CN117295507A - Chimeric antigen receptor T cell therapies - Google Patents

Chimeric antigen receptor T cell therapies Download PDF

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CN117295507A
CN117295507A CN202280034095.1A CN202280034095A CN117295507A CN 117295507 A CN117295507 A CN 117295507A CN 202280034095 A CN202280034095 A CN 202280034095A CN 117295507 A CN117295507 A CN 117295507A
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antigen
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R·S·加西亚
V·普拉卡斯
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Kite Pharma Inc
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Kite Pharma Inc
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Priority claimed from PCT/US2022/029047 external-priority patent/WO2022241151A2/en
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Abstract

The present disclosure provides methods of treating malignant tumors comprising administering an effective dose of an immune cell therapy (e.g., chimeric antigen receptor genetically modified T cell immunotherapy), and methods for making such immune therapies. Some aspects of the present disclosure relate to methods of determining a patient's objective response to immune cell immunotherapy based on patient levels and product attributes before and after administration of the immunotherapy to the patient.

Description

Chimeric antigen receptor T cell therapies
Cross Reference to Related Applications
The present application claims priority from U.S. provisional patent application No. 63/188,916, filed on day 14, 5, 2021, U.S. provisional patent application No. 63/248,941, filed on day 27, 9, 2021, and U.S. provisional patent application No. 63/328,364, filed on day 7, 4, 2022, each of which is incorporated herein by reference in its entirety.
Background
Human cancer essentially consists of normal cells that undergo genetic or epigenetic transformation to become abnormal cancer cells. In this way, cancer cells begin to express different proteins and other antigens than those expressed by normal cells. The innate immune system of the body can use these abnormal tumor antigens to specifically target and kill cancer cells. However, cancer cells employ various mechanisms to prevent immune cells (such as T and B lymphocytes) from successfully targeting cancer cells.
Human T cell therapy relies on enriched or modified human T cells to target and kill cancer cells in a patient. To increase the ability of T cells to target and kill specific cancer cells, methods have been developed to engineer T cells to express constructs that direct T cells to specific target cancer cells. Chimeric Antigen Receptors (CARs) comprising a binding domain capable of interacting with a particular tumor antigen allow T cells to target and kill cancer cells expressing the particular tumor antigen.
It is necessary to know how the properties of CAR positive T cells and the immune status of a patient relate to the clinical outcome of immunotherapy.
Disclosure of Invention
Provided herein are methods and uses of cells (e.g., engineered T cells) and/or compositions thereof for treating a subject having a disease or disorder, typically or including a cancer or tumor, such as leukemia or lymphoma. In some aspects, the methods and uses provide or achieve improved response and/or more durable response or efficacy and/or reduced risk of toxicity or other side effects in subjects treated with some methods as compared to certain alternative methods. In some embodiments, the methods comprise administering a specified or relative number of engineered cells, administering a defined ratio of specific types of cells, treating a specific patient population (such as those with specific risk profiles, stages, and/or past treatment history), administering additional therapeutic agents, and/or combinations thereof.
Methods are also provided that involve assessing specific parameters, e.g., expression of specific biomarkers or analytes, that may be correlated with a result, such as a therapeutic result, including a response, such as a Complete Response (CR) or a Partial Response (PR); or safety consequences such as toxicity, e.g., neurotoxicity or CRS, following administration of cell therapy. Methods of assessing the likelihood of a response and/or the likelihood of a risk of toxicity based on an assessment of a parameter, such as the expression of a biomarker or analyte, are also provided.
In one aspect, the present disclosure provides a method of treating a cancer that expresses a tumor antigen in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of CAR T cells that express an antigen binding molecule that recognizes the tumor antigen. In some embodiments, the cancer is leukemia or lymphoma. In some embodiments, the cancer is Mantle Cell Lymphoma (MCL). In some embodiments, MCL recurs/is refractory following systemic therapy for ≡2 lines. In some embodiments, the cancer is (recurrent/refractory) indolent non-hodgkin lymphoma (iNHL). In some embodiments, the cancer is Follicular Lymphoma (FL). In some embodiments, the cancer is Marginal Zone Lymphoma (MZL). In some embodiments, the subject has MCL (MCL POD 24) with a high risk profile of progression within 24 months from the first receiving anti-CD 20 containing chemotherapy. In some embodiments, the cancer is iNHL (iNHL POD 24) with a high risk of disease progression characteristic within 24 months of diagnosis. In some embodiments, the tumor antigen is CD19. In some embodiments, CAR T cell therapy is administered early. For example, CAR T-cell therapy can be administered as first line therapy and/or prior to progression.
The following embodiments are illustrative of the present disclosure and are not limiting:
1. a method of treating cancer non-hodgkin's lymphoma (NHL), comprising administering to a subject a therapeutically effective amount of immune cells against a tumor antigen.
2. The method of embodiment 1, wherein the subject is at high risk of disease progression.
3. The method of embodiment 1 or 2, wherein the NHL is Mantle Cell Lymphoma (MCL) or Indolent NHL (iNHL).
4. The method of embodiment 1 or 2, wherein the iNHL is an edge zone lymphoma (MZL) or Follicular Lymphoma (FL).
5. The method of embodiment 2, wherein the subject is at high risk if the subject shows disease progression within 24 months after the initial diagnosis.
6. The method of embodiment 2, wherein the subject is at high risk if the subject shows disease progression within 24 months from the first receipt of anti-CD 20 containing chemotherapy.
7. The method of embodiment 6, wherein the chemotherapy comprises an alkylating agent.
8. The method of any one of embodiments 1-7, wherein the immune cells are administered as a first line, a second line, a third line, a fourth line, a fifth line, or a sixth line therapy.
9. The method of any one of embodiments 1 to 8, wherein the immune cells are selected from Tumor Infiltrating Lymphocytes (TILs), NK cells, autologous T cells, allogeneic T cells, and engineered autologous T cells (eacts), and any combination thereof.
10. The method of embodiment 9, wherein the immune cell is a CAR T cell.
11. The method of embodiment 10, wherein the CAR T cell therapy comprises alemtuquor (axicabtagene ciloleucel) or brexucabtagene autoleucel/KTE-X19.
12. The method of embodiments 1, 2 or 9, wherein the therapeutically effective amount or effective dose of immune cells is at least about 10 4 Individual cells, at least about 10 5 Individual cells, at least about 10 6 Individual cells, at least about 10 7 Individual cells, at least about 10 8 Individual cells, at least about 10 9 Individual cells or at least about 10 10 Individual cells.
13. The method of embodiment 1, 2 or 9, wherein the therapeutically effective amount or effective dose of immune cells is about 10 4 Individual cells, about 10 5 Individual cells, about 10 6 Individual cells, about 10 7 Individual cells or about 10 8 Individual cells.
14. The method of embodiment 1, 2 or 9, wherein the therapeutically effective amount or effective dose of immune cells is about 2 x 10 6 Individual cells/kg, about 3X 10 6 Individual cells/kg, about 4X 10 6 Individual cells/kg, about 5X 10 6 Individual cells/kg, about 6X 10 6 Individual cells/kg, about 7X 10 6 Individual cells/kg, about 8X 10 6 Individual cells/kg, about 9X 10 6 Individual cells/kg, about 1X 10 7 Individual cells/kg, about 2X 10 7 Individual cells/kg, about 3X 10 7 Individual cells/kg, about 4X 10 7 Individual cells/kg, about 5X 10 7 Individual cells/kg, about 6X 10 7 Individual cells/kg, about 7X 10 7 Individual cells/kg, about 8X 10 7 Individual cells/kg or about 9X 10 7 Individual cells/kg.
15. The method of embodiment 1, 2 or 9, wherein the therapeutically effective amount or effective dose of immune cells is between about 1 x 10 per kg body weight 6 And about 2X 10 6 Between immune cells up to about 1X 10 8 Maximum dose of individual immune cells.
16. The method of embodiment 1, 2 or 9, wherein the therapeutically effective dose of immune cells is between 75 x 10 6 And 200X 10 6 Between individual immune cells.
17. The method of any one of embodiments 1 to 16, wherein the tumor antigen is selected from the group consisting of a tumor-associated surface antigen, 5T4, alpha Fetoprotein (AFP), B7-1 (CD 80),
B7-2 (CD 86), BCMA, B-human chorionic gonadotrophin, CA-125, carcinoembryonic antigen (CEA), CD123, CD133, CD138, CD19, CD20, CD22,
CD23、CD24、CD25、CD30、CD33、CD34、CD4、CD40、
CD44, CD56, CD8, CLL-1, c-Met, CMV-specific antigen, CS-1,
CSPG4, CTLA-4, DLL3, bissialoganglioside GD2, ductal epithelial mucin, EBV specific antigen, EGFR variant III (EGFRvIII), ELF2M, endoglin, ephrin B2, epidermal Growth Factor Receptor (EGFR), epithelial cell adhesion molecule (EpCAM), epithelial tumor antigen, erbB2 (HER 2/neu), fibroblast-related protein (fap), FLT3, folate binding protein, GD2, GD3, glioma-related antigen, glycosphingolipids, gp36, HBV specific antigen, HCV specific antigen, HER1-HER2, a combination of HER2-HER3, HERV-K, high molecular weight melanoma-related antigen (HMW-MAA), HIV-1 envelope glycoprotein gp41,
HPV specific antigen, human telomerase reverse transcriptase, IGFI receptor, IGF-II, IL-11Rα, IL-13R-a2, influenza virus specific antigen; CD38, insulin growth factor (IGFl) -l, enterocarboxylesterase, kappa chain, LAGA-la, lambda chain, lassa virus-specific antigen, lectin-reactive AFP, lineage-specific or tissue-specific antigen such as
CD3, MAGE-A1, major Histocompatibility Complex (MHC) molecules, major MHC molecules presenting tumor-specific peptide epitopes,
M-CSF, melanoma-associated antigen, mesothelin, MN-CA IX, MUC-1, mut hsp70-2, mutant p53, mutant ras, neutrophil elastase, NKG2D,
Nkp30, NY-ESO-1, p53, PAP, prostase, prostate Specific Antigen (PSA), prostate cancer tumor antigen-1 (PCTA-1), prostate specific antigen protein, STEAP1, STEAP2, PSMA, RAGE-1, ROR1, RU2
(AS), surface adhesion molecules, survivin and telomerase, TAG-72, additional domain a (EDA) and additional domain B (EDB) of fibronectin, al domain of tenascin-C (TnC Al), thyroglobulin, tumor matrix antigen, vascular endothelial growth factor receptor-2 (VEGFR 2), virus-specific surface antigens such AS HIV-specific antigens (such AS HIV gpl 20), and any derivatives or variants of these surface antigens.
18. The method of embodiment 17, wherein the target antigen is CD19.
19. The method of embodiment 1 or 2, wherein the NHL is diffuse large B-cell lymphoma (DLBCL) non-specific, primary mediastinum large B-cell lymphoma, high grade B-cell lymphoma, or DLBCL caused by follicular lymphoma.
20. The method of any one of embodiments 1 to 19, further comprising preconditioning the subject with one or more preconditioning agents.
21. The method of embodiment 20, wherein the subject is preconditioned with administration of an alkylating agent and/or a platinum-based agent.
22. The method of embodiment 21, wherein the alkylating agent is selected from the group consisting of: melphalan, chlorambucil, cyclophosphamide, dichloromethyldiethylamine, nitrogen mustard (HN 2), uratemustine, uracil nitrogen mustard, melphalan, chlorambucil, ifosfamide, bendamustine, carmustine, lomustine, streptozotocin, alkyl sulfonates, busulfan, thiotepa, or the like, or any combination thereof.
