of these embodiments are performed with autologous anti-CD19 CAR T cell therapy, with axicabtagene ciloleucel prepared as described in International Application PCT / US2016 / 057983, with or without an AKT inhibitor. In some embodiments, the therapy is used to treat ALL patients. In other embodiments, the therapy is used to treat any other cancer, as described elsewhere herein. In some embodiments, the patient is an adult patient with relapsed / refractory acute lymphoblastic leukemia (ALL). In some embodiments, the patient has undergone conditioning therapy. In some embodiments, the conditioning therapy is cyclophosphamide 500 mg / m on days -4, -3, and -2 of treatment (D0) for 3 days. 2 and fludarabine 30 mg / m 2 In some embodiments, these patients receive 0.5×10 6 , 1×10 6 Or 2 x 10 6 Axicabtagene ciloleucel will be administered at CAR+ cells / kg.

[0184] In some of these and other embodiments, the patient is administered steroids in grade 2 NE. In some embodiments, the patient is administered steroids in grade 3 NE. In some of these embodiments, the patient is administered tocilizumab alone or in combination with steroids in the setting of CRS. In some other embodiments, the patient is administered an anti-IL6 antibody (e.g., siltuximab) to manage CRS. In some other embodiments, the patient is administered an anti-IL6 antibody (e.g., siltuximab) to manage neurotoxicity (e.g., neurotoxic events). In some embodiments, the patient is refractory to anti-IL6 therapy. In some embodiments, the patient is administered siltuximab as a third line treatment for CRS after failure of both tocilizumab and corticosteroid therapy.

[0185] Neutralizing or reducing the CSF / CSFR1 axis to manage adverse effects, safety, and neurotoxicity In some embodiments, axicabtagene ciloleucel treatment is combined with an inhibitor or antagonist of a member of the colony-stimulating factor (CSF) protein family or their receptors (e.g., CSF1R, CSF2R) to manage adverse effects, safety, and / or neurotoxicity. These treatments can be used in any context, including but not limited to prophylaxis. They can also be used in combination with steroids. Inhibitors or antagonists used in combination treatments can be, but are not limited to, antibodies, neutralizing agents expressed in CAR-T cells, small molecules, and other agents. In some embodiments, other CD19-directed genetically modified autologous T cell immunotherapies (e.g., axicabtagene ciloleucel; other examples of such immunotherapies are described elsewhere herein) are combined with inhibitors or antagonists of a member of the colony-stimulating factor (CSF) protein family or their receptors (e.g., CSF1R, CSF2R).

[0186] In one embodiment, the CSF family member is GM-CSF (granulocyte-macrophage colony-stimulating factor, also known as CSF2). GM-CSF is a It can be produced by several hematopoietic and non-hematopoietic cell types and can activate / "prime" bone marrow populations to produce inflammatory mediators such as TNF and interleukin 1β (IL1β). In some embodiments, the GM-CSF inhibitor is an antibody that binds to and neutralizes circulating GM-CSF. In some embodiments, the antibody is selected from lenzilumab, namilumab (AMG203), GSK3196165 / MOR103 / Otilimab (GSK / MorphoSys), KB002 and KB003 (KaloBios), MT203 (Micromet and Nycomed), and MORAb-022 / gimsilumab (Morphotek). In some embodiments, the antibody is a biosimilar thereof. In some embodiments, the antagonist is E21R, a modified form of GM-CSF that antagonizes the function of GM-CSF. In some embodiments, the inhibitor / antagonist is a small molecule.

[0187] In one embodiment, the CSF family member is M-CSF (also known as macrophage colony stimulating factor or CSF1). Non-limiting examples of agents that inhibit or antagonize CSF1 include small molecules, antibodies, chimeric antigen receptors, fusion proteins, and other agents. In one embodiment, the CSF1 inhibitor or antagonist is an anti-CSF1 antibody. In one embodiment, the anti-CSF1 antibody is selected from those made by Roche (e.g., RG7155), Pfizer (PD-0360324), Novartis (MCS110 / Lanotuzumab), or a biosimilar version of any one of them.

[0188] In some embodiments, the inhibitor or antagonist inactivates the activity of either the GM-CSF-R-alpha (also known as CSF2R) or CSF1R receptor. In some embodiments, the inhibitor is selected from mavrilimumab (formerly CAM-3001), a fully human GM-CSF receptor alpha monoclonal antibody currently in development by MedImmune, Inc.; cabilalizumab (Five Prime Therapeutics); LY3022855 (IMC-CS4) (Eli Lilly), Emactuzumab, also known as RG7155 or RO5509554; FPA008, a humanized mAb (Five Prime / BMS); AMG820 (Amgen); ARRY-382 (Array Biopharma); MCS110 (Novartis); PLX3397 (Plexxikon); ELB041 / AFS98 / TG3003 (ElsaLys Bio, Transgene), SNDX-6352 (Syndax). In some embodiments, the inhibitor or antagonist is expressed in CAR-T cells. In some embodiments, the inhibitor is a small molecule (e.g., heteroaryl amides, quinolinones, pyrido-pyrimidos); BLZ945 (Novartis), PLX7486, ARRY-382, pexidiltinib (also known as PLX3397), or 5-((5-chloro-1H-pyrrolo[2,3-b]pyridin-3-yl)methyl)-N-06-(trifluoromethyl)pyridin-3-yl)methyl)pyridin-2-amine; GW 2580 (CAS 870483-87-7), KI20227 (CAS 623142-96-1), AC708 by Ambit Siosciences, or Cannarile et al., Journal for Immuno Therapy of Cancer (2017) 5:53 and U.S. Patent Application Publication No. 20180371093, which are incorporated by reference herein for the inhibitors they disclose.Additional neutralizing antibodies against GM-CSF or its receptor are described, for example, in "GM-CSF as a target in inflammatory / autoimmune disease: current evidence and future therapeutic potential" Hamilton, JA Expert. Rev. Clin. Immunol., 2015; and "Targeting GM-CSF in inflammatory diseases" Wicks, IP, Roberts, AW Nat. Rev. Rheumatol., 2016 It is described in the art.

[0189] The inhibitors or combinations of inhibitors of GM-CSF, CSF1, GM-CSFR or CSF1R may be administered in different dosages. Those skilled in the art can routinely determine their effective amounts and timing of administration. In one embodiment, mavrilimumab and / or lenzilumab may be used in combination with treatment with CD19-directed genetically modified autologous T-cell immunotherapy, which in one example may be axicabtagene ciloleucel. Other examples of such immunotherapy are described elsewhere herein.

[0190] In one embodiment, mavrilimumab is administered SQ, IV, or OP / IP on D0 of CAR-T cell treatment. In one embodiment, a single dose of 100 mg or 150 mg is administered SQ or IP. Mavrilimumab may be administered before, simultaneously with, or after axicabtagene ciloleucel. In one embodiment, both drugs are administered on day 0 (D0) of CAR-T cell treatment. In one embodiment, mavrilimumab is administered in a single dose of 30 mg. In one embodiment, mavrilimumab is administered in a single dose of 50 mg. In some embodiments, mavrilimumab is administered in a dose of 10-500 mg, 100-200 mg, 30-100 mg, or 30-150 mg. In one embodiment, mavrilimumab is administered at 1 mg / kg body weight, 2 mg / kg, 3 mg / kg, 4 mg / kg, 5 mg / kg, 6 mg / kg, 7 mg / kg, 8 mg / kg, 9 mg / kg, 10 mg / kg, 11 mg / kg, 12 mg / kg, 13 mg / kg, 14 mg / kg, 15 mg / kg, 16 mg / kg, 17 mg / kg, 18 mg / kg, 19 mg / kg, 20 mg / kg, 21 mg / kg, 22 mg / kg, 23 mg / kg, 24 mg / kg, 25 mg / kg, 26 mg / kg, 27 mg / kg, 28 mg / kg, 29 mg / kg, or 30 mg / kg subcutaneously, OP / IP, or IV, preferably once on day 0. In one embodiment, mavrilimumab is administered once at 3 mg / kg body weight, preferably on day 0, and mavrilimumab can be administered in a single dose or multiple doses (e.g., weekly, biweekly). In one embodiment, mavrilimumab is administered SQ, IV, or IP as a single dose of 0.01-10.0 mg / kg.

[0191] In one embodiment, lenzilumab is administered by IP infusion at 10 mg / kg (up to 600 mg), 20 mg / kg (up to 1200 mg), or 30 mg / kg (up to 1800 mg). Lenzilumab may be administered before, simultaneously, or after axicabtagene ciloleucel. In one embodiment, both drugs are administered on D0. In one embodiment, lenzilumab is administered as a single dose of 0.01-50.0 mg / kg. In one embodiment, the single dose is 1, 3, 10, 50, or 100 mg / kg. In one embodiment, the single dose is 200 mg, 400 mg, or 600 mg. In one embodiment, lenzilumab is administered in weekly doses of 10, 50, or 100 mg / kg.

[0192] In one embodiment, an inhibitor or antagonist of GM-CSF, M-CSF, GM-CSF-R-alpha (also known as CSF2R) or CSF1R receptor (e.g., mavrilimumab or lenzilumab) can be administered any day between day -5 and day 5 relative to the initiation of treatment with CD19-directed genetically modified autologous T cell immunotherapy (D0). In one embodiment, the inhibitor is administered in one or more daily doses any day between day -2 and day 10. In one embodiment, the inhibitor is administered one week or more after immunotherapy has begun (D0).

[0193] Without wishing to be bound by any theory, it is hypothesized that inhibition of the GM-CSF receptor (e.g., mavrilimumab) or BM-CSF (e.g., lenzilumab) may neutralize the production of several cytokines from myeloid cells or compartments, thus reducing the degree of neuroinflammation. Under such circumstances, the incidence of grade 1, 2, 3 or higher neurological or adverse events may be reduced, which may be observed or measured by several parameters, such as the level of GM-CSF suppression or inhibition, the level of CAR T cells, and the level of cytokines in the blood.

[0194] In some embodiments, axicabtagene ciloleucel treatment is combined with treatment with an anti-CD20 agent. In some embodiments, the agent is rituximab. In some embodiments, the agent is an antibody selected from Arzera, Gazyva, ibritumomab tiuxetan, obinutuzumab, ofatumumab, ocrelizumab, veltuzumab, Rituxan Hycera, Rituxan, Bexxar, Zevalin, and biosimilars thereof. These antibodies are described in JML Casan, J. Wong, MJ Northcott, and S. Opat (2018) Anti-CD20 monoclonal antibodies: reviewing a revolution, Hum Vaccin Immunother. 2018;14(12):2820-2841, and FHDu, EA Mills, Y. Mao-Draayer (2017) Next-generation anti-CD20 monoclonal antibodies in autoimmune disease treatment. Autoimmun Highlights. 2017;8:1-12. doi:10.1007 / s13317-017-0100-y, both of which are incorporated by reference in their entirety, particularly with respect to exemplary dosing regimens and cancers within the scope of this disclosure.

[0195] Dosage form and strength In some embodiments, the CD19-directed genetically modified autologous T cell immunotherapy is available as a cell suspension for infusion. In some embodiments, a single dose of CD19-directed genetically modified autologous T cell immunotherapy is approximately 1×10 per kg of body weight in approximately 68 mL of suspension in an infusion bag. 6 ~Approx. 2×10 6 Viable CAR-positive T cells (or up to 2 × 10 for patients weighing ≥ 100 kg) 8In some embodiments, the CD19-directed genetically modified autologous T cell immunotherapy comprises a target dose of CAR-positive viable T cells). In some embodiments, the CD19-directed genetically modified autologous T cell immunotherapy is axicabtagene ciloleucel (YESCARTA®). In some embodiments, a single dose of the CD19-directed genetically modified autologous T cell immunotherapy is present in the container. Such containers can be sterile. In some embodiments, the container is an infusion bag. In some embodiments, the volume of the infusion bag is about 100 mL, 150 mL, 200 mL, 250 mL, 300 mL, 500 mL, 750 mL, 1,000 mL, 1,500 mL, 2,000 mL, or 3,000 mL.

[0196] Risk Evaluation and Mitigation Strategy (REMS) Due to the risk of CRS and neurotoxicity, in some embodiments, CD19-directed genetically modified autologous T cell immunotherapy may be available through a restricted program under a Risk Evaluation Mitigation Strategy (REMS). Exemplary components of a REMS are as follows: Healthcare facilities that dispense and administer CD19-directed genetically modified autologous T-cell immunotherapies must be registered and must comply with REMS requirements. Qualified medical facilities must have tocilizumab immediately available on-site and must ensure that, if needed for the treatment of CRS, a minimum of two doses of tocilizumab are available for each patient to be infused within two hours after the infusion of the CD19-directed genetically modified autologous T-cell immunotherapy. Qualified medical facilities must ensure that health care providers who prescribe, dispense or administer CD19-directed genetically modified autologous T-cell immunotherapy are trained in the management of CRS and neurotoxicity.

[0197] Cytokine release syndrome (CRS) In some embodiments, the medical facility verifies that two doses of tocilizumab are available prior to infusion of the CD19-directed genetically modified autologous T cell immunotherapy. In some embodiments, the medical facility verifies that four doses of tocilizumab are available prior to infusion of the CD19-directed genetically modified autologous T cell immunotherapy. In some embodiments, the method comprises monitoring the patient daily for at least seven days at a certified medical facility after infusion for signs and symptoms of CRS. In some embodiments, the method comprises monitoring the patient daily for at least seven to ten days at a certified medical facility after infusion for signs and symptoms of CRS. In some embodiments, the method comprises monitoring the patient daily for at least eight days at a certified medical facility after infusion for signs and symptoms of CRS. In some embodiments, the method comprises monitoring the patient daily for at least nine days at a certified medical facility after infusion for signs and symptoms of CRS. In some embodiments, the method comprises monitoring the patient daily for at least ten days at a certified medical facility after infusion for signs and symptoms of CRS. In some embodiments, the method includes monitoring the patient for signs or symptoms of CRS for 4 weeks after infusion. In some embodiments, the method includes advising the patient to seek immediate medical attention whenever signs or symptoms of CRS occur. In some embodiments, the method includes initiating treatment with supportive care, tocilizumab, or tocilizumab and corticosteroids as indicated at the first sign of CRS. Examples of supportive care can be found in the following references, which are incorporated herein by reference in their entirety: Jennifer N. Brudno, "Clinical Risk Assessment and Treatment of CRS," vol. 14, no. 1, pp. 1111-1115, 2002; ... and James N. Kochenderfer(2016),Toxicities of chimeric antigen receptor T cells:recognition and management,Blood 2016 127:3321-3330;Lara L Riegler,Gavin P Jones,and Daniel W Lee(2019)Current approaches in the grading and management of cytokine release syndrome after chimeric antigen receptor T-cell therapy,Ther Clin Risk Manag.2019;15:323-335; and Bradley D Hunter Caron A Jacobson(2019)CAR T-Cell Associated Neurotoxicity:Mechanisms,Clinicopathologic Correlates,and Future Directions,JNCI:Journal of the National Cancer Such references, such as those summarized in the Journal of the American Chemical Society, can be found by those skilled in the art.

[0198] Neurotoxicity In some embodiments, the method includes monitoring the patient daily for at least 7 days at a certified medical facility after infusion for signs and symptoms of neurotoxicity. In some embodiments, the method includes monitoring the patient daily for at least 7-10 days at a certified medical facility after infusion for signs and symptoms of CRS. In some embodiments, the method includes monitoring the patient daily for at least 10 days at a certified medical facility after infusion for signs and symptoms of CRS. In some embodiments, the method includes monitoring the patient for 4 weeks after infusion for signs or symptoms of neurotoxicity and treating promptly.

[0199] Hypersensitivity reactions Allergic reactions may occur following infusion of CD19-directed genetically modified autologous T-cell immunotherapy. In some embodiments, severe hypersensitivity reactions, including anaphylaxis, may be due to dimethylsulfoxide (DMSO) or residual gentamicin in the CD19-directed genetically modified autologous T-cell immunotherapy.

[0200] Viral reactivation In some embodiments, reactivation of Hepatitis B Virus (HBV) may occur in patients treated with drugs directed against B cells, which may potentially result in fulminant hepatitis, liver failure and death. In some embodiments, the method includes screening for HBV, HCV and HIV according to clinical guidelines prior to harvesting the cells for manufacturing.

[0201] Prolonged cytopenias In some embodiments, patients may exhibit cytopenias for several weeks following lymphodepleting chemotherapy and infusion of CD19-directed genetically modified autologous T-cell immunotherapy. In some embodiments, the method includes monitoring blood counts following infusion of CD19-directed genetically modified autologous T-cell immunotherapy.

[0202] Hypogammaglobulinemia In some embodiments, B cell aplasia and hypogammaglobulinemia may occur in patients undergoing treatment with CD19-directed genetically modified autologous T cell immunotherapy. In some embodiments, the methods include monitoring immunoglobulin levels following treatment with CD19-directed genetically modified autologous T cell immunotherapy and managing using infection precautions, antibiotic prophylaxis, and immunoglobulin replacement.

[0203] In some embodiments, vaccination with live viral vaccines is not recommended for at least 6 weeks prior to initiation of lymphodepleting chemotherapy, during CD19-directed genetically modified autologous T cell immunotherapy treatment, and until immune recovery following CD19-directed genetically modified autologous T cell immunotherapy treatment. Secondary malignancies In some embodiments, patients treated with CD19-directed genetically modified autologous T cell immunotherapy may develop secondary malignancies. In some embodiments, the method includes lifelong monitoring for secondary malignancies.

[0204] Tumor lysis syndrome (TLS) Patients treated with CD19-directed genetically modified autologous T cell immunotherapy may develop TLS, which may be severe.To minimize the risk of TLS, in some embodiments, the method includes evaluating patients for elevated uric acid or high tumor burden, and administering allopurinol or alternative prophylaxis before injecting axicabtagene ciloleucel.The signs and symptoms of TLS should be monitored, and events should be managed according to standard guidelines.

[0205] Effects on ability to drive and use machinery Due to the possibility of neurological events such as altered mental status or seizures, patients receiving CD19-directed genetically modified autologous T-cell immunotherapy are at risk for altered or decreased consciousness or coordination for 8 weeks following infusion of the CD19-directed genetically modified autologous T-cell immunotherapy. In some embodiments, the methods include advising the patient to refrain from driving and engaging in hazardous occupations or activities, such as operating heavy or potentially dangerous machinery, during this initial period.