23. The method of embodiment 21, wherein the platinum-based preconditioning agent is selected from the group consisting of: platinum, cisplatin, carboplatin, nedaplatin, oxaliplatin, satraplatin, triplatinum tetranitrate, procarbazine, altretamine, triazene, dacarbazine, mitozolomide, temozolomide, dacarbazine, temozolomide, and any combination thereof.
24. The method of embodiment 20, wherein the preconditioning agent comprises cyclophosphamide and fludarabine.
25. The method of any one of embodiments 20 to 24, wherein the administration of the one or more preconditioning agents begins at least seven days, at least six days, at least five days, at least four days, at least three days, at least two days, or at least one day prior to the administration of the cell therapy.
The following additional embodiments are illustrative of the present disclosure and are not limiting:
1. a method of treating cancer in a subject in need thereof, wherein the cancer is non-hodgkin's lymphoma (NHL) or relapsed/refractory B-precursor acute lymphoblastic leukemia or relapsed/refractory B-cell non-hodgkin's lymphoma (R/R B-ALL), the method comprising administering to the subject a therapeutically effective amount of immune cells that are anti-tumor antigens; optionally, wherein the subject is a pediatric or adolescent subject.
2. The method of claim 1, wherein the subject is at high risk of disease progression.
3. The method of claim 1 or 2, wherein the NHL is Mantle Cell Lymphoma (MCL) or Indolent NHL (iNHL).
4. The method of claim 1 or 2, wherein the iNHL is an edge zone lymphoma (MZL) or Follicular Lymphoma (FL).
5. The method of claim 2, wherein the subject is at high risk if the subject shows disease progression within 24 months after initial diagnosis.
6. The method of claim 2, wherein the subject is at high risk if the subject shows disease progression within 24 months from the first receipt of anti-CD 20 containing chemotherapy.
7. The method of claim 6, wherein the chemotherapy comprises an alkylating agent.
8. The method of any one of claims 1 to 7, wherein the immune cells are administered as a first line, a second line, a third line, a fourth line, a fifth line, and/or a sixth line therapy, or prior to disease progression.
9. The method of any one of claims 1 to 8, wherein the immune cells are selected from Tumor Infiltrating Lymphocytes (TILs), NK cells, autologous T cells, allogeneic T cells, and engineered autologous T cells (eacts), and any combination thereof.
10. The method of claim 9, wherein the immune cell is a CAR T cell.
11. The method of claim 10, wherein the CAR T cell therapy comprises alemtuzite or brexucabtagene autoleucel/KTE-X-19.
12. The method of claim 1, 2 or 9, wherein the therapeutically effective amount or effective dose of immune cells is at least about 10 4 Individual cells, at least about 10 5 Individual cells, at least about 10 6 Individual cells, at least about 10 7 Individual cells, at least about 10 8 Individual cells, at least about 10 9 Individual cells or at least about 10 10 Individual cells.
13. The method of claim 1, 2 or 9, wherein the therapeutically effective amount or effective dose of immune cells is about 10 4 Individual cells, about 10 5 Individual cells, about 10 6 Individual cells, about 10 7 Individual cells or about 10 8 Individual cells.
14. The method of claim 1, 2 or 9, wherein the therapeutically effective amount or effective dose of immune cells is about 2 x 10 6 Individual cells/kg, about 3X 10 6 Individual cells/kg, about 4X 10 6 Individual cells/kg, about 5X 10 6 Individual cells/kg, about 6X 10 6 Individual cells/kg, about 7X 10 6 Individual cells/kg, about 8X 10 6 Individual cells/kg, about 9X 10 6 Individual cells/kg, about
1×10 7 Individual cells/kg, about 2X 10 7 Individual cells/kg, about 3X 10 7 Individual cells/kg, about 4X 10 7 Individual cells/kg, about 5X 10 7 Individual cells/kg, about 6X 10 7 Individual cells/kg, about 7X 10 7 Individual cells/kg, about 8X 10 7 Individual cells/kg or about 9X 10 7 Individual cells/kg.
15. The method of claim 1, 2 or 9, wherein the therapeutically effective amount or effective dose of immune cells is between about 1 x 10 per kg body weight 6 And about 2X 10 6 Between immune cells up to about 1X 10 8 Maximum dose of individual immune cells.
16. The method of claim 1, 2 or 9, wherein the immune cells are in a therapeutically effective doseBetween 75X 10 6 And 200X 10 6 Between individual immune cells.
17. The method of any one of claims 1 to 16, wherein the tumor antigen is selected from the group consisting of a tumor-associated surface antigen, 5T4, alpha Fetoprotein (AFP), B7-1 (CD 80),
B7-2 (CD 86), BCMA, B-human chorionic gonadotrophin, CA-125, carcinoembryonic antigen (CEA), CD123, CD133, CD138, CD19, CD20, CD22,
CD23、CD24、CD25、CD30、CD33、CD34、CD4、CD40、
CD44, CD56, CD8, CLL-1, c-Met, CMV-specific antigen, CS-1,
CSPG4, CTLA-4, DLL3, bissialoganglioside GD2, ductal epithelial mucin, EBV specific antigen, EGFR variant III (EGFRvIII), ELF2M, endoglin, ephrin B2, epidermal Growth Factor Receptor (EGFR), epithelial cell adhesion molecule (EpCAM), epithelial tumor antigen, erbB2 (HER 2/neu), fibroblast-related protein (fap), FLT3, folate binding protein, GD2, GD3, glioma-related antigen, glycosphingolipids, gp36, HBV specific antigen, HCV specific antigen, HER1-HER2, a combination of HER2-HER3, HERV-K, high molecular weight melanoma-related antigen (HMW-MAA), HIV-1 envelope glycoprotein gp41,
HPV specific antigen, human telomerase reverse transcriptase, IGFI receptor, IGF-II, IL-11Rα, IL-13R-a2, influenza virus specific antigen; CD38, insulin growth factor (IGFl) -l, enterocarboxylesterase, kappa chain, LAGA-la, lambda chain, lassa virus-specific antigen, lectin-reactive AFP, lineage-specific or tissue-specific antigen such as
CD3, MAGE-A1, major Histocompatibility Complex (MHC) molecules presenting tumor specific peptide epitopes, M-CSF, melanoma associated antigens, mesothelin, MN-CA IX, MUC-1, mut hsp70-2, mutant p53, mutant ras, neutrophil elastase, NKG2D, nkp, NY-ESO-1, p53, PAP, prostase, prostate Specific Antigen (PSA), prostate cancer tumor antigen-1 (PCTA-1), prostate specific antigen protein, STEAP1, STEAP2, PSMA, RAGE-1, ROR1, RU2 (AS), surface adhesion molecules, survivin and telomerase, TAG-72, additional domain A (EDA) and additional domain B (EDB) of fibronectin, and Al domain of thrombopoietin-C (TnC Al), thyroxine, vascular growth factor, surface antigen (VEGFR-2), surface antigen specific antigen such AS HIV-specific antigen (VEGFR-20), or surface antigen-specific variants such AS any of these.
18. The method of claim 17, wherein the target antigen is CD19.
19. The method of claim 1 or 2, wherein the NHL is diffuse large B-cell lymphoma (DLBCL) non-specific, primary mediastinum large B-cell lymphoma, high grade B-cell lymphoma, or DLBCL caused by follicular lymphoma.
20. The method of any one of claims 1 to 19, further comprising preconditioning the subject with one or more preconditioning agents.
21. The method of claim 20, wherein the subject is preconditioned with administration of an alkylating agent and/or a platinum-based agent.
22. The method of claim 21, wherein the alkylating agent is selected from the group consisting of: melphalan, chlorambucil, cyclophosphamide, dichloromethyldiethylamine, nitrogen mustard (HN 2), uratemustine, uracil nitrogen mustard, melphalan, chlorambucil, ifosfamide, bendamustine, carmustine, lomustine, streptozotocin, alkyl sulfonates, busulfan, thiotepa, or the like, or any combination thereof.
23. The method of claim 21, wherein the platinum-based preconditioning agent is selected from the group consisting of: platinum, cisplatin, carboplatin, nedaplatin, oxaliplatin, satraplatin, triplatinum tetranitrate, procarbazine, altretamine, triazene, dacarbazine, mitozolomide, temozolomide, dacarbazine, temozolomide, and any combination thereof.
24. The method of claim 20, wherein the preconditioning agent comprises cyclophosphamide and fludarabine.
25. The method of any one of claims 20 to 24, wherein administration of the one or more preconditioning agents begins at least seven days, at least six days, at least five days, at least four days, at least three days, at least two days, or at least one day prior to administration of the cell therapy.
26. The method of claim 1, wherein the subject is administered allogeneic stem cell therapy (alloSCT) after treatment with the anti-CD 19 CAR T cell therapy.
27. The method of claim 1, wherein the subject has a tumor burden.
28. The method of claim 1, wherein tolizumab is administered only in the case of cytokine release syndrome for management of neurological events and/or beginning to use steroids for management of level 2 neurological events.
The following additional embodiments are illustrative of the present disclosure and are not limiting:
one embodiment of the present disclosure relates to a method of treating recurrent/refractory B precursor acute lymphoblastic leukemia in a subject, the method comprising administering to the subject a therapeutically effective amount of immune cells against a tumor antigen, wherein the subject is a pediatric or adolescent subject.
One embodiment of the present disclosure relates to the above method, wherein the therapeutically effective amount of immune cells is between about 1 x 10 per kg body weight 6 And about 2X 10 6 Between individual immune cells.
One embodiment of the present disclosure relates to the above method, wherein the immune cells are administered in a total volume of between about 40ml to 68 ml.
One embodiment of the present disclosure relates to the above method, wherein the immune cells are administered in a total volume of about 40 ml.
One embodiment of the present disclosure relates to the above method, wherein the therapeutically effective amount of the immune cells is about 1 x 10 per kg body weight 6 And (3) immune cells.
One embodiment of the present disclosure relates to the above method, wherein the immune cells are administered as a first line, a second line, a third line, a fourth line, a fifth line, or a sixth line therapy, or prior to disease progression.
One embodiment of the present disclosure is directed to the above method, wherein the tumor antigen is selected from the group consisting of tumor-associated surface antigen, 5T4, alpha Fetoprotein (AFP), B7-1 (CD 80), B7-2 (CD 86), BCMA, B-human chorionic gonadotrophin, CA-125, carcinoembryonic antigen (CEA), CD123, CD133, CD138, CD19, CD20, CD22, CD23, CD24, CD25, CD30, CD33, CD34, CD4, CD40, CD44, CD56, CD8, CLL-1, c-Met, CMV-specific antigen, CS-1, CSPG4, CTLA-4, DLL3, bisialoganglioside GD2, catheter epithelial mucin, EBV-specific antigen, EGFR variant III (EGFRvIII), ELF2M, endothelial glycoprotein, hepadin B2, epidermal Growth Factor Receptor (EGFR), epithelial cell adhesion molecule (EpCAM), epithelial tumor antigen, epF 2 (HER 2/neu), fibroblast-related protein (fap), FLT3, GD2, GD-3, GD-binding antigen, HEIL-3, HEIL-specific antigen, HIV-1, HIV-2, HIV-specific antigen, HIV-1, HIV-receptor, HIV-specific antigen; CD38, insulin growth factor (IGFl) -l, enterocarboxylesterase, kappa chain, LAGA-la, lambda chain, lasa-specific antigen, lectin-reactive AFP, lineage-specific or tissue-specific antigen such AS CD3, MAGE-A1, major Histocompatibility Complex (MHC) molecule, major Histocompatibility Complex (MHC) molecule presenting tumor-specific peptide epitopes, M-CSF, melanoma-associated antigen, mesothelin, MN-CA IX, MUC-1, mut hsp70-2, mutant p53, mutant ras, neutrophil elastase, NKG2D, nkp, NY-ESO-1, p53, PAP prostases, prostate Specific Antigen (PSA), prostate cancer tumor antigen-1 (PCTA-1), prostate specific antigen proteins, STEAP1, STEAP2, PSMA, RAGE-1, ROR1, RU2 (AS), surface adhesion molecules, survivin and telomerase, TAG-72, the Extra Domain A (EDA) and Extra Domain B (EDB) of fibronectin and the Al domain of tenascin-C (TnC Al), thyroglobulin, tumor matrix antigen, vascular endothelial growth factor receptor-2 (VEGFR 2), virus-specific surface antigens such AS HIV-specific antigens (such AS HIV gpl 20), and any derivatives or variants of these surface antigens.