[0206] Storage and handling In some embodiments, the CD19-directed genetically modified autologous T cell immunotherapy is provided in an infusion bag containing about 68 mL of a frozen suspension of genetically modified autologous T cells in 5% DMSO and 2.5% albumin (human). In some embodiments, the CD19-directed genetically modified autologous T cell immunotherapy is provided in about 68 mL of a frozen suspension of genetically modified autologous T cells in 5% DMSO and 2.5% albumin (human) (NDC In some embodiments, the CD19-directed genetically modified autologous T-cell immunotherapy includes Cryostar CS10. In some embodiments, the CD19-directed genetically modified autologous T-cell immunotherapy includes 300 mg sodium per infusion. In some embodiments, the CD19-directed genetically modified autologous T-cell immunotherapy includes an infusion bag containing about 50-100 mL, 50-90 mL, 50-80 mL, 50-70 mL, 60-70 mL, 60-75 mL, or 65-75 mL of a suspension of genetically modified autologous T-cells in 5% DMSO and 2.5% albumin (human). In some embodiments, the CD19-directed genetically modified autologous T cell immunotherapy is provided in an infusion bag containing less than 100 mL, less than 90 mL, less than 80 mL, less than 70 mL, less than 70 mL, less than 72 mL, or less than 75 mL of a suspension of genetically modified autologous T cells in 5% DMSO and 2.5% albumin (human). In some embodiments, the CD19-directed genetically modified autologous T cell immunotherapy is provided in an infusion bag containing more than 50 mL, more than 60 mL, more than 65 mL, more than 66 mL, more than 67 mL, or more than 68 mL of a suspension of genetically modified autologous T cells in 5% DMSO and 2.5% albumin (human). In some embodiments, the suspension is frozen.

[0207] In some embodiments, the CD19-directed genetically modified autologous T cell immunotherapy infusion bag is supplied as an ethylene-vinyl acetate cryopreservation bag containing approximately 68 mL of cell dispersion with an additional sealed tube and two available spike ports.

[0208] In some embodiments, the CD19-directed genetically modified autologous T-cell immunotherapy infusion bag is individually packaged in a metal cassette. In some embodiments, the CD19-directed genetically modified autologous T-cell immunotherapy infusion bag is individually packaged in a metal cassette (NDC 71287-119-02). In some embodiments, the CD19-directed genetically modified autologous T-cell immunotherapy infusion bag is stored in the vapor phase of liquid nitrogen. In some embodiments, the CD19-directed genetically modified autologous T-cell immunotherapy infusion bag is supplied as a liquid nitrogen dry shipper.

[0209] In some embodiments, the method includes matching the patient's identity with the patient identifiers on the cassette and infusion bag upon receipt. In some embodiments, the CD19-directed genetically modified autologous T-cell immunotherapy is frozen and stored in the vapor phase of liquid nitrogen (minus 150° C. or below). In some embodiments, the CD19-directed genetically modified autologous T-cell immunotherapy is thawed prior to use.

[0210] CAR-T cell therapy may be associated with various adverse events, for example, cerebral edema may be detected in the treatment with CD19 CAR-T described herein. In one aspect, the present application provides a method of recovering from cerebral edema after CD19 CAR-T therapy, comprising administering an immunosuppressant drug that depletes T lymphocytes, such as antithymocyte globulin (ATG). In one embodiment, ATG is administered IV at 2 mg / kg / day. In one embodiment, ATG is administered with methylprednisolone and / or tocilizumab to reduce systemic cytokine storm. In one embodiment, ATG is rabbit ATG or equine ATG. In another embodiment, ATG may be administered in one or more doses.

[0211] The following non-limiting examples and data illustrate various aspects and features of the disclosed cell and therapeutic methods and uses. In some embodiments, use of the methods and compounds of the present invention provides surprising, unexpected and contrary results and data. The utility of the disclosed methods is illustrated by the use of several cells or compounds that can be used therein, but one of skill in the art will appreciate that various other cells or compounds can be used therein, provided they are within the scope of the present disclosure. It will be understood that comparable results can be obtained using combinations of the above. In the context of these examples, ASCT means autologous stem cell transplant, DLBCL means diffuse large B-cell lymphoma, ECOG means Eastern Cooperative Oncology Group performance status, IPI means International Prognostic Index, PD means progressive disease, PMBCL means primary mediastinal B-cell lymphoma, SPD means sum of two-way products; TFL means transformed follicular lymphoma. EXAMPLES

[0212] Example 1 Leukapheresis is a procedure to collect peripheral blood mononuclear cells (PBMCs). In this example, subjects or patients with relapsed or refractory aggressive B-cell non-Hodgkin's lymphoma undergo leukapheresis to obtain T cells for the production of axicabtagene ciloleucel. Eligible patients may have disease resistant to their most recent treatment or may have relapsed within one year after autologous hematopoietic stem cell transplantation (HSCT).

[0213] After leukapheresis, patients received a lymphodepleting regimen, which consisted of cyclophosphamide 500 mg / m 2 IV and fludarabine 30 mg / m 2 IV administration of 2×10 6 Dose of CAR-positive viable T cells / kg (maximum tolerated dose: 2 × 10 8The subject may be administered by IV infusion on days 5, 4, and 3 prior to receiving axicabtagene ciloleucel with 100 mg / kg of body weight (cells) (day 0). Additionally, the subject may be administered mavrilimumab or lenzilumab. Mavrilimumab may be administered at a dose of 100 mg or 150 mg. Mavrilimumab may be administered SQ, OP / IP, or IV once on day 0 at 3 mg / kg body weight. Lenzilumab may be administered at a dose of 10 mg / kg (up to 600 mg), 20 mg / kg (up to 1200 mg), or 30 mg / kg (up to 1800 mg). Mavrilimumab and lenzilumab may be administered on day 0 prior to axicabtagene ciloleucel, simultaneously with axicabtagene ciloleucel, or after receiving axicabtagene ciloleucel.

[0214] Neurological assessments may be performed on days 0, 1, and every other day over the 7-day observation period. Common neurotoxicities may include encephalopathy, headache, tremors, dizziness, aphasia, delirium, insomnia, and anxiety.

[0215] The grading criteria can be the NCI Common Terminology Criteria for Adverse Events (CTCAE) version 5.0 or other guidelines. To assess whether a subject exhibits reduced or regulated neurological events, several parameters can be examined at different time points, including levels of cytokines, GS-CSF, CD19 CAR-T cells, anti-tumor effects, and / or cytokine / chemokine milieu, for example, every month for 28 days or for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 years.

[0216] Without being bound by any theory, it is hypothesized that inhibition of GM-CSF receptors (such as mavrilimumab) or BM-CSF (such as lenzilumab) may neutralize the production of several cytokines from myeloid cells or compartments, thus reducing the degree of neuroinflammation. Under such circumstances, the incidence of grade 1, 2, 3 or higher neurological or adverse events may be reduced. This may be observed or measured by several parameters, such as the level of GM-CSF suppression or inhibition, the level of CAR T cells, and the level of cytokines in the blood.

[0217] Additionally, PK and PD analyses can be performed on blood (anti-CD19 CAR T cell levels) or serum (cytokines) as potential predictive markers of efficacy and safety of axicabtagene ciloleucel. Cytokines and chemokines include those involved in homeostasis, pro-inflammatory and immunomodulatory cytokines IL-2, IL-6, IL-10, IL-12p40 / p70, IL-15, IL-17a, TNF-α, IFN-γ, and GM-CSF; acute phase reactants such as CRP; chemokines IL-8, MCP-1, and MIP-1α, and IP-10; and HLH-associated markers such as ferritin and IL-2Rα.

[0218] Efficacy is established based on complete response (CR) rate and duration of response (DOR) as determined by an independent review committee. Efficacy can be determined or measured using parameters such as objective response rate (ORR), complete response rate, partial response rate, and / or duration of response. ORR can be determined by the revised International Working Group criteria of 2007 as assessed by an independent review committee or other similar guidelines. Response duration parameters can include number of responders, DOR (months) (median, 95% CI, range), DOR if best response is CR and DOR if best response is PR (months, median, 95% CI, range), and median follow-up (months) for DOR. CR, complete response; DOR, duration of response; NE, not estimable; PR, partial response. Among all responders, DOR is measured from the date of first objective response to the date of progression or death due to relapse or toxicity. Some patient selection parameters include: 1. Relapsed or refractory large B-cell lymphoma after 2 or more lines of systemic therapy, including histologically confirmed diffuse large B-cell lymphoma (DLBCL) not otherwise specified, primary mediastinal large B-cell lymphoma, high-grade B-cell lymphoma, and DLBCL arising from follicular lymphoma. 2. Subject must have received sufficient prior therapy, including at least: Anti-CD20 monoclonal antibody (unless the investigator determines the tumor is CD20 negative), and · Anthracycline-containing chemotherapy regimens; 3. At least one measurable lesion according to the Lugano response criteria for malignant lymphoma (Cheson et al., 2014). 4. No known or suspected central nervous system involvement by lymphoma 5. ECOG performance status 0 or 1 6. Adequate bone marrow function as evidenced by: ANC≧1000 / uL ·Platelets ≧75,000 / uL Absolute lymphocyte count ≥ 100 / uL 7. Adequate renal, hepatic, cardiac and pulmonary function as evidenced by: Creatinine clearance (Cockcroft Gault) ≧60mL / min ·Serum ALT / AST≦2.5ULN Total bilirubin ≦1.5mg / dL (excluding subjects with Gilbert syndrome) Cardiac ejection fraction ≥ 50% with no evidence of clinically significant pericardial effusion as determined by ECHO and no clinically significant ECG findings No clinically significant pleural effusion Baseline oxygen saturation >92% on room air