One embodiment of the present disclosure relates to the above method, wherein the target antigen is CD19.
One embodiment of the present disclosure relates to the above method, further comprising preconditioning the subject with one or more preconditioning agents, wherein the one or more preconditioning agents are selected from at least one of an alkylating agent and a platinum-based agent, wherein the alkylating agent is selected from the group consisting of: melphalan, chlorambucil, cyclophosphamide, dichloromethyldiethylamine, nitrogen mustard (HN 2), uratemustine, uracil nitrogen mustard, melphalan, chlorambucil, ifosfamide, bendamustine, carmustine, lomustine, streptozotocin, alkyl sulfonates, busulfan, thiotepa or the like, and any combination thereof, and wherein the platinum-based preconditioning agent is selected from the group consisting of: platinum, cisplatin, carboplatin, nedaplatin, oxaliplatin, satraplatin, triplatinum tetranitrate, procarbazine, altretamine, triazene, dacarbazine, mitozolomide, temozolomide, dacarbazine, temozolomide, and any combination thereof.
One embodiment of the present disclosure relates to the above method, wherein the preconditioning agent comprises cyclophosphamide and fludarabine.
One embodiment of the present disclosure relates to the above method, wherein the cyclophosphamide is present at a concentration of between 200mg/m 2 Day and 2000mg/m 2 Dose between/day, and wherein the fludarabine is used at a dose of between 20mg/m 2 Day and 900mg/m 2 Dosage between/day.
One embodiment of the present disclosure relates to the above method, wherein said administration of said one or more preconditioning agents is initiated at least seven days, at least six days, at least five days, at least four days, at least three days, at least two days, or at least one day prior to administration of said immune cells.
One embodiment of the present disclosure relates to the above method, wherein the subject has a high tumor burden.
One embodiment of the present disclosure relates to the above method, comprising at least one of the following steps: tozucchini is administered only in the case of cytokine release syndrome for management of neurological events, and corticosteroids are administered for management of grade 2 neurological events.
One embodiment of the present disclosure relates to the above method, wherein the subject is at high risk of disease progression, wherein the subject is at high risk if the subject shows disease progression within 24 months after the initial diagnosis.
One embodiment of the present disclosure relates to the above method, wherein the immune cells are selected from Tumor Infiltrating Lymphocytes (TILs), NK cells, autologous T cells, allogeneic T cells, and engineered autologous T cells (eacts), and any combination thereof.
One embodiment of the present disclosure relates to the above method, wherein the immune cell is a CAR T cell.
One embodiment of the present disclosure relates to a method of treating cancer in a subject in need thereof, wherein the cancer is non-hodgkin's lymphoma (NHL) or relapsed/refractory B precursor acute lymphoblastic leukemia or relapsed/refractory B cell non-hodgkin's lymphoma (R/R B-ALL), the method comprising administering to the subject a therapeutically effective amount of immune cells that are anti-tumor antigens, and wherein the immune cells are autologous T cells that express the anti-CD 19 Chimeric Antigen Receptor (CAR).
One embodiment of the present disclosure relates to the above method, wherein the cancer is NHL and the NHL is Mantle Cell Lymphoma (MCL) or Indolent NHL (iNHL).
One embodiment of the present disclosure relates to the above method, wherein the iNHL is an edge zone lymphoma (MZL) or Follicular Lymphoma (FL).
One embodiment of the present disclosure relates to the above method, wherein the cancer is NHL, and the NHL is diffuse large B-cell lymphoma (DLBCL) non-specific finger, primary mediastinum large B-cell lymphoma, high grade B-cell lymphoma, or DLBCL caused by follicular lymphoma.
Drawings
FIG. 1. Design of clinical trial-2. A. Administration after leukapheresis and not less than 5 days before initiation of conditioning chemotherapy; PET-CT is required after bridging. b bone marrow biopsies are completed at screening and if positive, incomplete or if uncertain, biopsies are required to confirm CR. AE, adverse events; a CAR, chimeric antigen receptor; CR, complete reaction; CRS, cytokine release syndrome; IV, intravenous; MCL, mantle cell lymphoma; ORR, objective response rate; PO, oral administration; R/R, recurrent/refractory.
Figure 2. ORR in patients with and without MCL POD24 was assessed by IRRC. Evaluated by IRRC according to the rugano classification (Lugano Classification). 7a One patient was not rated. CR, complete reaction; IRRC, independent radiological review committee; ORR, objective response rate; PD, progressive disease; with POD24, disease progression after initial diagnosis<24 months; without POD24, disease progression is not less than 24 months after initial diagnosis; PR, partial reaction; SD, disease is stable.
FIGS. 3A, 3B and 3C show the duration of the reaction (DR) (3A), progression Free Survival (PFS) (3B) and Overall Survival (OS) (3C) according to the MCL POD24 status. a Patients with a response. DOR, duration of reaction; NE, not estimated; OS, overall survival; PFS, progression free survival; with POD24, disease progression after initial diagnosis<24 months; without POD24, disease progression was > 24 months after initial diagnosis.
Figure 4 CAR T-cell expansion in patients with and without MCL POD 24.
Fig. 5. B cells detectable over time in patients with and without MCL POD 24.
FIG. 6 design of clinical trial-5 clinical trial. a From the end of the last treatment>Patients with stable disease (no recurrence) for 1 year are not eligible. b The single anti-CD 20 antibody was not counted as a line number of treatments for eligibility. c Patients with evaluable efficacy include 80 or more subjectsThe FL patients treated, which were followed for no less than 18 months after axi-cel infusion, and MZL patients treated, which were followed for no less than 4 weeks after axi-cel infusion, until the date of data cutoff. Axi-cel, alemtujopsis; a CAR, chimeric antigen receptor; FL, follicular lymphoma; iNHL, indolent non-hodgkin's lymphoma; IV, intravenous; a mAb, monoclonal antibody; MZL, marginal zone lymphoma; POD24, disease progression from first receiving anti-CD 20 containing chemotherapy <24 months; R/R, recurrent/refractory.
Fig. 7. ORR in patients with and without iNHL POD24 was assessed by IRRC. Evaluation by IRRC was performed according to the rukino classification. (Cheson BD, et al J Clin Oncol.2014; 32:3059-68). a Of the 5 patients reported as ND, 4 (1 FL without POD24;3 MZL) were disease-free according to IRRC at baseline and after baseline, but the investigator considered disease; 1 patient with FL and POD24 died prior to the first disease assessment. CR, complete reaction; FL, follicular lymphoma; IRRC, independent radiological review committee; MZL, marginal zone lymphoma; ND, incomplete/undefined; ORR, overall reaction rate; iNHL POD24, disease progression from first receiving anti-CD 20 containing chemotherapy<24 months; PD, progressive disease; PR, partial reaction; SD, disease is stable.
FIGS. 8A, 8B and 8C DOR (8A), PFS (8B) and OS (8C) in the iNHL POD24 state. DOR, duration of reaction; FL, follicular lymphoma; mo, month; MZL, marginal zone lymphoma; NE, not estimated; NR, not reached; OS, overall survival; PFS, progression free survival; POD24, disease progression <24 months from the first receipt of anti-CD 20 containing chemotherapy.
Fig. 9A, 9B and 9C CAR T-cell expansion (9A and 9B) and pre-treatment serum analyte (9C) in patients with FL in iNHL POD24 status. P-values were calculated using Wilcoxon rank sum test. a Data from the first 2 patients with FL were not available for retreatment. AUC (AUC) 0-28 Area under the curve between day 0 and day 28; a CAR, chimeric antigen receptor; CCL, chemokine (C-C motif) ligand; FL, follicular lymphoma; LOQ, limit of quantitation; MDC, macrophage-derived chemokine; POD24, from the first timeChemotherapy with anti-CD 20 contributes to disease progression<24 months; TARC, thymus and activation regulating chemokines;
FIG. 10A, FIG. 10B, FIG. 10C, FIG. 10D and FIG. 10E. Correlation between AUC (10A), response (10B), MRD rate (10C) and toxicity (10D and 10E) of CAR gene copy number/. Mu.g DNA.
Fig. 11 additional cytokine and inflammatory marker levels over time. CRP, C-reactive protein; CXCL10, C-X-C motif chemokine ligand 10; GM-CSF, granulocyte-macrophage colony stimulating factor; IL, interleukin; MCP, monocyte attractant protein; rα, receptor α; RA, receptor antagonists; SAA, serum amyloid A; VCAM, vascular cell adhesion molecule.
Fig. 12. Subgroup analysis of overall remission rate. BM, bone marrow; ORR, overall remission rate; SCT, stem cell transplantation.
Detailed Description
Definition of the definition
For easier understanding of the present disclosure, certain terms are first defined below. Additional definitions of the following terms and other terms are set forth throughout the specification.
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.
As used herein, the term "or" is understood to be inclusive and to encompass both "or" and "unless specified otherwise or apparent from the context.
The term "and/or" as used herein shall be taken to mean a specific disclosure of each of the two specified features or components, with or without the other. Thus, the term "and/or" as used in phrases such as "a and/or B" herein is intended to include a and B; a or B; a (alone); and B (alone). Also, the term "and/or" as used in phrases such as "A, B and/or C" is intended to encompass each of the following aspects: 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).
As used herein, the terms "for example" and "i.e." are used by way of example only, are not intended to be limiting, and should not be construed to refer to only those items explicitly recited in the specification.
The terms "or more," "at least," "exceeding," etc. such as "at least one" are understood to include, 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, 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, or 150, 200, 300, 400, 500, 600, 700, 800, 900, 1000, 2000, 3000, 4000, 5000, or more than the value. But also any larger numbers or scores therebetween.
Conversely, the term "no more than" includes every value that is less than the recited value. For example, "no more than 100 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, 53, 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. But also any smaller numbers or scores therebetween.
The terms "plurality," "at least two," "two or more," "at least a second," and the like are understood to include, but are not limited to, at least 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, or 150, 200, 300, 400, 500, 600, 700, 800, 900, 1000, 2000, 3000, 4000, 5000, or more. But also any larger numbers or scores therebetween.
Throughout this specification the word "comprise", or variations such as "comprises" or "comprising", will be understood to imply 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. It will be understood that wherever aspects are described herein in the language "comprising," other similar aspects are also provided as described in terms of "consisting of and/or" consisting essentially of. The term "consisting of … …" excludes any element, step or component not specified in the claims. Relates to Gray,53F.2d 520,11USPQ 255 (CCPA 1931); one-sided Davis,80uspq 448,450 (bd.app.1948) ("consisting of … …" is defined as "closed claims to include materials other than those recited except for impurities normally associated therewith"). The term "consisting essentially of … …" limits the scope of the claims to the specified materials or steps as well as those materials or steps that do not materially affect the basic and novel characteristics of the claimed invention.
Unless specifically stated or apparent from the context, as used herein, the term "about" refers to a value or composition that is within an acceptable error range for the particular value or composition as determined by one of ordinary skill in the art, which will depend in part on how the value or composition is measured or determined, i.e., the limitations of the measurement system. For example, in accordance with the practice in the art, "about" or "approximately" may mean within one or more than one standard deviation. "about" or "approximately" may mean a range of up to 10% (i.e., ±10%). Thus, "about" may be understood to be within 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, 0.5%, 0.1%, 0.05%, 0.01% or 0.001% of the stated value. For example, about 5mg may include any amount between 4.5mg and 5.5 mg. Furthermore, these terms may mean at most one order of magnitude or at most 5 times a certain value, especially for biological systems or processes. When a particular value or composition is provided in this disclosure, unless otherwise indicated, the meaning of "about" or "approximately" should be assumed to be within an acceptable error range for the particular value or composition.
As described herein, any concentration range, percentage range, ratio range, or integer range should be understood to include the value of any integer within the range, as well as fractions thereof (such as tenths and hundredths of integers) as appropriate, unless otherwise indicated.
Units, prefixes, and symbols as used herein are provided using forms accepted by their international system of units (SI). The numerical range includes the numbers defining the range.
Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure pertains. For example, juo, "simplified dictionary of biomedical and molecular biology (The Concise Dictionary of Biomedicine and Molecular Biology)", 2 nd edition, 2001, CRC Press (CRC Press); "dictionary of cell and molecular biology (The Dictionary of Cell & Molecular Biology)", 5 th edition, 2013, academic Press (Academic Press); and "oxford biochemical and molecular biological dictionary (The Oxford Dictionary Of Biochemistry And Molecular Biology)", by emmack et al, 2 nd edition, 2006, oxford university press (Oxford University Press) provides one of skill in the art with a general dictionary of many terms used in this disclosure.
"administering" refers to physically introducing an agent into a subject using any of a variety of methods and delivery systems known to those of skill in the art. Exemplary routes of administration for the formulations disclosed herein include intravenous, intramuscular, subcutaneous, intraperitoneal, spinal, or other parenteral routes of administration (e.g., by injection or infusion). Exemplary routes of administration for the compositions disclosed herein include intravenous, intramuscular, subcutaneous, intraperitoneal, spinal, or other parenteral routes of administration (e.g., by injection or infusion). As used herein, the phrase "parenteral administration" means modes of administration other than enteral and topical administration (typically by injection), and includes, but is not limited to, intravenous, intramuscular, intraarterial, intrathecal, intralymphatic, intralesional, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous, subcuticular, intra-articular, subcapsular, subarachnoid, intraspinal, epidural and intrasternal injection and infusion, and in vivo electroporation. In some embodiments, the formulation is administered by a non-parenteral route (e.g., orally). Other non-parenteral routes include topical, epidermal or mucosal routes of administration, such as intranasal, vaginal, rectal, sublingual or topical. Administration may also be performed, for example, once, multiple times, and/or over one or more extended periods of time. In one embodiment, the CAR T cell therapy is administered via an "infusion product" comprising CAR T cells.
The term "antibody" (Ab) includes, but is not limited to, glycoprotein immunoglobulins that specifically bind to an antigen. In general, an antibody may comprise at least two heavy (H) chains and two light (L) chains, or antigen binding molecules thereof, that are interconnected by disulfide bonds. Each H chain comprises a heavy chain variable region (abbreviated herein as VH) and a heavy chain constant region. The heavy chain constant region comprises three constant domains, CH1, CH2 and CH3. Each light chain comprises a light chain variable region (abbreviated herein as VL) and a light chain constant region. The light chain constant region comprises one constant domain CL. VH and VL regions can be further subdivided into regions of higher variability, termed Complementarity Determining Regions (CDRs), interspersed with regions that are more conserved, termed Framework Regions (FR). Each VH and VL comprises 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 comprise binding domains that interact with antigens. The constant region of an Ab may mediate binding of an immunoglobulin to host tissues or factors including various cells of the immune system (e.g., effector cells) and the first component of the classical complement system (C1 q).
Antibodies can 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 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, endosomes, antibody fusions (sometimes referred to herein as "antibody conjugates"), heteroconjugate antibodies, single domain antibodies, monovalent antibodies, single chain antibodies, or single chain Fv (scFv), camelized antibodies, affibodies, fab fragments, F (ab') 2 fragments, disulfide-linked Fv (sdFv), anti-idiotype (anti-Id) antibodies (including, for example, anti-Id antibodies), miniantibodies, domain antibodies, synthetic antibodies (sometimes referred to herein as "antibody mimetics"), and antigen binding fragments of any of the above. In some embodiments, the antibodies described herein refer to a population of polyclonal antibodies.
An "antigen binding molecule," "antigen binding portion," or "antibody fragment" refers to any molecule that comprises an antigen binding portion (e.g., CDR) of an antibody from which the molecule is derived. The antigen binding molecule may comprise an antigen Complementarity Determining Region (CDR). Examples of antibody fragments include, but are not limited to, fab ', F (ab') 2, and Fv fragments, dabs, linear antibodies, scFv antibodies, and multispecific antibodies formed from antigen-binding molecules. A peptibody (i.e., an Fc fusion molecule comprising a peptide binding domain) is 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 or viral or bacterial antigen on a cell involved in a hyperproliferative disease. In some embodiments, the antigen binding molecule binds CD19. In further embodiments, the antigen binding molecule is an antibody fragment that specifically binds an antigen, including one or more Complementarity Determining Regions (CDRs) thereof. 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 a high affinity polymer (avimer).
An "antigen" refers to any molecule that causes an immune response or is capable of being bound by an antibody or antigen binding molecule. The immune response may involve antibody production, or activation of specific immunocompetent cells, or both. Those skilled in the art will readily appreciate that any macromolecule (including almost all proteins or peptides) may be used as an antigen. The antigen may be expressed endogenously, i.e. by genomic DNA, or may be expressed recombinantly. The antigen may be specific for a tissue (such as a cancer cell), or it may be broadly expressed. In addition, fragments of larger molecules may act as antigens. In some embodiments, the antigen is a tumor antigen.
The term "neutralizing" refers to an antigen binding molecule, scFv, antibody or fragment thereof that binds a ligand and prevents or reduces the biological effect of the ligand. In some embodiments, the antigen binding molecule, scFv, antibody, or fragment thereof directly blocks the binding site on the ligand, or alters the binding capacity of the ligand by indirect means (e.g., structural or energy changes in the ligand). In some embodiments, the antigen binding molecule, scFv, antibody, or fragment thereof prevents the protein to which it binds from performing a biological function.
The term "autologous" refers to any material derived from the same individual and later reintroduced into that individual. For example, the engineered autologous cell therapies (eACT TM ) The method involves collecting lymphocytes from a patient, then engineering them to express, for example, a CAR construct, and then administering back to the same patient.
The term "allogeneic" refers to any material derived from one individual and subsequently introduced into another individual of the same species, such as allogeneic T cell transplantation.
In one embodiment, the CAR T cell therapy comprises "alemtuquor therapy". "treatment with Alkylrensaine" was performed as a 2X 10 single infusion 6 Target dose of anti-CD 19CAR T cells/kg intravenously administered anti-CD 19CAR transduced autologous T cells. For subjects with a body weight greater than 100kg, 2X 10 can be administered 8 Maximum plateau dose of individual anti-CD 19CAR T cells. anti-CD 19CAR T cells are autologous human T cells that have been engineered to express an extracellular single chain variable fragment (scFv) with specificity for CD19 linked to an intracellular signaling moiety consisting of signaling domains from CD28 and CD3 zeta (CD 3-zeta) molecules arranged in tandem, anti-CD 19CAR vector constructs have been designed, optimized and initially tested at the surgical branch of the national cancer institute (NCI, IND 13871) (Kochenderfer et al, J immunother.2009;32 (7): 689-702; kochenderfer et al, blood.2010;116 (19): 3875-86). The scFv is derived from the variable region of the anti-CD 19 monoclonal antibody FMC63 (Nicholson et al, molecular immunology 1997;34 (16-17): 1157-65). A portion of the CD28 co-stimulatory molecule was added because the murine model indicated that this is important for anti-tumor effect and persistence of anti-CD 19CAR T cells (Kowolik et al, cancer Res.2006;66 (22): 10995-1004). The signaling domain of the CD3- ζ chain is used for T cell activation. These fragments were cloned into murine stem cell virus (MSGV 1) based vectors for genetic engineering of autologous T cells. The CAR construct is transduced into the genome of T cells by a retroviral vector. Briefly, peripheral Blood Mononuclear Cells (PBMCs) were obtained by leukocyte apheresis and Ficoll isolation. Peripheral blood mononuclear cells are activated by incubation with an anti-CD 3 antibody in the presence of recombinant interleukin 2 (IL-2). The stimulated cells were transduced with a retroviral vector containing the anti-CD 19CAR gene and proliferated in culture to produce enough engineered T cells for administration. The alopecie is a subject specific product.
In a real worldIn embodiments, CAR T cell therapy comprises KTE-X19, an autologous anti-CD 19 Chimeric Antigen Receptor (CAR) T cell therapy approved in the united states and the european union for the treatment of relapsed/refractory (R/R) MCL. ((brexucabtagene autoleucel) prescription information Kite Pharma, inc;2021;Summary of product characteristics (autologous anti-CD 19 transduced cd3+ cells) Kite Pharma EU b.v.; 2021). The method of manufacture of KTE-X19 was modified with respect to the method of manufacture of alemtujopsis to enrich CD4 positively + /CD8 + Cells remove circulating lymphoma cells.
In one embodiment, the product is characterized by a cellular composition. Cells can be labeled with fluorescent conjugated antibodies to CD3 (pan T cell marker), CD14, CD19 (B cell marker), CD45 (pan white cell marker) and CD56 (activation and NK markers) and evaluated by flow cytometry. Cell viability may be assessed using negative staining with a viability dye (SYTOX near infrared). The lower limit of quantification (LLOQ) of the assay may be 0.2% and 5% for NK cells and monocytes. The percentage of NK cells (NK cells CD 45) + 、CD14 - 、CD3 - And CD56 + The method comprises the steps of carrying out a first treatment on the surface of the T cell is CD45 + 、CD14 - And CD3 - ). The median percentages of NK cells from 23 batches of alopecie and 97 batches of KTE-X19 can be 1.9% (range 0.8% -3.2%) and 0.1% (range 0.0% -2.8%), respectively. CD3 from the same batch of Alkylrensaine and KTE-X19 - The median percentages of cellular impurities may be 2.4% (range 0.9% -4.6%) and 0.5% (range 0.3% -3.9%), respectively. The results of KTE-X19 (brexucabtagene autoleucel, TECARTUS) and Alkerese (YESCARTA) in terms of cell viability may be greater than or equal to 72% and greater than or equal to 80%, respectively; the results in terms of anti-CD 19 CAR expression can be 24% or more and 15% or more, respectively; the results in terms of IFN-gamma production may be ≡190pg/mL and ≡520pg/mL, respectively; in CD3 + The results in terms of percentage of cells can be respectivelyMore than or equal to 90 percent and more than or equal to 85 percent. Brexucabtagene autoleucel may consist essentially of CD3+ T cells (99.3% + -0.8%), which may be further described as CD4+ (37.9% + -16.5%) and CD8+ (59.3% + -16.5%) subsets.
The terms "transduction" and "transduced" refer to the process of introducing exogenous DNA into a cell by a viral vector (see Jones et al, "Genetics: principles and analysis," Boston: jones & Bartlett publication (1998)). In some embodiments, the vector is a retroviral vector, a DNA vector, an RNA vector, an adenovirus vector, a baculovirus vector, an Epstein-Barr virus vector, a papilloma virus vector, a vaccinia virus vector, a herpes simplex virus vector, an adenovirus-associated vector, a lentiviral vector, or any combination thereof.