[0219] Example 2 This study included patients with R / R LBCL (including DLBCL, PMBCL, TFL, and high-grade B-cell lymphoma) who had been treated with ≥2 prior lines of therapy. Minimal prior therapy included anti-CD20 antibodies or anthracycline-containing chemotherapy. Patients underwent leukapheresis and received conditioning chemotherapy with cyclophosphamide 500 mg / m 2 and fludarabine 30 mg / m 2 on days -4, -3, and -2, followed by 2 x 10 6 Patients with high disease burden at screening or baseline received bridging therapy (between leukapheresis and conditioning chemotherapy), which included dexamethasone (20–40 mg / kg). mg / day or equivalent for 1-4 days) in combination with rituximab (375 mg / m weekly for 3 weeks). 2 ), or rituximab (375 mg / m 2 for 1 day) combined with methylprednisolone (1 g / m2 for 1–3 days) 2 ) or bendamustine (90 mg / m2 for 2 days 2) and both bendamustine and rituximab were started on the same day. The study applied modified adverse event management (or modified AE management), in which patients showing grade 1 or higher CRS (cytokine release syndrome) received either tocilizumab or corticosteroids, patients showing grade 1 or higher NE (neurological events) received corticosteroids, and patients showing grade 2 or higher NE (neurological events) received either tocilizumab or corticosteroids. Corticosteroids included oral or IV dexamethasone 10 mg 1 to 4 times daily or IV methylprednisone 1 g daily. Tocilizumab was not administered prophylactically. The primary endpoints were the incidence and severity of CRS and NE. It was hypothesized that early use or intervention such as corticosteroids in less severe cases (e.g., grade 1 CRS or NE) could facilitate management of severe CRS and NE and potentially reduce such incidence without affecting response rates. Sixty-seven percent of patients received bridging therapy (therapy between leukapheresis and conditioning chemotherapy). All patients had evidence of disease after bridging therapy documented by baseline positron emission tomography / computed tomography scan (PET-CT). Eighty-six percent of patients had DLBCL (diffuse large B-cell lymphoma) and 14% had TFL (transformed follicular lymphoma). Median age was 63 years (y) (range, 36-73), and 67% were male. Eighty-one percent had stage III-IV disease. Seventy-six percent were R / R to second-line or later therapy, and 10% relapsed after autologous stem cell transplant. Sixty-seven percent had received three or more prior lines of therapy. The two-way sum of product SPD (range) tumor burden was 1915 (360-11,487). Grade ≥3 AEs were observed in 95% of patients. The most common grade ≥3 AEs were decreased neutrophil count (33%), anemia (29%), neutropenia (29%), and pyrexia (24%). Grade 1 or 2 NE was observed in 48% of patients, and grade ≥3 NE was observed in 10% of patients. Grade ≥3 NE symptoms included somnolence (10%), confusion (10%), encephalopathy (5%), and aphasia (5%).No grade 4 or higher NE was observed. Grade 1 CRS was observed in 33% of patients, and grade 2 CRS in 67% of patients. No patients had grade 3 or higher CRS. At a median follow-up of 7.7 months, the ORR was 81%, with 62% CR and 19% partial response (PR). 5% of patients had stable disease (SD) and 14% progressive disease (PD). In the previous study, patients received the same treatment and different AE management, in which patients with grade 2 or higher CRS and patients with grade 2 or higher NE received either tocilizumab or corticosteroids (26% of patients received steroids and 43% received tocilizumab). Results of 2 years of follow-up showed that the objective response rate (ORR) was 83%, the complete response (CR) rate was 58%, 11% of patients had grade 3 or higher CRS, and 32% of patients had NE. Taken together, the above results suggested that early use of corticosteroids did not affect treatment success and reduced the incidence of severe CRS and / or NE.

[0220] Example 3 In this study, patients with R / RB cell ALL, >5% BM (bone marrow) blasts, and ECOG 0-1 were randomized to receive 2×10 6 , 1×10 6 , or 0.5 × 10 6 CD19 CAR T cells / kg, in which autologous anti-CD19 CAR T cells carrying the same CAR construct as axicabtagene ciloleucel were administered after conditioning chemotherapy (cyclophosphamide: 900 mg / m 2 Fludarabine: 25 mg / m 2 , -4, -3, -2) were prepared according to the method described in International Application No. PCT / US2016 / 057983, a process that removes circulating tumor cells.

[0221] 1×10 6Modified adverse event management (or modified AE management) was implemented for patients in the dose cohort, with corticosteroids at the onset of grade ≥2 NE (instead of grade ≥3), and tocilizumab used for active toxicity only. The primary endpoint was dose-limiting toxicity (DLT) rate. Patients with prior allogeneic SCT >100 days and / or prior blinatumomab were included. Median age was 46 years (range, 18-77). Thirty patients (66%) had received ≥3 prior therapies, and the median pre-treatment BM blasts was 70% (range, 0-97). Six, 23, and 16 patients had ≥2, 1, and 0.5 × 10 6 Patients received 1 × 10 cells / kg. There were no DLTs in DLT-evaluable patients. The most common grade ≥ 3 AEs were hypotension (38%), pyrexia (38%), and thrombocytopenia (31%). Of the 41 patients, 68% had a CR / CRi and 73% had undetectable MRD. 6 Of 19 patients treated with 1 × 10 cells / kg, 16 (84%) had a CR / CRi (CR with incomplete hematologic recovery) and median event-free survival was 15 months. 6 In the nine patients treated with cells / kg and modified AE management, two (22%) had grade 3 CRS and one (11%) had grade 3 NE. No grade 4 or higher CRS or NE was observed with modified AE management. These patients also demonstrated a reduction in the median duration of NE, reduced rates of grade 3 / 4 NE, grade 3 or higher TEAEs (treatment-emergent adverse events), CD19 CAR T-cell administration-related TEAEs, and CRS.

[0222] 1×10 6 In patients treated with CD19 CAR-T cells / kg; 84% who had a CR or CRi were MRD negative. ORR was consistent across key covariates including resistant subgroups and prior transplant. Revised AE management guidelines calling for early steroid intervention for neurotoxicity and use of tocilizumab only in the setting of CRS reduced the incidence and severity of CRS and NE and shortened the duration of NE. [Table 3]

[0223] Example 4 This study will enroll patients with R / R LBCL (including DLBCL, PMBCL, TFL and high-grade B-cell lymphoma) in an open-label study evaluating the safety and efficacy of axicabtagene ciloleucel in combination with other therapeutic agents. Cohort 1: Patients will receive rituximab (RITUXAN®) 375 mg / m on day -5. 2 (intravenous) and fludarabine 30 mg / m on days -5, -4, and -3. 2 and cyclophosphamide 500 mg / m 2 Patients will receive conditioning chemotherapy with 2 × 10 6 Patients will receive axicabtagene ciloleucel at a target dose of anti-CD19 CAR T cells / kg. Patients will also receive 5 additional doses of rituximab at 28-day intervals after CAR T cell infusion. Cohort 2: Patients will receive lenalidomide 10 mg (REVLIMID®) daily starting 7 days after leukapheresis and continuing until day 3 after axicabtagene ciloleucel infusion. Fludarabine 30 mg / m 2 and cyclophosphamide 500 mg / m 2 Conditioning chemotherapy with 2×10 6 Patients will receive axicabtagene ciloleucel at a target dose of 1000 anti-CD19 CAR T cells / kg. Patients will also receive lenalidomide 20 mg every 28 days (21 days / 28 days on treatment) for five additional cycles after CAR T cell infusion. In both cohorts, some patients will receive Mesnex per institutional guidelines and may be administered according to the package insert.

[0224] The primary outcome measure is the complete response (CR) rate. The CR rate is defined as the incidence of CR according to the Lugano classification. Secondary outcome measures include, but are not limited to, the percentage of participants experiencing adverse events; the percentage of participants experiencing clinically significant changes in safety laboratory values; objective response rate (ORR, incidence of either CR or partial response (PR) according to the Lugano classification); duration of response (DOR, for participants experiencing an objective response, the time from first objective response to disease progression according to the Lugano classification); progression-free survival (PFS, the time from the date of infusion to the date of disease progression according to the Lugano classification); overall survival (OS, the time from infusion to the date of death); and the level of axicabtageneciloleucel in the blood.

[0225] Some of the patient selection parameters include: histologically proven large B-cell lymphoma (including diffuse large B-cell lymphoma (DLBCL) not otherwise specified, primary mediastinal large B-cell lymphoma, DLBCL arising from follicular lymphoma, and high-grade B-cell lymphoma); chemotherapy-resistant disease (including non-response to first-line therapy (primary refractory disease), excluding individuals who are intolerant to first-line chemotherapy: progressive disease (PD) as best response to first-line therapy, stable disease (SD) as best response after at least four cycles of first-line therapy (e.g., four cycles of RCHOP) with duration of SD within 6 months of the last dose of therapy), second-line or later. non-response to treatment (includes PD as best response to the most recent treatment regimen and SD as best response after at least two cycles of last-line therapy, with SD duration within 6 months of the last dose of therapy); resistance after autologous stem cell transplant (ASCT) (includes disease progression or recurrence within 12 months after ASCT (biopsy-confirmed recurrence in individuals who relapsed; if salvage therapy was administered after ASCT, the individual did not respond to or subsequently relapsed from last-line therapy); at least one measurable lesion by Lugano classification (lesions previously treated with radiation therapy are considered measurable only if progression is documented after radiation therapy has been completed); individuals must have at least one anti-CD20 Patients must have received sufficient prior therapy, including mAb and / or anthracycline-containing chemotherapy regimens; no evidence, suspicion, or history of lymphoma involvement in the central nervous system (CNS) or detectable cerebrospinal fluid malignant cells or brain metastases; at least 2 weeks or 5 half-lives, whichever is shorter, must have elapsed since any prior systemic therapy at the time the individual is scheduled for leukapheresis; toxicities from prior therapy must have stabilized and resolved to grade 1 or less (excluding clinically insignificant toxicities, e.g., alopecia); Eastern Cooperative Oncology Group (ECOG) performance status of 0 or is 1; absolute neutrophil count (ANC) ≥ 1500 / μL. Growth factors within 7 days prior to screening fail to meet ANC eligibility criteria; platelet count ≥ 100,000 / μL. Transfusions within 7 days prior to screening fail to meet platelet eligibility criteria; absolute lymphocyte count ≥ 100 / μL; adequate renal, hepatic, pulmonary and cardiac function, defined as creatinine clearance (estimated by Cockcroft Gault) ≥ 60 mL / min, serum alanine aminotransferase (ALT) / aspartate aminotransferase (AST) ≤ 2.5 upper limit of normal (ULN), total bilirubin ≤ 1.5 mg / dL (except for individuals with Gilbert's syndrome), cardiac ejection fraction ≥ 50%, and no evidence of pericardial effusion, no clinically significant pleural effusion, baseline oxygen saturation > 92% on room air; individuals must be able to comply with the requirements of the REVLIMID REMS®.

[0226] Some of the patient exclusion criteria include: known CD19-negative or CD20-negative tumors; history of Richter's transformation of chronic lymphocytic leukemia (CLL); prior lenalidomide or other immunomodulatory imide drug (IMiD) treatment; prior CAR therapy or other genetically modified T cell therapy; hypersensitivity to rituximab; history of severe immediate hypersensitivity reactions due to aminoglycosides; presence or suspicion of fungal, bacterial, viral, or other infections that are uncontrolled or require IV antimicrobials for management, uncomplicated urinary tract infections (UTIs) and uncomplicated bacterial pharyngitis are permitted after consultation with the sponsor's medical monitor if responsive to active treatment; history of human immunodeficiency virus (HIV) infection or acute or chronic hepatitis B or C infection. Individuals with a history of hepatitis infection should be considered for the most recent Infectious Diseases. Must be cured of their infection as determined by standard serologic and genetic testing per International Diseases Society of America (IDSA) guidelines or applicable national guidelines; history of non-melanoma skin cancer in situ (e.g., cervical, bladder, breast) or low-grade (Gleason ≦6) malignancy other than prostate cancer, or surveillance with no planned treatment (unless disease-free for ≥3 years); autologous SCT within 6 weeks of planned enrollment; prior solid organ transplant, including prior allogeneic stem cell transplant (SCT); prior CD19-targeted therapy; in the investigator's judgment, the individual is unlikely to complete all protocol-required study participation or treatment.

[0227] Example 5 The anti-CD19 CAR-T cell product used in CD19 CAR-T was manufactured by obtaining leukocytes from the leukapheresis product, enriched by selecting CD4+ / CD8+ T cells, activated by culture with anti-CD3 and anti-CD28 antibodies, and transduced with a retroviral vector containing the anti-CD19 CAR gene.

[0228] This study described the treatment of a subject with stage IV relapsed / refractory pleomorphic mantle cell lymphoma (MCL) (ibrutinib resistance). The subject was previously treated with rituximab + bendamustine (partial remission) with maintenance rituximab (progression), followed by 2 cycles of rituximab + bendamustine (mixed response) with 3 doses of maintenance rituximab, approximately 17 cycles of acalabrutinib (complete response, then progression), and 1 cycle of ibrutinib (progression), which was completed 3 weeks prior to leukapheresis. There was no history of neurological disease. Prior to CD19 CAR-T infusion, the patient had an ECOG performance status score of 0, non-mass disease, and a Ki-67 proliferation index of 80%-90%. After leukapheresis, the patient received conditioning chemotherapy followed by infusion of CD19 CAR-T. Prophylactic levetiracetam was initiated prior to the infusion and there were no signs of fever or infection.

[0229] Twenty-four hours after CD19 CAR-T infusion, grade 1 CRS was observed, and The patient was treated with siltuximab (8 mg / kg intravenous (IV)). Vancomycin and aztreonam (1 gm each IV twice daily) were administered and grade 2 encephalopathy was observed. By day 3, the subject had leukopenia with a white blood cell count of 3.7 K / uL (normal absolute neutrophil count). On day 4, grade 2 CRS was treated with siltuximab (11 mg / kg IV). Methylprednisolone 500 mg IV twice daily was initiated. Some clinical improvement was observed, but the subject's condition worsened and a second dose of tocilizumab was administered. Grade 4 encephalopathy was observed. Mannitol 20% (0.25 mg / kg) every 6 hours was initiated along with steroids and cerebral edema management guidelines were followed. By day 5, grade 3 hypertransaminaemia was observed. The cerebrospinal fluid (CSF) was clear and the opening pressure was 20 cmH. 2 O was elevated, CSF protein (600 mg / dL) and glucose (106 mg / dL) were elevated, and cytology showed increased leukocytes (24 cells / μL) and atypical and activated lymphocytes (predominantly T cells) mixed with histiocytes and neutrophils (78%). Intrathecal hydrocortisone and ara-C were administered because of worsening neurological symptoms. On day 6, EEG showed bihemispheric cerebral cortical dysfunction, and MRI showed cerebral edema and sulcal hyperintensities. At neurosurgery, an external ventricular drain was placed to treat the cerebral edema. Prophylactic acyclovir was initiated to reduce the risk of infection.

[0230] ATG (2 mg / kg / day IV) was administered with increased methylprednisolone (1 g twice daily) and a third dose of tocilizumab to suppress the systemic cytokine storm. On day 7, MRI showed stable changes and a second dose of ATG was administered. Improvement in hypertransaminasemia was noted. A third dose of ATG was administered on day 8, after which hypertransaminasemia resolved. Over the next 11 days, tapered doses of methylprednisolone were administered with clinical improvement. The ventricular drain was removed on day 14, and encephalopathy resolved. MRI on day 20 showed persistent but significantly improved periventricular white matter T2 hyperintensities and resolution of abnormal signals in the dorsal brainstem / thalamus. The patient was discharged to a rehabilitation facility on tapered doses of oral steroids. Two months after CD19 CAR-T, brain MRI findings had completely resolved. The patient remained on study and after 24 months of follow-up, the patient is in complete remission with no persistent neurological or cognitive deficits.

[0231] IL-2, a homeostatic cytokine produced by activated CAR T cells, was measured at 16.7 pg / mL in serum on day 3 and may be related to CAR T cell proliferation. The median at this same time point in the broader cohort was 4.7 pg / mL (IQR, 2.2-10.0), representing a 3.6-fold increase. After ATG (day 7), IL-2 was suppressed (below the limit of detection in the assay used), indicating a potential impact of ATG on anti-CD19 CAR T cell activity. Further serum biomarker analysis after CD19 CAR-T indicated an increase in controlled CAR T cell and myeloid-related activity. A decrease in serum interferon (INF)-γ was observed between day 3 (after CD19 CAR-T) and day 7 (after ATG) compared to the broader population. Serum IFN-γ, a Th1 cytokine and hallmark of CAR-T cell activity, was 584.4 pg / mL on day 3 and declined to 7.5 pg / mL (lower limit of detection) on day 7 after ATG administration. In contrast, median IFN-γ in the broader cohort was 97.8 pg / mL (IQR, 24.4 to 262.4) on day 3 and 187.7 pg / mL (IQR, 20 to 1243.6) on day 7.

[0232] Monocyte chemotactic protein 1 (MCP-1; CCL2), a marker of myeloid-associated activity associated with CD19-directed CAR T cytotoxicity, was 1500 pg / ml (above the limit of quantification) on day 3 (cohort median, 712.3 pg / ml [IQR, 517.6-1142.4]), suggesting the possibility of excessive myeloid activity after infusion as a contributing factor. On day 7, serum MCP-1 had decreased to 96.9 pg / mL (cohort median, 463.8 [IQR, 271.4-954.1]), further demonstrating the influence of ATG in suppressing myeloid-associated inflammation. It was done.

[0233] Analysis of cytokines in CSF collected on day 8 showed elevated levels of IFNγ-inducible protein 10 (CXCL10) and MCP-1, as well as elevated levels of intercellular adhesion molecule 1, IL-1 receptor antagonist, and IL-2 receptor alpha, compared with those measured in serum on day 7 (Table S1). Elevated levels of these cytokines in the CSF are consistent with CAR T cell and bone marrow trafficking to the central nervous system (CNS) and may contribute to NE. The subject achieved a profound and durable response, undergoing a complete recovery without neurological deficit. Improvement in cerebral edema was observed after administration of ATG, which correlated with improved transaminemia. Pharmacokinetic and pharmacodynamic results showed that ATG could contribute to the resolution of cerebral edema together with other measures, including corticosteroids with siltuximab or tocilizumab, IL-6 or IL-6 receptor blockade, and ventriculostomy. This result may lead to revision of the adverse event management guidelines of NE and CRSS.

[0234] Example 6 In cohort 1+2 (C1+2) of a trial of axicabtagene ciloleucel in patients with refractory LBCL, grade ≥ 3 cytokine release syndrome (CRS) and neurological events (NE) occurred in 11% and 32% of patients, respectively. Patients were treated with fludarabine 30 mg / m on days -5, -4, and -3. 2and cyclophosphamide 500 mg / m 2 On day 0, axicabtagene ciloleucel was added at 2 × 10 6 Patients received conditioning chemotherapy with a target dose of anti-CD19 CAR T cells / kg. In C1+2, the objective response rate (ORR) was 83% with a complete response (CR) rate of 58%. A nonrandomized safety expansion cohort was added (cohort 4 [C4]) to evaluate the effect of early steroid use on the rate of CRS and NE. Early results suggested that early use of steroids may help reduce the incidence of severe CRS and NE without affecting response rates or CAR T cell expansion.