"cancer" refers to a broad group of various diseases characterized by uncontrolled growth of abnormal cells in the body. Unregulated cell division and growth results in the formation of malignant tumors that invade adjacent tissues and can also metastasize to distal parts of the body through the lymphatic system or blood flow. "cancer" or "cancerous tissue" may include tumors. In this application, the term cancer is synonymous with malignant tumor. 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 leukocyte malignancies. In some embodiments, the methods disclosed herein can be used to reduce tumor size derived from, for example, tumors such as: bone cancer, pancreatic cancer, skin cancer, head and neck cancer, cutaneous or intraocular malignant melanoma, uterine cancer, ovarian cancer, rectal cancer, anal region cancer, gastric cancer, testicular cancer, uterine cancer, fallopian tube cancer, 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, endocrine system cancer, thyroid cancer, parathyroid cancer, adrenal gland cancer, soft tissue sarcoma, urinary tract cancer, penile cancer, chronic or acute leukemia, acute myelogenous leukemia, chronic myelogenous leukemia, acute Lymphocytic Leukemia (ALL) (including non-T-cell ALL), chronic Lymphocytic Leukemia (CLL), childhood solid tumors, lymphocytic lymphomas, bladder cancer, kidney or ureter cancer, renal pelvis cancer, central Nervous System (CNS) tumors, primary CNS lymphomas, tumor angiogenesis, spinal cord axis tumors, brain stem gliomas, pituitary adenomas, kaposi's sarcoma, epidermoid carcinoma, squamous cell carcinoma, T cell lymphomas, environmentally induced cancers (including those caused by asbestos), other B cell malignancies, and combinations thereof. In some embodiments, the cancer is multiple myeloma. In some embodiments, the cancer is NHL. A particular cancer may be responsive to chemotherapy or radiation therapy, or the cancer may be refractory. Refractory cancer refers to a cancer that is not amenable to surgical intervention and that is initially unresponsive to chemotherapy or radiation therapy, or that becomes unresponsive over time.
As used herein, "anti-tumor effect" refers to a biological effect that may be manifested 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 or progression-free survival, an increase in life expectancy, or an improvement in various physiological symptoms associated with a tumor. Antitumor effect may also refer to the prevention of the occurrence of tumors, such as vaccines.
As used herein, "cytokine" refers to a non-antibody protein released by one cell in response to contact with a specific antigen, wherein the cytokine interacts with a second cell to mediate a response in the second cell. As used herein, "cytokine" refers to a protein released by one cell population that acts as an intercellular mediator on another cell. Cytokines may be expressed endogenously by the cells or administered to the subject. Cytokines can be released by immune cells (including macrophages, B cells, T cells, and mast cells) to spread the immune response. Cytokines can induce various responses in the recipient cells. Cytokines may include homeostatic cytokines, chemokines, pro-inflammatory cytokines, effectors, and acute phase proteins. For example, steady state cytokines, including Interleukins (IL) 7 and IL-15, promote immune cell survival and proliferation, and pro-inflammatory 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 pro-inflammatory 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 cell 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).
A "chemokine" is a cytokine that mediates chemotaxis or directed movement of cells. Examples of chemokines include, but are not limited to, IL-8, IL-16, eosinophil-activating chemokine-3, macrophage-derived chemokine (MDC or CCL 22), monocyte-chemotactic protein 1 (MCP-1 or CCL 2), MCP-4, macrophage inflammatory protein 1 alpha (MIP-1 alpha, MIP-1 a), MIP-1 beta (MIP-1 b), gamma-inducible protein 10 (IP-10), and thymus activation-regulating chemokine (TARC or CCL 17).
As used herein, "chimeric receptor" refers to an engineered surface expression molecule capable of recognizing a particular molecule. Chimeric Antigen Receptors (CARs) and engineered T Cell Receptors (TCRs) comprising a binding domain capable of interacting with a particular tumor antigen allow T cells to target and kill cancer cells expressing the particular tumor antigen. In one embodiment, T cell therapy is based on T cells engineered to express a Chimeric Antigen Receptor (CAR) or a T Cell Receptor (TCR) comprising (i) an antigen binding molecule, (ii) a co-stimulatory domain, and (iii) an activation domain. The co-stimulatory domain may comprise an extracellular domain, a transmembrane domain and an intracellular domain, wherein the extracellular domain comprises a hinge domain that may be truncated.
A "therapeutically effective amount," "effective dose," "effective amount," or "therapeutically effective dose" of a therapeutic agent (e.g., an engineered CAR T cell, a small molecule, "agent" described in the specification) is any amount that, when used alone or in combination with another therapeutic agent, protects a subject from onset of a disease or promotes regression of a disease (as evidenced by a reduction in the severity of disease symptoms, an increase in the frequency and duration of asymptomatic disease periods, or prevention of a disorder or disability resulting from the disease). Such terms may be used interchangeably. The ability of a therapeutic agent to promote disease regression can be assessed using a variety of methods known to the skilled artisan, such as in human subjects during clinical trials, in animal model systems that predict efficacy in humans, or by assaying the activity of the agent in an in vitro assay. The therapeutically effective amount and dosage regimen can be determined empirically by testing in known in vitro or in vivo (e.g., animal model) systems.
The term "combination" refers to a fixed combination in the form of a dosage unit, or a combined administration, wherein the compounds of the present disclosure and the combination partner (e.g., another drug as explained below, also referred to as a "therapeutic agent" or "agent") may be administered simultaneously, independently, or separately over time intervals, particularly where these time intervals allow the combination partners to exhibit a synergistic (e.g., synergistic) effect. The individual components may be packaged in a kit or individually. One or both components (e.g., powder or liquid) may be reconstituted or diluted to the desired dosage prior to administration. As used herein, the terms "co-administration" or "combined administration" and the like are intended to encompass administration of the selected combination partners to a single subject (e.g., patient) in need thereof, and are intended to include treatment regimens in which the agents do not have to be administered by the same route of administration or simultaneously.
The term "product" or "infusion product" is used interchangeably herein and refers to a T cell composition administered to a subject in need thereof. Typically, in CAR T cell therapy, the T cell composition is administered as an infusion product.
As used herein, the term "lymphocyte" includes Natural Killer (NK) cells, T cells, or B cells. NK cells are a cytotoxic (cytotoxic) lymphocyte that represents a major component of the innate immune system. NK cells exclude tumor and virus-infected cells. It acts through processes of apoptosis or programmed cell death. They are 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 (TCRs) distinguish themselves from other lymphocyte types. Thymus is a specialized organ of the immune system, mainly responsible for the maturation of T cells. T cells are of six types, namely: helper T cells (e.g., cd4+ cells), cytotoxic T cells (also known as TC, cytotoxic T lymphocytes, CTLs, T killer cells, cytolytic T cells, cd8+ T cells, or killer T cells), memory T cells ((i) stem memory TSCM cells (e.g., naive cells) are CD45RO-, ccr7+, cd45ra+, cd62l+ (L-selectin), cd27+, cd28+, and IL-7rα+, but they also express a large number of CD95, IL-2rβ, CXCR3, and LFA-1, and exhibit many functional attributes specific to memory cells); (ii) Central memory TCM cells express L-selectin and CCR7, which secrete IL-2 but not ifnγ or IL-4, and (iii) however, effector memory TEM cells 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 γδ T cells. B cells, on the other hand, play a major role in humoral immunity (with antibody involvement). It produces antibodies and antigens and functions as an Antigen Presenting Cell (APC) and is transformed into memory B cells after activation by antigen interaction. In mammals, immature B cells are formed in bone marrow from which their name is derived.
In the context of the present disclosure, the terms "TN", "T naive" and ccr7+cd45ra+ actually refer to cells that are more like stem-like memory cells than typical naive T cells. Thus, in the examples and claims, T is as follows N All references to (c) refer to cells that are experimentally selected by their characterization as ccr7+cd45ra+ cells only, and should be so interpreted. They are better named in the context of the present disclosure as stem-like memory cells, but they should be called ccr7+cd45ra+ cells. Further characterization of the stem-like memory cells can be carried out, for example, using Arihara Y, jacobsen CA, armand P et al Journal for ImmunoTherapy of cancer.2019;7 (1) the process described in P210.
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 region or non-coding region or a portion thereof, or inserting a coding region or a portion thereof. In some embodiments, the modified cell is a lymphocyte, such as a T cell, which can be obtained from a patient or donor. The cells can be modified to express an exogenous construct, such as a Chimeric Antigen Receptor (CAR) or T Cell Receptor (TCR), that is incorporated into the cell genome.
By "immune response" is meant 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 produced by any of these cells or the liver, including abs, cytokines, and complement, resulting in selective targeting, binding, damage, destruction, and/or elimination of invasive pathogens from the vertebrate body, pathogen-infected cells or tissues, cancer cells or other abnormal cells, or normal human cells or tissues in the case of autoimmune or pathological inflammation.
The term "immunotherapy" refers to the treatment of a subject suffering from a disease or at risk of developing a disease or recurrence by a method comprising inducing, enhancing, suppressing, or otherwise altering an immune response. Examples of immunotherapy include, but are not limited to, T cell therapy. T cell therapies may include adoptive T cell therapy, tumor Infiltrating Lymphocyte (TIL) immunotherapy, autologous cell therapy, engineered autologous cell therapy (eACT) TM ) And allogeneic T cell transplantation. However, one of skill in the art will recognize that the conditioning methods disclosed herein will enhance the efficacy of any transplanted T cell therapy. Examples of T cell therapies are described in U.S. patent publications 2014/0154228 and 2002/0006409, U.S. patent 7,741,465, U.S. patent 6,319,494, U.S. patent 5,728,388 and international publication WO 2008/081035. In some embodiments, the immunotherapy comprises CAR T cell therapy. In some embodiments, the CAR T cell therapy product is via Infusion and administration.
The T cells for immunotherapy may be from any source known in the art. For example, T cells may be differentiated from a population of hematopoietic stem cells in vitro, or may be obtained from a subject. T cells may be obtained, for example, from Peripheral Blood Mononuclear Cells (PBMCs), bone marrow, lymph node tissue, cord blood, thymus tissue, tissue from an infection site, ascites, pleural effusion, spleen tissue, and tumors. In addition, T cells may be derived from one or more T cell lines available in the art. Various techniques known to the skilled artisan (such as FICOLL TM Isolation and/or apheresis) to obtain T cells from a blood unit collected from a subject. Additional methods for isolating T cells for T cell therapy are disclosed in U.S. patent publication 2013/0287748, which is incorporated herein by reference in its entirety.
The term "engineered autologous cell therapy" or "eACT TM "(also known as adoptive cell transfer) is a process by which T cells of the patient themselves are collected and then these cells are genetically engineered to recognize and target one or more antigens expressed on the cell surface of one or more specific tumor cells or malignant tumors. T cells can be engineered to express, for example, chimeric Antigen Receptors (CARs). CAR positive (+) T cells are engineered to express extracellular single chain variable region fragments (scFv) specific for a particular tumor antigen, linked to an intracellular signaling portion comprising at least one costimulatory domain and at least one activation domain. The CAR scFv can be designed to target, for example, CD19 being a transmembrane protein expressed by cells in the B cell lineage, including ALL normal B cells and B cell malignancies, including but not limited to Diffuse Large B Cell Lymphoma (DLBCL) non-specific, primary mediastinum large B cell lymphoma, high grade B cell lymphoma, and DLBCL, NHL, CLL and non-T cell ALL caused by follicular lymphoma. Exemplary CAR T cell therapies and constructs are described in U.S. patent publications 2013/0287748, 2014/0227237, 2014/0099309, and 2014/0050708, and these references are incorporated by reference in their entirety.
As used herein, "patient" or "subject" includes any person suffering from cancer (e.g., lymphoma or leukemia). The terms "subject" and "patient" are used interchangeably herein.
As used herein, the term "in vitro cell" refers to any cell that is cultured ex vivo. In particular, the in vitro cells may comprise T cells. The term "in vivo" refers to being within a patient.