[0235] Eligible patients will have been leukapheresised and may have optionally received bridging chemotherapy (not permitted for C1+2) and will receive 2×10 6 Conditioning chemotherapy (fludarabine and cyclophosphamide) was received prior to axicabtagene ciloleucel infusion at a target dose of anti-CD19 CAR T cells / kg. Patients in C4 received early steroid intervention starting with grade 1 NE and grade 1 CRS that did not improve after 3 days of supportive care. Primary endpoints were the incidence and severity of CRS and NE. Further endpoints were efficacy outcomes and biomarker analysis, including blood levels of CAR T cells and inflammatory markers. ORR and CAR T cell levels in C1+2 and 4 were compared across quartiles of tumor burden, and values ​​were determined by C1+2.

[0236] Forty-one patients were treated with a median follow-up of 8.7 months (range, 2.9–13.9 mo). Only one patient was lost to follow-up >6 months due to delayed dosing after the primary analysis trigger. All patients (68%) who received bridging therapy prior to treatment had evidence of disease after bridging, documented by a new baseline PET / CT scan. Disease types were diverse and included DLBCL (63%), TFL (24%), PMBCL (5%), and HGBCL (7%). Nearly half of all patients (49%) had ECOG1, 70% had stage III / IV disease, 68% were refractory to second-line or later therapy, 12% were relapsed to second-line or later therapy, 63% had prior third-line or later therapy, and 20% relapsed after ASCT. Overall, patients enrolled in C4 had lower median tumor burden by sum of two-way products (SPD; C4: 2100 mm2; C1+2: 3723 mm2) and lower pre-Tx serum LDH levels compared to C1+2.

[0237] Compared with C1+2, more patients in C4 received steroids and tocilizumab (73% and 76% vs. 27% and 43%). Early steroid use appears to affect the percentage of patients with severe CRS or NE, with fewer patients in C4 experiencing grade ≥3 CRS (2%) and NE (17%) than previously observed in C1+2. ORR in C4 was 73%, with a CR rate of 51%. Fifty-four percent of patients maintained a sustained response at ≥6 months of follow-up, compared with a sustained response rate of 44% in the primary analysis of C1+2 (also at ≥6 months of follow-up). Patients in C4 generally had lower SPD than patients in C1+2, but responses were comparable between cohorts when assessed by tumor burden. Median DOR was 8.9 months, consistent with that observed in the primary analysis of cohorts 1+2 (8.1 months; Locke, AACR 2017). Median PFS was 11.7 months, and median OS was not reached. CAR T cell proliferation was comparable between C1+2 and 4, i.e., CAR peak levels of 42 cells / μL blood in C1+2 vs. 59 cells / μL in C4, and C1+2 had a median CAR AUC of 462 cells / μL × days vs. 512 cells / μL in C4. CAR T cell proliferation was also comparable between cohorts when adjusted for tumor burden. Of note, C4 vs. 1+2 appeared to have lower levels of key biomarkers associated with severe NE, including ferritin (pre-Tx) and IL-2 (post-Tx). Early steroid use appears to reduce the rate of CAR T cell therapy-related CRS and NE in C4 compared with C1+2, without a clinically meaningful impact on efficacy at a median follow-up of 8.7 months.

[0238] Example 7 In a previous study, the median follow-up for axicabtagene ciloleucel treatment was 27.1 months, with an overall response rate of 83%, with 39% of treated patients experiencing a durable response. Tumor biopsies obtained before cell therapy and at the time of recurrence were analyzed.

[0239] Tumor tissue samples from patients in cohorts 1 and 2 of the same study described above were analyzed for protein expression of B-cell lineage markers (CD19, CD20, PAX5, CD79a, and CD22) by multiplex immunohistochemistry (IHC), followed by multiplex immunofluorescence (IF) staining and confocal microscopy of representative cases. Pretreatment tissue samples were available from 96 patients and 21 were available after relapse. Paired pretreatment and postrelapse samples were available for 16 patients. H-scores for CD19 and CD20 were derived based on the percentage and intensity of antigen expression. A score of 0-5 was considered negative and a score of 6-300 was considered positive. CD19 splice variants were assessed by RNA sequencing.

[0240] Among all patients with available post-relapse samples, 7 / 21 (33%) showed loss of CD19 expression. Analysis of 16 paired pre- and post-treatment samples showed loss of CD19 expression in 4 patients (25%) who relapsed after axicabtagene ciloleucel. 19 post-relapse tumor samples were evaluable for other B-cell lineage markers and showed preservation of CD20, CD22, and CD79a as well as the B-cell lineage transcription factor PAX5, including samples in which CD19 expression was lost or substantially reduced. Multiplex IF showed that CD19 and CD20 were expressed on the cell membrane, and analysis revealed the presence of malignant cells with different relative expression levels of these two antigens within a given biopsy. Of the 96 pre-treatment tumor samples, IHC analysis showed that CD20 was expressed in nearly all samples along with CD19, even though all patients had relapsed after previously receiving a rituximab-based regimen. The expression levels of CD19 and CD20 in these tumor biopsies obtained before cell therapy did not correlate with each other.

[0241] RNA sequencing was similar to that previously described in B-cell acute lymphoblastic leukemia. showed alternative splicing of CD19 with loss of exon 2 and / or exon 5 / 6 in diffuse large B-cell lymphoma tumors at baseline and / or relapse. Furthermore, several novel splice junctions have been identified. Correlations between H-score and CD19 splice forms and clinical outcomes including response and progression-free survival were analyzed. In this cohort of patients who relapsed after axicabtagene ciloleucel, reduced CD19 expression by IHC compared to pretreatment was common, which may be related to alternative splicing and selection of variants lacking the target epitope. Furthermore, data showed that expression of alternative B-cell lineage antigens was preserved. CD20 cell surface expression was increased in most tumors despite preceding rituximab-based therapy.

[0242] Example 8 This is a study to conduct a post-approval evaluation of axicabtagene ciloleucel and follow patients for 15 years through an established cell therapy registry. The overall median age was 61 years, 101 (34%) patients were 65 years or older, and 197 (67%) patients were male. Baseline clinical characteristics included Eastern Cooperative Oncology Group (ECOG) performance score 0-1 (77%), transformed lymphoma (27%), double-hit lymphoma (36%), previous autologous transplant (34%), and prior chemotherapy-refractory disease (66%). Median time from diagnosis to infusion was 18 months (range 2-274 months). Overall response rate (ORR) was 70% (complete response [CR] 52% and partial response [PR] 18%). Patients aged 65 years or older performed generally similarly to those aged <65 years, with slightly better CR rates (62% vs. 46%, p=0.03), but similar overall response rates (CR+PR, 75% vs. 67%, p=0.26). Cytokine release syndrome (CRS) of any grade was reported in 83% of patients. The incidence of grade ≥3 CRS was 11% according to Lee et al. 2014 and 14% according to the American Society for Transplant and Cellular Therapy (ASTCT) consensus grading. The median time to any grade CRS was 3 days (range, 1–17 days), and 94% of CRS cases resolved with a median duration of 7 days (range, 1–121 days). Among CRS patients, tocilizumab, corticosteroids, and siltuximab were used in 70%, 26%, and 1% of cases, respectively. Neurological adverse events (AEs) of any grade occurring after axicabtagene ciloleucel infusion were reported in 181 (61%) patients. One patient was reported to have died due to cerebral edema. The median time to onset of any grade neurological AE was 6 days (range, 1-82 days), and 88% had resolved by the time of data submission with a median duration of 8 days (range, 1-105 days). Corticosteroids were used in 56% of patients for treatment. Patients aged 65 years or older had similar CRS (85% vs. 82%, p=0.62), grade ≥3 CRS (13% vs. 9%, p=0.62), and neurological AEs (68% vs. 58%, p=0.13) compared with patients <65 years.Prolonged cytopenias (thrombocytopenia and neutropenia), defined by failure to resolve within 30 days after administration of axicabtagene ciloleucel, occurred in 7% of patients. Preliminary data reveal that subsequent neoplasms were reported in six patients (2%): myelodysplasia (n=3), lung cancer (n=1), neuroendocrine tumor (n=1), and cutaneous squamous cell carcinoma (n=1).

[0243] Example 9 This was a single-arm, phase 1 / 2 trial in adults with refractory diffuse large B-cell lymphoma (DLBCL), primary mediastinal B-cell lymphoma (PMBCL), or transformed follicular lymphoma (TFL). Refractory disease was defined as failure to respond to last chemotherapy or relapse within 12 months after autologous stem cell therapy (ASCT). Patients were also required to have an Eastern Cooperative Oncology Group (ECOG) performance status of 0 or 1 and to have received anti-CD20 monoclonal antibody therapy. Patients were also required to have received prior treatment with both an anti-cancer antibody and an anthracycline-containing regimen. 2 / day) and fludarabine (30 mg / m 2 Following a 3-day conditioning regimen of 100 mg / day, patients received 1 dose of axicabtagene ciloleucel (axi-cel) (2 × 10 6 Patients received 100 mg / kg of CAR T cells. Patients were followed for a minimum of 24 months from axicabtagene ciloleucel infusion (median follow-up was 27.1 months). Safety evaluations included all treated patients enrolled in phase 1 and phase 2 (age 65 and older, n=27; age <65, n=81). Efficacy evaluations included patients treated in phase 2 only (age 65 and older, n=24; age <65, n=77).

[0244] Baseline characteristics were largely similar between patients aged 65 years and younger, with a few exceptions (Table 4). Most notably, patients aged 65 years and older were numerically more likely to have an International Prognostic Index (IPI) score of 3-4 than the younger age group, which is attributable to age >60 being a component of IPI scoring (Shipp MA,H. et al., A predictive model for aggressive non-Hodgkin's lymphoma. The International Non-Hodgkin's Lymphoma Prognostic Factors Project. N Engl J Med. 1993;329(14):987-994). Because patients aged 65 years and older were less likely to be considered for ASCT, a larger proportion of patients younger than 65 years underwent ASCT. In vivo CAR T cell expansion was similar for patients aged 65 years and older and those younger than 65 years (median peak expansion, 43.0 and 35.3 CAR T cells / μL of blood, respectively; P=0.769; median area under the curve from day 0 to day 28, 562.0 and 448.4 CAR T cells / μL, respectively; P=0.983). [Table 4]

[0245] AE, adverse event; CRS, cytokine release syndrome; SPD, two-way sum of products; *The most common grade ≥3 AEs occurring in ≥25% of any age group are shown. † Neutropenia includes the terms neutropenia, febrile neutropenia, and neutropenia. ‡Thrombocytopenia includes the terms thrombocytopenia and thrombocytopenia. §Symptoms indicated are those occurring in 10% or more of patients in any age group. ||Patients in response at data cutoff. Efficacy outcomes were analyzed. There were a total of 24 patients aged 65 years or older and 77 patients aged <65 years in this analysis. Investigator-assessed objective response rates (ORR) were similar (92% vs. 81%, respectively). Patients aged 65 years or older were numerically more likely than patients <65 years to have a complete response (CR) as their best response (75% vs. 53%, respectively) and a durable response at data cutoff (42% vs. 38%, respectively). The distribution for partial response (PR) was 17% vs. 27%, respectively, and 24-month overall survival was 54% vs. 49%, respectively.

[0246] Similar trends were observed when evaluating response rates by histologic subtype among patients aged ≥65 vs. <65 years. Objective response rates were 92% vs. 81% for all LBCL, 88% vs. 82% for DLBCL, and 100% vs. 78% for TFL patients. Complete response rates were 75% vs. 53% for all LBCL, 65% vs. 50% for DLBCL, 100% vs. 56% for TFL patients, and 75% for PMBCL patients <65 years.

[0247] Median duration of response was 12.0 vs. 8.1 months in patients aged 65 years or older vs. <65 years, respectively. Median progression-free survival (95% CI) was 13.2 vs. 5.6 months, respectively. Approximately 50% of patients in both age groups were alive 24 months after treatment (54% vs. 49%, respectively). Efficacy outcomes in this study (ZUMA-1) were comparable to those reported in the 2017 SCHOLAR-1 study, a pooled retrospective analysis of 636 DLBCL patients who were refractory to chemotherapy or relapsed within 12 months of ASCT. (Crump M et al., Outcomes in refractory diffuse large B-cell lymphoma: results from the international SCHOLAR-1 study. Blood. 2017;130(16):1800-1808). For patients aged 65 years and older and younger than 65 years, available treatments in the pre-CAR T cell era resulted in ORRs of only 19% and 27%, respectively, and 2-year survival rates of 19% and 20%, respectively.

[0248] Adverse events (AEs) are summarized in Table 4. The most common grade 3 or higher AE was cytopenia, which occurred in similar proportions in treated patients aged ≥65 years and <65 years. The most common grade 3 or higher cytopenia present at or after 93 days post-treatment was neutropenia, which was reported in 15% and 10% of patients ≥65 years and <65 years, respectively. The rate of grade 3 or higher cytokine release syndrome (CRS) was 7% vs. 12%, respectively. Grade 3 or higher neurological events were observed in 44% vs. 28%, respectively. Several neurological event-related symptoms, such as grade 3 or higher delirium and encephalopathy, were observed in numerically higher proportions in patients ≥65 years than in patients <65 years, which may be commensurate with age. Grade 3 or higher infections were reported in 19% vs. 31%, respectively. A total of 26% vs. 32%, respectively, received intravenous immunoglobulin therapy at the investigator's discretion. As previously reported, grade 5 AEs were observed in a total of 4 patients (4% of each age group). Locke FL et al., Long-term safety and activity of axicabtagene ciloleucel in refractory large B-cell lymphoma (ZUMA-1): a single-arm, multicentre, phase 1-2 trial. Lancet Oncol. 2019;20(1):31-42; Neelapu SS et al., Axicabtagene ciloleucel CAR-T-cell therapy in refractory large B-cell lymphoma. N Engl J Med.2017;377(26):2531~2544;Locke FL et al., Phase 1 results of ZUMA-1:a multicenter study of KTE-C19 anti-CD19 CAR Tcell therapy in refractory aggressive lymphoma.Mol Ther.2017;25(1):285-295.

[0249] These safety findings are consistent with a pooled analysis of 214 patients with large B-cell lymphoma enrolled in clinical trials of tisagenlecleucel and axicabtagene ciloleucel, which reported similar rates of grade 3 or higher CRS in patients aged 65 years and younger. Sharma P et al., A US Food and Drug Administration age-based pooled analysis of cytokine release syndrome and neurotoxicity in subjects with relapsed / refractory lymphoma treated with chimeric antigen receptor (CAR) Tcell therapy. Blood. 2018. 132(Suppl 1):4201-4201. Compared to this analysis of ZUMA-1, a subgroup analysis of 300 patients treated with axicabtagene ciloleucel in the standard of care setting showed similar ORR and CRS rates in patients aged 65 years or older and those aged <65 years, but slightly higher rates of complete response and all-grade neurological events. (Sano D. et al., Safety and efficacy of axicabtagene ciloleucel (axi-cel) in older patients: Results from US Lymphoma CAR-T Consortium. Hematological Oncology. 2019; 37(S2): 304-305).

[0250] In this subgroup analysis of the 2-year follow-up of ZUMA-1, axicabtagene ciloleucel induced a high proportion of durable responses with a manageable safety profile, regardless of age. No age-related differences in efficacy, pharmacokinetic profile, or safety were observed, suggesting that age alone cannot limit the use of axicabtagene ciloleucel. Overall, axicabtagene ciloleucel demonstrated substantial clinical benefit in elderly patients with refractory large B-cell lymphoma, a population with limited treatment options. (Nastoupil LJ, et al., Axicabtagene Ciloleucel (Axi-cel) CD19 Chimeric Antigen Receptor (CAR) T-Cell Therapy for Relapsed / Refractory large B-Cell Lymphoma: Real World Experience.Blood.2018:132(Suppl 1):abstract 91).

Claims

1. 1. A method of treating relapsed or refractory B-cell lymphoma or ALL in a subject in need of treatment, comprising administering to the subject a therapeutically effective amount of a CD19-directed genetically modified T-cell immunotherapy.

2. 10. The method of claim 1, further comprising monitoring the patient after infusion for signs and symptoms of adverse reactions and / or safety.

3. 3. The method of claim 1 or 2, wherein the adverse reaction is selected from the group consisting of cytokine release syndrome (CRS), neurotoxicity, hypersensitivity reactions, serious infections, cytopenias, and hypogammaglobulinemia.

4. 4. The method of any one of claims 1 to 3, wherein the signs and symptoms of the adverse reaction are selected from the group consisting of fever, hypotension, tachycardia, hypoxia, and chills, and include cardiac arrhythmias (including atrial fibrillation and ventricular tachycardia), cardiac arrest, heart failure, renal insufficiency, capillary leak syndrome, hypotension, hypoxia, organ toxicity, hemophagocytic lymphohistiocytosis / macrophage activation syndrome (HLH / MAS), seizures, encephalopathy, headache, tremor, dizziness, aphasia, delirium, insomnia anxiety, anaphylaxis, febrile neutropenia, thrombocytopenia, neutropenia, and anemia.

5. 5. The method of any one of claims 2 to 4, wherein the adverse reaction is (1) neurotoxicity, optionally wherein symptoms of the neurotoxicity are encephalopathy, headache, tremors, dizziness, aphasia, delirium, insomnia, and / or anxiety, and / or (2) cytopenia.

6. (i) administering an effective amount of an inhibitor or combination of inhibitors of GM-CSF, CSF1, GM-CSFR, or CSF1R to treat the symptoms of the adverse reaction; (ii) administering an effective amount of an immunosuppressant to deplete T lymphocytes, e.g., antithymocyte globulin (ATG), preferably wherein the adverse reaction is cerebral edema; (iii) administering an effective amount of a steroid (e.g., a corticosteroid) and / or tocilizumab to treat the symptoms of the adverse reaction; (iv) administering a non-sedating anti-seizure medication for seizure prevention; (v) administering at least one of erythropoietin, darbepoetin alfa, platelet transfusions, filgrastim, or pegfilgrastim, and / or administering tocilizumab; (vi) administering antithymocyte globulin, and / or The method of any one of claims 1 to 5, further comprising (vii) administering rituximab.

7. (i) the GM-CSF inhibitor is selected from lenzilumab, namilumab (AMG203), GSK3196165 / MOR103 / otilimab (GSK / MorphoSys), KB002 and KB003 (KaloBios), MT203 (Micromet and Nycomed), MORAb-022 / dimcirumab (Morphotek), or a biosimilar of any one of them, E21R, and a small molecule; (ii) the CSF1 inhibitor is selected from RG7155, PD-0360324, MCS110 / Lanotuzumab), or a biosimilar version of any one of them, and a small molecule; and / or (iii) the GM-CSFR inhibitor and the CSF1R inhibitor are selected from the group consisting of mavrilimumab (formerly CAM-3001; MedImmune, Inc.), caviralizumab (Five Prime Therapeutics), LY3022855 (IMC-CS4) ( 7. The method of claim 6, wherein the therapeutic agent is selected from Emactuzumab, also known as RG7155 or RO5509554, FPA008 (Five Prime / BMS), AMG820 (Amgen), ARRY-382 (Array Biopharma), MCS110 (Novartis), PLX3397 (Plexxikon), ELB041 / AFS98 / TG3003 (ElsaLys Bio, Transgene), SNDX-6352 (Syndax), a biosimilar version of any one of them, and a small molecule.