The terms "peptide," "polypeptide," and "protein" are used interchangeably and refer to a compound consisting of amino acid residues covalently linked by peptide bonds. The protein or peptide contains at least two amino acids, and there is no limitation on the maximum number of amino acids that can constitute the protein or peptide sequence. Polypeptides include any peptide or protein comprising two or more amino acids linked to each other by peptide bonds. As used herein, the term refers to both short chains (which are also commonly referred to in the art as, for example, peptides, oligopeptides, and oligomers) and long chains (which are commonly referred to in the art as proteins, many types of proteins). "Polypeptides" include, for example, biologically active fragments, substantially homologous polypeptides, oligopeptides, homodimers, heterodimers, variants of polypeptides, modified polypeptides, derivatives, analogs, fusion proteins, and the like. The polypeptides include natural peptides, recombinant peptides, synthetic peptides, or combinations thereof.
As used herein, "stimulation" refers to a primary response induced by the binding of a stimulatory molecule to its cognate ligand, wherein the binding mediates a signaling event. A "stimulatory molecule" is a molecule on a T cell (e.g., a T Cell Receptor (TCR)/CD 3 complex) that specifically binds to a cognate stimulatory ligand presented on an antigen presenting cell. A "stimulatory ligand" is a ligand that, when presented on an antigen presenting cell (e.g., APC, dendritic cell, B cell, etc.), specifically binds to a stimulatory molecule on a T cell, thereby mediating a primary response of the T cell (including, but not limited to, activation, initiation of an immune response, proliferation, etc.). Stimulating ligands include, but are not limited to, anti-CD 3 antibodies, peptide-loaded MHC class I molecules, superagonist anti-CD 2 antibodies, and superagonist anti-CD 28 antibodies.
As used herein, a "co-stimulatory signal" refers to a signal that, in combination with a primary signal such as a TCR/CD3 linkage, causes a T cell response (such as, but not limited to, proliferation and/or up-or down-regulation of a key molecule).
As used herein, a "costimulatory ligand" includes a molecule on an antigen presenting cell that specifically binds to a cognate costimulatory molecule on a T cell. Binding of the costimulatory ligand provides a signal that mediates T cell responses (including but not limited to proliferation, activation, differentiation, etc.). The co-stimulatory ligand induces a signal other than the primary signal provided by the stimulatory molecule, e.g., through binding of the T Cell Receptor (TCR)/CD 3 complex to the Major Histocompatibility Complex (MHC) molecule loaded with the peptide. Co-stimulatory ligands may include, but are not limited to, 3/TR6, 4-1BB ligand, agonists or antibodies that bind to Toll ligand receptors, B7-1 (CD 80), B7-2 (CD 86), CD30 ligand, CD40, CD7, CD70, CD83, herpes virus invasion mediator (HVEM), human leukocyte antigen G (HLA-G), ILT4, immunoglobulin-like transcript (ILT) 3, inducible co-stimulatory ligand (ICOS-L), intercellular adhesion molecule (ICAM), ligand that specifically binds to B7-H3, lymphotoxin beta receptor, MHC class I chain-associated protein A (MICA), MHC class I chain-associated protein B (MICB), OX40 ligand, PD-L2, or Programmed Death (PD) L1. In certain embodiments, the costimulatory ligand comprises, but is not limited to, an antibody that specifically binds to a costimulatory molecule present on a T cell, 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 (NKG 2C), OX40, PD-1, or tumor necrosis factor superfamily member 14 (TNFSF 14 or LIGHT).
A "costimulatory molecule" is a cognate binding partner on a T cell that specifically binds to a costimulatory ligand, thereby mediating a costimulatory response of 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 (SLAMF 8), BTLA, CD33, CD45, CD100 (SEMA 4D), CD103, CD134, CD137, CD154, CD16, CD160 (BY 55), CD18, CD19 ase:Sub>A, CD2, CD22, CD247, CD27, CD276 (B7-H3), CD28, CD29, CD3 (α; betase:Sub>A; deltase:Sub>A; ε,. Gammase:Sub>A.; zetase:Sub>A.), CD30, CD37, CD4, CD40, CD49 ase:Sub>A, CD49D, CD f, CD5, CD64, CD69, CD7, CD80, CD83 ligand, CD84, CD86, CD8 alphase:Sub>A, CD8 betase:Sub>A, CD9, CD96 (Tactive), CD11 ase:Sub>A, CD11B, CD11c, CD11D, CDS, CEACAM1, CRT AM, DAP-10, DNAM1 (CD 226), fc gammase:Sub>A receptor, GADS, GITR, HVEM (LIGHTR), IA4, ICAM-1, ICOS, ig alphase:Sub>A (CD 79 ase:Sub>A), IL2 Rbetase:Sub>A, IL2 Rgammase:Sub>A, IL7 Ralphase:Sub>A, integrin, ITGA4, ITGA6, ITGAD, ITGAE, ITGAL, ITGAM, ITGAX, ITGB2, ITGB7, ITGBL, KIRDS2, LAT, LFA-1, LIGHT (tumor necrosis factor superfamily member 14; TNFSF 14), LTBR, ly9 (CD 229), lymphocyte function associated antigen-1 (LFA-1 (CD 11 ase:Sub>A/CD 18), MHC 2, SL35G 2, SL35F 1, SL35F 6, SL35F, SL35, SLF, SL35, SLF-6, SL35, SLF, SLSL35, SLF-6, SLSLSL35, SL35, SLF-6, SLSLSLSLF-6, SLSLSLSLSL35, or combination thereof.
The terms "decrease" and "decrease" are used interchangeably herein and indicate any change that is less than the original value. "decrease" and "decrease" are relative terms that require a comparison between before and after measurement. "decrease" and "decrease" include complete depletion. Similarly, the term "increase" means any change above the original value. "increasing", "higher" and "lower" are relative terms that require comparison between before and after measurement and/or between reference standards. In some embodiments, the reference value is obtained from a value of a general population, which may be a general patient population. In some embodiments, the reference value is from a quartile analysis of a general patient population.
"treatment" of a subject refers to any type of intervention or procedure performed on the subject, or administration of an active agent to the subject, with the purpose of reversing, alleviating, ameliorating, inhibiting, slowing or preventing the onset, progression, development, severity or recurrence of symptoms, complications or disorders associated with the disease, or biochemical indicators. In some embodiments, "treating" includes partial remission. In another embodiment, "treating" or "treatment" includes complete remission. In some embodiments, the treatment may be prophylactic, in which case the treatment is administered prior to any symptoms of the disorder being observed. As used herein, the term "preventing" refers to the prevention or protective treatment of a disease or condition. Prevention of a symptom, disease, or condition may include, for example, reducing (e.g., alleviating) one or more symptoms of the disease or condition relative to a reference level (e.g., symptoms in a similar subject to which no treatment was administered). Prevention may also include delaying the onset of one or more symptoms of the disease or disease state, for example, relative to a reference level (e.g., onset of symptoms in a similar subject to which no treatment was administered). In embodiments, the disease is a disease described herein. In some embodiments, the disease is cancer. In some embodiments, the disease state is CRS or neurotoxicity. In some embodiments, the indicator of improved or successful treatment comprises determining that the associated score on a toxicity grading scale (e.g., CRS or neurotoxicity grading scale) is not exhibited, such as a score of less than 3, or a change in grading or severity on a grading scale as discussed herein, such as a change from score 4 to score 3, or a change from score 4 to score 2, 1, or 0.
As used herein, the term "multifunctional T cell" refers to a cell that co-secretes at least two proteins from a pre-specified panel of each cell, coupled with the amount of each protein produced (i.e., the number of proteins secreted and the combination of intensity at which). In some embodiments, a single cell functional profile is determined for each evaluable population of engineered T cells. The spectra can be classified into the groups of effectors (granzymes B, IFN-gamma, MIP-1 alpha, perforins, TNF-alpha, TNF-beta), stimuli (GM-CSF, IL-2, IL-5, IL-7, IL-8, IL-9, IL-12, IL-15, IL-21), modulators (IL-4, IL-10, IL-13, IL-22, TGF-beta 1, sCD137, sCD 40L), chemoattractions (CCL-11, IP-10, MIP-1 beta, RANTES) and inflammatory (IL-1 b, IL-6, IL-17A, IL-17F, MCP-1, MCP-4). In some embodiments, the functional profile of each cell enables other metrics to be calculated, including the breakdown of each sample according to the cell's versatility (i.e., how many percentages of the cells are secreting multiple cytokines as compared to non-secreting or monofunctional cells), and the breakdown of the sample by functional groups (i.e., which mono-and multi-functional groups are being secreted by cells in the sample, and their frequency).
As used herein, "bone marrow cells" are a subset of leukocytes, including granulocytes, monocytes, macrophages and dendritic cells.
As used herein, the term "quartile" is a statistical term describing the division of observations into four defined intervals based on data values and how they are compared to the entire set of observations.
As used herein, the term "study day 0" is defined as the day on which the subject received the first CAR T cell infusion. The day prior to study day 0 will be study day-1. Any day after the entry into the group and prior to study day-1 will be consecutive and negative integer values.
As used herein, the term "sustained response" refers to a subject that is in a sustained response in a follow-up of at least one year after CAR T cell infusion. In one embodiment, the "duration of response" (DOR) is defined only for subjects experiencing an objective response and is the time from the first objective response to disease progression (Cheson et al, 2014) or disease-related death (based on prior occurrence).
As used herein, the term "relapse" refers to a subject who achieves a Complete Response (CR) or a Partial Response (PR) and subsequently experiences disease progression.
As used herein, the term "non-responsive" refers to a subject that has never experienced CR or PR following CAR T cell infusion.
As used herein, the term "objective response" refers to Complete Response (CR), partial Response (PR), or no response. Objective responses can be assessed according to revised IWG malignancy response criteria (Cheson et al, J Clin Oncol.2007;25 (5): 579-86).
As used herein, the term "complete response" refers to complete regression of a disease that becomes undetectable by radiological imaging and clinical laboratory assessment. There was no evidence of cancer at a given time.
As used herein, the term "partial response" refers to a tumor that is reduced by more than 30% but not completely regressed.
As used herein, an "objective response rate" (ORR) is determined according to International Working Group (IWG) 2007 standard (Cheson et al J Clin Oncol.2007;25 (5): 579-86).
As used herein, "Progression Free Survival (PFS)" may be defined as the time from the date of T cell infusion to the date of disease progression or death due to any cause. Progress was defined in terms of the evaluation of the response by researchers defined by the IWG standard (Cheson et al, J Clin Oncol.2007;25 (5): 579-86).
The term "Overall Survival (OS)" may be defined as the time from the date of T cell infusion to the date of death due to any cause.
As used herein, expansion and persistence of CAR T cells in peripheral blood can be monitored by qPCR analysis, for example using CAR-specific primers directed against the scFv portion of the CAR (e.g., the heavy chain of the CD19 binding domain) and its hinge/CD 28 transmembrane domain. Alternatively, it can be measured by calculating the number of CAR cells per unit blood volume.
As used herein, the predetermined blood draw of CAR T cells can be prior to CAR T cell infusion, day 7, week 2 (day 14), week 4 (day 28), month 3 (day 90), month 6 (day 180), month 12 (day 360), and month 24 (day 720).
As used herein, "peak of CAR T cells" is defined as the maximum absolute number of car+pbmcs/μl reached in serum after day 0.
As used herein, "time to peak of CAR T cells" is defined as the number of days from day 0 to the day of peak of CAR T cells.
As used herein, "area under the curve (AUC) of CAR T cell levels from day 0 to day 28" is defined as the area under the curve in the plot relative to CAR T cell levels from day 0 to day 28 for the planned visit. The AUC measures the total level of CAR T cells over time.
As used herein, the predetermined blood withdrawal of cytokines is prior to or on the day of conditioning chemotherapy (day-5), day 0, day 1, day 3, day 5, day 7, every other day to hospitalization (if any), week 2 (day 14), and week 4 (day 28).
As used herein, the "baseline" of a cytokine is defined as the last value measured prior to conditioning chemotherapy.
As used herein, fold change from baseline on day X is defined as
As used herein, the "peak post-baseline cytokine" is defined as the maximum level of cytokine in serum reached after baseline (day-5) until day 28.
As used herein, the "time to peak cytokine" following CAR T cell infusion is defined as the number of days from day 0 to the day of peak cytokine.
As used herein, "area under the curve (AUC) of cytokine levels from day-5 to day 28" is defined as the area under the curve in the plot of cytokine levels relative to planned visits from day-5 to day 28. The AUC measures the total level of cytokines over time. Whereas cytokines and car+ T cells are measured at certain discrete time points, the AUC can be estimated using trapezoidal rules.
As used herein, adverse events (TEAEs) occurring in treatment are defined as Adverse Events (AEs) occurring at or after the first dose of conditioning days. Adverse events may be coded with the regulatory active medical dictionary (MedDRA) version 22.0 and ranked using the National Cancer Institute (NCI) adverse event common terminology standard (CTCAE) version 4.03. Cytokine Release Syndrome (CRS) events can be ranked according to syndrome levels by Lee and colleagues (Lee et al, 2014blood.2014;124 (2): 188-95). Individual CRS symptoms may be graded according to CTCAE 4.03. Neurological events can be identified with search strategies based on known neurological toxicities associated with CAR T immunotherapy, such as, for example, topp, MS et al Lancet oncology.2015;16 57-66.
Various aspects of the disclosure are described in further detail in the following subsections.
Pre-treatment attributes
The pre-treatment properties of the apheresis and immune cells (also referred to herein as engineered cells, such as T cells) and patient immune factors measured from patient samples can be used to assess the probability of clinical outcome (including response and toxicity). Attributes associated with clinical outcome may be tumor-related parameters (e.g., tumor burden, serum LDH as a hypoxia/cell death marker, inflammatory markers associated with tumor burden and bone marrow cell activity), T cell attributes (e.g., T cell adaptability, functionality (especially T1-related ifnγ production), and total number of infused CD 8T cells), and CAR T cell engraftment as measured by peak CAR T cell levels in blood at early time points.
Information inferred from T cell properties and pre-patient treatment properties can be used to determine, refine, or prepare a therapeutically effective dose suitable for treating a malignancy (e.g., cancer). In addition, some T cell attributes and pre-patient treatment attributes can be used to determine whether a patient will develop adverse events (e.g., neurotoxicity (NT), cytokine Release Syndrome (CRS)) following treatment with engineered Chimeric Antigen Receptor (CAR) immunotherapy. Thus, an effective adverse event management strategy (e.g., based on the measured levels of one or more attributes, administration of tolizumab, corticosteroid therapy, or anti-epileptic drugs for toxicity prevention) may be determined.
In some embodiments, the pre-treatment property is a property of an engineered T cell comprising one or more chimeric antigen receptors. In some embodiments, the pre-treatment property is T cell transduction rate, primary T cell phenotype, number of CAR T cells and T cell subsets, CAR T cell fitness, T cell functionality, T cell versatility, number of differentiated car+cd8+ T cells, number of ccr7+cd45ra+ T cells, CD4/CD8 ratio, IFN- γ in the co-culture.
In some embodiments, the pre-treatment property is measured from a sample obtained from the patient (e.g., cerebrospinal fluid (CSF), blood, serum, or tissue biopsy). In some embodiments, the one or more pre-treatment attributes are tumor burden, IL-6 level, or LDH level.
T cell adaptation
In some embodiments, the innate cell adaptation is assessed based on the ability of CAR T cells to expand during in vitro non-specific stimulation (e.g., shorter doubling time), the differentiation status of CAR T cells (favorable juvenile phenotype), the level of specialized CAR T cell subpopulations in the CAR T cell population (e.g., the number of CD8 and naive-like CD8 cells (e.g., cd8+ccr7+cd45ra+ T cells) in infusion products), and the CAR T cell expansion rate in vivo.
In one embodiment, T cell adaptation is the ability of cells to rapidly expand. In the context of engineered T cells, in one embodiment, T cell adaptation is a measurement of how fast an engineered T cell population expands prior to treatment. T cell adaptation, as described herein, is a property of engineered T cells that is associated with clinical outcome. In some embodiments, T cell fitness is measured by doubling time or rate of expansion. An exemplary derivation of T cell "fitness" as measured by T cell population Doubling Time (DT) during the preparation process is shown below.
Duration can be defined as the total manufacturing time frame minus three days (essentially days of cultured product cells after transduction and prior to collection and cryopreservation). Recombinant IL-2 (after non-specific stimulation with, for example, an anti-CD 3 antibody) can be used to drive expansion of polyclonal T cells to achieve a target dose. The shorter the DT, the higher the engineered T cell adaptation. The in vitro amplification rate can be calculated using the following formula.
Amplification rate = ln (2)/doubling time
In the above case, the amplification rate is provided in units of "rate/day" or "/day".
In some embodiments, the in vivo rate of expansion is measured by enumerating CAR cells per unit blood volume. In some embodiments, the in vivo amplification rate is measured by CAR gene copy number/μg host DNA. In some embodiments, the in vivo rate of expansion is measured by enumerating CAR cells per unit blood volume.
As described herein, higher peak expansion of CAR T cells in peripheral blood (typically occurring within 2 weeks after CAR T cell infusion) may be associated with objective responses and persistent responses, which are defined as sustained responses with minimal follow-up for 1 year. The peak number of CAR T cells in the blood correlates with the response. Cumulative CAR T cell levels measured by area under the curve (AUC) in blood over the first 28 days may also be correlated with better objective and sustained response to therapy. In some embodiments, the CAR T cell level is calculated by enumerating the number of CAR T cells per unit blood volume. In one embodiment, higher peak expansion of CAR T cells in peripheral blood means that the peak expansion value falls within a higher quartile. In some embodiments, the in vivo rate of expansion is measured by enumerating CAR cells per unit blood volume. In some embodiments, the in vivo amplification rate is measured by CAR gene copy number/μg host DNA.
As described herein, the inherent ability of T cells to expand as measured prior to treatment as measured by product doubling time is a major attribute of product T cell adaptability. DT is most strongly correlated with the frequency of T cell differentiation subpopulations in the final infusion bag relative to other product characteristics. In particular, DT is positively correlated with the frequency of effector memory T (TEM) cells and negatively correlated with the frequency of naive T (TN) cells. In one embodiment (e.g., alemtuquor), T identified as CCR7+CD45RA+ cells N The cells are in fact stem-like memory cells rather than the typical naive T cells. As described herein, baseline tumor burden is positively correlated with differentiation phenotype in the final infusion product. As described herein, product composition and clinical manifestations are associated with a patient's pre-treatment immune status. Thus, in one embodiment, the present disclosure provides a method of reducing tumor burden following treatment with CAR T cells, comprising administering an infusion product comprising an increased frequency of naive T (TN) cells in the infusion product relative to a reference value. In another embodiment, the present disclosure provides methods of predicting or estimating the differentiation phenotype of a final infusion productA method comprising measuring a baseline tumor burden in a patient to obtain a value and estimating or predicting a differentiation phenotype based on the value. In one embodiment, measuring further comprises preparing an effective dose of CAR T cells in the final product based on the value.
T cell phenotype
As described herein, T cell phenotype in the manufacturing starting material (apheresis material) can be correlated with T cell fitness (DT). The total% of Tn-like cells and Tcm cells (ccr7+ cells) are inversely related to DT. The% of Tem (CCR 7-CD45 RA-) cells is directly related to DT. Thus, in some embodiments, the pre-treatment attribute is% of Tn-like cells and Tcm cells. In some embodiments, the% of Tn-like cells and Tcm cells is determined by the percentage of ccr7+ cells. In some embodiments, the percentage of ccr7+ cells is measured by flow cytometry.
In some embodiments, the pre-treatment attribute is% of Tem (CCR 7-CD45 RA-) cells. In some embodiments, the% of Tem cells is determined by the percentage of CCR7-CD45 RA-cells. In some embodiments, the percentage of CCR7-CD45 RA-cells is measured by flow cytometry.
As described herein, the greater the proportion of effector memory T cells in the singulated product within the total cd3+ T cells or CD4 and CD8 subsets, the longer the product doubling time. As described herein, the younger the T cell phenotype in the starting material, but the better the product T cell adaptability. As described herein, represents an immunocompetent T expressing a costimulatory molecule N CD27+CD28+T of cell subsets N Cells are positively correlated with product doubling time. As described herein, there is a direct link between all major phenotype groups, including the proportion of T cell subsets defined by differentiation markers in the CD3, CD4 and CD8 subgroups in the apheresis product relative to the final product phenotype. As described herein, with CD25 hi The proportion of T cells expressed by CD4 (which may represent regulatory T cells in the apheresis material) is inversely related to the CD 8T cell output in the product. As described herein, tumor burden following CAR T cell treatment is positively correlated with the differentiation phenotype of the final product.
T1 function
Engineered T cells may be characterized by their immune function. The methods of the present disclosure provide for measurement of levels of cytokine production ex vivo. In some embodiments, the cytokine is selected from the group consisting of IFN gamma, TNFa, IL-12, MIP1 beta, MIP1 alpha, IL-2, IL-4, IL-5 and IL-13. In some embodiments, T cell function is measured by the level of Th1 cytokines.
In some embodiments, the Th1 cytokine is selected from the group consisting of IFN gamma, TNFa and IL-12. In some embodiments, T cell function is measured by ifnγ production levels. In some embodiments, excess T cell ifnγ (pre-treatment attribute) and post-treatment T1 activity are attributes that can be used to determine whether a patient will experience an adverse event (e.g., neurotoxicity). In some embodiments, ifnγ levels produced by the engineered CAR T cells are measured by co-culturing prior to administration of the engineered CAR T cells.
Other immune cell product characteristics
In some embodiments, the immune cell product administered to the subject has several other product characteristics related to its efficacy. In some embodiments, the product features are selected from: total number of CAR T cells per μl; total number of T cells per μl; transduction rate,%; IFN-gamma levels, pg/mL; vitality,%; CD4/CD8 ratio; naive (ccr7+cd45ra+) T cells,%; central memory (ccr7+cd45ra-) T cells,%; (T) N +T CM )/(T EM +T EFF ) Ratio (T) CM A central memory T cell; t (T) EFF Effector T cells; t (T) EM Effector memory T cells; t (T) N Naive T cells).
Chimeric antigen receptor
Chimeric Antigen Receptors (CARs) are genetically engineered receptors. These engineered receptors can be inserted into and expressed by immune cells, including T cells and other lymphocytes, according to techniques known in the art. Using CARs, a single receptor can be programmed to recognize both a specific antigen and, upon binding to that antigen, activate immune cells to attack and destroy cells carrying that antigen. When these antigens are present on tumor cells, CAR-expressing immune cells can target and kill tumor cells. Chimeric antigen receptors can incorporate co-stimulatory (signaling) domains to increase their potency. See U.S. Pat. Nos. 7,741,465 and 6,319,494, and Krause et al and Finney et al (supra), song et al Blood 119:696-706 (2012); kalos et al, sci.Transl.Med.3:95 (2011); porter et al, N.Engl.J.Med.365:725-33 (2011), and Gross et al, annu.Rev.Pharmacol.Toxicol.56:59-83 (2016).
In some embodiments, the co-stimulatory domain comprising a truncated hinge domain ("THD") further comprises some or all of an immunoglobulin family member, such as IgG1, igG2, igG3, igG4, igA, igD, igE, igM, or a fragment thereof.