8. 8. The method of claim 6 or 7, wherein at least one of said inhibitors of GM-CSF, CSF1, GM-CSFR, or CSF1R or a combination of inhibitors is administered on the same day as the CD19-directed genetically modified T cell immunotherapy.

9. 9. The method of any one of claims 6 to 8, wherein at least one of said inhibitors of GM-CSF, CSF1, GM-CSFR, or CSF1R or a combination of inhibitors is administered more than once following administration of CD19-directed genetically modified T cell immunotherapy.

10. 10. The method of any one of claims 6 to 9, wherein at least one of said inhibitors of GM-CSF, CSF1, GM-CSFR or CSF1R or a combination of inhibitors is administered 1 to 12 hours prior to, and optionally simultaneously with, administration of the CD19-directed genetically modified T cell immunotherapy.

11. 11. The method of any one of claims 6 to 10, wherein at least one of said inhibitors of GM-CSF, CSF1, GM-CSFR, or CSF1R or a combination of inhibitors is administered more than once following administration of CD19-directed genetically modified T cell immunotherapy.

12. 12. The method of any one of claims 6 to 11, wherein the inhibitor or combination of inhibitors is selected from lenzilumab and mavrilimumab.

13. CD19-directed genetically modified T cell immunotherapy was administered at 2 × 10 ６ Target dose of CAR-positive viable T cells / kg (maximum tolerated dose: 2×10 ８ 14. The method of any one of claims 1-13, wherein the medicamentous sera are administered at 3 mg / kg body weight as a single IV infusion of 100 mg / mL of erythrocyte sedimentation medium (e.g., erythrocytes), and mavrilimumab is administered once subcutaneously, OP / IP, or IV on day 0 at 3 mg / kg body weight.

14. The method of any one of claims 2 to 13, wherein the adverse reaction is cytokine release syndrome (CRS).

15. 15. The method of any one of claims 2 to 14, comprising monitoring for signs and symptoms of cytokine release syndrome (CRS) for about 7 days following infusion, optionally at least daily.

16. (i) If greater than Grade 2 cytokine release syndrome (CRS) is observed, administer tocilizumab IV at a dose of approximately 8 mg / kg over 1 hour, repeating every 8 hours as needed if unresponsive to increased IV fluids or supplemental oxygen; (ii) if the CRS symptoms observed in (b) have not improved 24 hours after (c), administer approximately 1 mg / kg methylprednisolone IV twice daily or an equivalent dose of dexamethasone and continue corticosteroids until events are Grade 1 or less, then taper over 3 days; (iii) If Grade 3 CRS is observed in (b), administer tocilizumab IV at a dose of 8 mg / kg over 1 hour, repeating tocilizumab every 8 hours as needed if unresponsive to increased IV fluids or supplemental oxygen, and methylprednisolone 1 mg / kg or an equivalent dose of dexamethasone twice daily and continue corticosteroids until events are grade 1 or less, then taper over 3 days; (iv) if CRS grade 4 is observed in (b), administer tocilizumab IV at a dose of about 8 mg / kg over 1 hour, repeating every 8 hours as needed if unresponsive to increasing IV fluids or supplemental oxygen, and administer methylprednisolone IV at about 1,000 mg per day for 3 days.

17. 17. The method of any one of claims 1 to 16, further comprising measuring cytokine and chemokine levels before and / or after said one or more administrations, optionally measuring the levels of at least one of IL-2, IL-6, IL-8, IL-10, IL-12p40 / p70, IL-15, IL-17a, TNF-α, IFN-γ, GM-CSF, and sIL2Rα.

18. 18. The method of any one of claims 2 to 17, wherein the adverse reaction is (1) neurotoxicity, optionally wherein symptoms of the neurotoxicity are encephalopathy, headache, tremors, dizziness, aphasia, delirium, insomnia, and / or anxiety, and / or (2) cytopenia.

19. administering non-sedating anti-seizure medications to prevent seizures; 19. The method of any one of claims 1 to 18, further comprising administering at least one of erythropoietin, darbepoetin alfa, platelet transfusions, filgrastim, or pegfilgrastim, and / or administering tocilizumab.

20. 20. The method of any one of claims 2 to 19, wherein if cytokine release syndrome (CRS) and / or neurotoxicity is observed, the cytokine release syndrome (CRS) and / or neurotoxicity is managed according to Table 1, Table 2, Example 2, and / or Example 3.

21. 21. The method of any one of claims 1 to 20, wherein the patient further receives 5 additional doses of rituximab at 28 day intervals following the infusion of the CD19-directed genetically modified T cell immunotherapy.

22. The patient receives rituximab 375 mg / m on day -5. ２ and fludarabine 30 mg / m on days −5, −4, and −3. ２ and cyclophosphamide 500 mg / m ２ 22. The method of claim 21 , wherein the patient receives conditioning chemotherapy with, followed by a two day rest on day −2 and day −1, followed by administration of anti-CD19 T cell immunotherapy.

23. 23. The method of any one of claims 1-22, wherein the patient further receives lenalidomide 20 mg at 28 day intervals (21 days / 28 days on treatment) for five additional cycles following anti-CD19 T cell immunotherapy.

24. The patient received lenalidomide 10 mg daily starting 7 days after leukapheresis and continuing until day 3 of anti-CD19 T-cell immunotherapy, followed by fludarabine 30 mg / m on days -5, -4, and -3. ２ and cyclophosphamide 500 mg / m ２ After conditioning chemotherapy with Anti-CD19 T cell immunotherapy was then administered at 2 × 10 ６ 24. The method of claim 23, administered at a target dose of anti-CD19 CAR T cells / kg.

25. The method of any one of claims 1 to 24, wherein the patient is further administered mesna.

26. The CD19-directed genetically modified T cell immunotherapy comprises administration of T cells genetically engineered to express an anti-CD19 chimeric antigen receptor (anti-CD19 CAR T cells). Item 26. The method according to any one of items 1 to 25.

27. 27. The method of claim 26, wherein the T cells are genetically modified ex vivo by retroviral transduction to express a chimeric antigen receptor (CAR) comprising an anti-CD19 single chain variable fragment (scFv) linked to the costimulatory domains of CD28 and CD3 zeta.

28. The method of any one of claims 1 to 27, wherein said CD19-directed genetically modified T cell immunotherapy comprises CD19-directed genetically modified T cell immunotherapy.

29. CD19-directed genetically modified T cell immunotherapy was administered on day 0 at 2×10 ６ Target dose of CAR-positive viable T cells / kg (maximum tolerated dose: 2×10 ８ and / or mavrilimumab is administered as a single IV infusion of 1 mg / kg body weight, 2 mg / kg body weight, 3 mg / kg body weight, 4 mg / kg body weight, 5 mg / kg body weight, 6 mg / kg body weight, 7 mg / kg body weight, 8 mg / kg body weight, 9 mg / kg body weight, 10 mg / kg body weight, 11 mg / kg body weight, 12 mg / kg body weight, 13 mg / kg body weight, 14 mg / kg body weight, 15 mg / kg body weight, 16 mg / kg body weight, 17 mg / kg body weight, 18 mg / kg body weight, 19 mg / kg body weight, 20 mg / kg body weight, 21 mg / kg body weight, 22 mg / kg body weight, 23 mg / kg body weight, 24 mg / kg body weight, 25 mg / kg body weight, 26 mg / kg body weight, 27 mg / kg body weight, 28 mg / kg body weight, 29 mg / kg body weight, 30 mg / kg body weight, 31 mg / kg body weight, 32 mg / kg body weight, 33 mg / kg body weight, 34 mg / kg body weight, 35 mg / kg body weight, 36 mg / kg body weight, 37 mg / kg body weight, 38 mg / kg body weight, 39 mg / kg body weight, 40 mg / kg body weight, 41 mg / kg body weight, 42 mg / kg body weight, 43 mg / kg body weight, 44 mg / kg body weight, 45 mg / kg body weight, 46 mg / kg body weight, 47 mg / kg body weight, 48 mg / kg body weight, 49 mg / kg body weight, 50 mg / kg body weight, 51 mg / kg body weight, 52 mg / kg body weight, 53 mg / kg body weight, 54 mg / kg body weight, 55 mg / kg body weight, 56 mg / kg 29. The method of any one of claims 1 to 28, wherein the compound is administered once at 7 mg / kg body weight, 18 mg / kg body weight, 19 mg / kg body weight, 20 mg / kg body weight, 21 mg / kg body weight, 22 mg / kg body weight, 23 mg / kg body weight, 24 mg / kg body weight, 25 mg / kg body weight, 26 mg / kg body weight, 27 mg / kg body weight, 28 mg / kg body weight, 29 mg / kg body weight, or 30 mg / kg, administered subcutaneously, OP / IP, or IV.

30. 30 mg / m ２ / day fludarabine and 500 mg / m ２ 30. The method of any one of claims 1-29, further comprising administering a conditioning chemotherapy comprising cyclophosphamide / day on day -5 to day -3.

31. The method of any one of claims 1 to 30, wherein the patient is refractory to first line therapy, refractory to second or subsequent line therapy, or refractory following autologous stem cell transplantation.

32. The method of any one of claims 1 to 31, wherein the patient has previously received an anti-CD20 mAb and an anthracycline-containing chemotherapy regimen.

33. 33. The method of any one of claims 1 to 32, wherein the relapsed or refractory large B-cell lymphoma is diffuse large B-cell lymphoma not otherwise specified (DLBCL), primary mediastinal large B-cell lymphoma, high-grade B-cell lymphoma, transformed follicular lymphoma, mantle cell lymphoma or diffuse large B-cell lymphoma arising from follicular lymphoma.

34. The method of any one of claims 1 to 33, wherein the patient is over 65 years of age.

Images