In some embodiments, THD is derived from human intact hinge domain ("CHD"). In other embodiments, the THD is derived from rodent, murine, or primate (e.g., non-human primate) CHD of the co-stimulatory protein. In some embodiments, the THD is derived from chimeric CHD of a costimulatory protein.
The co-stimulatory domain of a CAR of the present disclosure may also comprise a transmembrane domain and/or an intracellular signaling domain. The transmembrane domain can be fused to the extracellular domain of the CAR. The co-stimulatory domain may be similarly fused to the intracellular domain of the CAR. In some embodiments, a transmembrane domain that naturally associates with one of the domains in the CAR is used. In some cases, the transmembrane domains are selected or modified by amino acid substitutions to avoid binding of such domains to transmembrane domains of the same or different surface membrane proteins to minimize interactions with other members of the receptor complex. The transmembrane domain may be derived from natural or synthetic sources. Where the source is a natural source, the domain may be derived from any membrane-bound protein or transmembrane protein. The transmembrane region particularly useful in the present disclosure may be derived from (i.e., comprising) 4-1BB/CD137, activated NK cell receptor, immunoglobulin, B7-H3, BAFFR, BLAME (SLAMF 8), BTLA, CD100 (SEMA 4D), CD103, CD160 (BY 55), CD18, CD19 ase:Sub>A, CD2, CD247, CD27, CD276 (B7-H3), CD28, CD29, CD3 deltase:Sub>A, CD3 epsilon, CD3 gammase:Sub>A, CD3 zetase:Sub>A, CD30, CD4, CD40, CD49 ase:Sub>A, CD49D, CD f, CD69, CD7, CD84, CD8 alphase:Sub>A, CD8 betase:Sub>A, CD96 (Tactile), CD11 ase:Sub>A, CD11B, CD11c, CD11D, CDS, CEACAM1, CRT AM, cytokine receptor, DAP-10, DNAM1 (CD 226), fc gammase:Sub>A receptor, GADS, GITR, HVEM (LIGHTR), CD40 IA4, ICAM-1, igα (CD 79 ase:Sub>A), IL-2rβ, IL-2rγ, IL-7rα, inducible T cell costimulatory factor (ICOS), integrin, ITGA4, ITGA6, ITGAD, ITGAE, ITGAL, ITGAM, ITGAX, ITGB2, ITGB7, ITGBl, kirs 2, LAT, LFA-1, ligand that specifically binds CD83, LIGHT, LTBR, ly (CD 229), lymphocyte function-associated antigen 1 (LFA-1; CD11 ase:Sub>A/CD 18), MHC class 1 molecules, NKG2C, NKG2D, NKp30, NKp44, NKp46, NKp80 (KLRF 1), OX-40, PAG/Cbp, programmed death-1 (PD-1), PSGL1, SELPLG (CD 162), signaling lymphocyte activating molecules (SLAM proteins), SLAM (SLAMF 1); CD150, IPO-3), SLAMF4 (CD 244, 2B 4), SLAMF6 (NTB-A, lyl 08), SLAMF7, SLP-76, TNF receptor protein, TNFR2, TNFSF14, toll ligand receptor, TRANCE/RANKL, VLA1 or VLA-6, or fragments, truncated forms or combinations thereof.
Optionally, the short linker can form a link between any or some of the extracellular, transmembrane, and intracellular domains of the CAR. In some embodiments, the linker may be derived from a repeat of glycine-serine (SEQ ID NO: 2) (G4S) n or GSTSGSGKPGSGEGSTKG (SEQ ID NO: 1). In some embodiments, the linker comprises 3-20 amino acids and an amino acid sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to GSTSGSGKPGSGEGSTKG (SEQ ID NO: 1).
The linkers described herein may also be used as peptide tags. The linker peptide sequence may be of any suitable length to link one or more proteins of interest, and is preferably designed to be flexible enough to allow for proper folding and/or function and/or activity of the one or both peptides to which it is linked. Thus, the linker peptide may be no more than 10, no more than 11, no more than 12, no more than 13, no more than 14, no more than 15, no more than 16, no more than 17, no more than 18, no more than 19, or no more than 20 amino acids in length. In some embodiments, the linker peptide comprises a length of at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, or at least 20 amino acids. In some embodiments, the linker comprises at least 7 and no more than 20 amino acids, at least 7 and no more than 19 amino acids, at least 7 and no more than 18 amino acids, at least 7 and no more than 17 amino acids, at least 7 and no more than 16 amino acids, at least 7 and no more than 15 amino acids, at least 7 and no more than 14 amino acids, at least 7 and no more than 13 amino acids, at least 7 and no more than 12 amino acids, or at least 7 and no more than 11 amino acids. In certain embodiments, the linker comprises 15-17 amino acids, and in particular embodiments, 16 amino acids. In some embodiments, the linker comprises 10-20 amino acids. In some embodiments, the linker comprises 14-19 amino acids. In some embodiments, the linker comprises 15-17 amino acids. In some embodiments, the linker comprises 15-16 amino acids. In some embodiments, the linker comprises 16 amino acids. In some embodiments, the linker comprises 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 amino acids.
In some embodiments, spacer domains are used. In some embodiments, the spacer domain is derived from CD4, CD8a, CD8b, CD28T, 4-1BB or other molecules described herein. In some embodiments, the spacer domain may include a chemically induced dimer to control expression upon addition of a small molecule. In some embodiments, no spacer is used.
The intracellular (signaling) domain of the engineered T cells of the present disclosure can provide signaling to an activation domain that then activates at least one normal effector function of the immune cell. Effector functions of T cells may be, for example, cytolytic activity or helper activity, including secretion of cytokines.
In certain embodiments, suitable intracellular signaling domains include (i.e., including) but not limited to 4-1BB/CD137, activated NK cell receptor, immunoglobulin, B7-H3, BAFFR, BLAME (SLAMF 8), BTLA, CD100 (SEMA 4D), CD103, CD160 (BY 55), CD18, CD19 ase:Sub>A, CD2, CD247, CD27, CD276 (B7-H3), CD28, CD29, CD3 deltase:Sub>A, CD3 epsilon, CD3 gammase:Sub>A, CD30, CD4, CD40, CD49 ase:Sub>A, CD49D, CD f, CD69, CD7, CD84, CD8 alphase:Sub>A, CD8 betase:Sub>A, CD96 (Tactile), CD11 ase:Sub>A, CD11B, CD11c, CD11D, CDS, CEACAM1, CRT AM, cytokine receptor, DAP-10, DNAM1 (CD 226), fc gammase:Sub>A receptor, GADS, GITR, HVEM (LIGHTR), CD40 IA4, ICAM-1, igα (CD 79 ase:Sub>A), IL-2rβ, IL-2rγ, IL-7rα, inducible T cell costimulatory factor (ICOS), integrin, ITGA4, ITGA6, ITGAD, ITGAE, ITGAL, ITGAM, ITGAX, ITGB2, ITGB7, ITGBl, kirs 2, LAT, ligand that specifically binds CD83, LIGHT, LTBR, ly (CD 229), lyl08, lymphocyte function-associated antigen 1 (LFA-1; CD11 ase:Sub>A/CD 18), MHC class 1 molecules, NKG2C, NKG2D, NKp30, NKp44, NKp46, NKp80 (KLRF 1), OX-40, PAG/Cbp, programmed death-1 (PD-1), PSGL1, SELPLG (CD 162), signaling lymphocyte activating molecules (SLAM proteins), SLAM (SLAMF 1); CD150; IPO-3), SLAMF4 (CD 244;2B 4), SLAMF6 (NTB-A), SLAMF7, SLP-76, TNF receptor protein, TNFR2, TNFSF14, toll ligand receptor, TRANCE/RANKL, VLA1 or VLA-6, or fragments, truncated forms thereof, or combinations thereof.
Antigen binding molecules
A suitable CAR can bind to an antigen (such as a cell surface antigen) by incorporating an antigen binding molecule that interacts with the targeted antigen. In some embodiments, the antigen binding molecule is an antibody fragment thereof, e.g., one or more single chain antibody fragments ("scFv"). scFv is a single chain antibody fragment having antibody heavy and light chain variable regions linked together. See U.S. Pat. Nos. 7,741,465 and 6,319,494, eshhar et al, cancer Immunol Immunotherapy (1997) 45:131-136.scFv retain the ability of the parent antibody to specifically interact with the target antigen. scFv are useful in chimeric antigen receptors because they can be engineered to be expressed as part of a single chain with other CAR components. As above. See also Krause et al, J.Exp.Med., volume 188, stage 4, 1998 (619-626); finney et al Journal of Immunology,1998,161:2791-2797. It will be appreciated that the antigen binding molecule is typically contained within the extracellular portion of the CAR such that it is capable of recognizing and binding to the antigen of interest. Bispecific and multispecific CARs are contemplated within the scope of the present disclosure to have specificity for more than one target of interest.
In some embodiments, the polynucleotide encodes a CAR comprising a (truncated) hinge domain and an antigen binding molecule that specifically binds to a target antigen. In some embodiments, the target antigen is a tumor antigen. In some embodiments, the antigen is selected from the group consisting of tumor-associated surface antigens, such as 5T4, alpha Fetoprotein (AFP), B7-1 (CD 80), B7-2 (CD 86), BCMA, B-human chorionic gonadotrophin, CA-125, carcinoembryonic antigen (CEA), CD123, CD133, CD138, CD19, CD20, CD22, CD23, CD24, CD25, CD30, CD34, CD4, CD40, CD44, CD56, CD8, CLL-1, c-Met, CMV-specific antigen, CS-1, CSPG4, CTLA-4, DLL3, disialoganglioside GD2, ductal-epithelial mucin, EBV-specific antigen, EGFR variant III (ELF 2M), endoglin, hepatins B2, epidermal Growth Factor Receptor (EGFR) epithelial cell adhesion molecule (EpCAM), epithelial tumor antigen, erbB2 (HER 2/neu), fibroblast-related protein (fap), FLT3, folate binding protein, GD2, GD3, glioma-related antigen, glycosphingolipids, gp36, HBV-specific antigen, HCV-specific antigen, HER1-HER2, HER2-HER3 combination, HERV-K, high molecular weight melanoma-related antigen (HMW-MAA), HIV-1 envelope glycoprotein gp41, HPV-specific antigen, human telomerase reverse transcriptase, IGFI receptor, IGF-II, IL-11 ra, IL-13R-a2, influenza virus-specific antigen; CD38, insulin growth factor (IGFl) -l, enterocarboxylesterase, kappa chain, LAGA-la, lambda chain, lasa-specific antigen, lectin-reactive AFP, lineage-specific or tissue-specific antigen such AS CD3, MAGE-A1, major Histocompatibility Complex (MHC) molecule, major Histocompatibility Complex (MHC) molecule presenting tumor-specific peptide epitopes, M-CSF, melanoma-associated antigen, mesothelin, MN-CA IX, MUC-1, mut hsp70-2, mutant p53, mutant ras, neutrophil elastase, NKG2D, nkp, NY-ESO-1, p53, PAP prostases, prostate Specific Antigen (PSA), prostate cancer tumor antigen-1 (PCTA-1), prostate specific antigen proteins, STEAP1, STEAP2, PSMA, RAGE-1, ROR1, RU2 (AS), surface adhesion molecules, survivin and telomerase, TAG-72, the Extra Domain A (EDA) and Extra Domain B (EDB) of fibronectin and the Al domain of tenascin-C (TnC Al), thyroglobulin, tumor matrix antigen, vascular endothelial growth factor receptor-2 (VEGFR 2), virus-specific surface antigens such AS HIV-specific antigens (such AS HIV gpl 20), and any derivatives or variants of these surface antigens.
The present disclosure is further illustrated by the following examples, which should not be construed as further limiting. The disclosure provided herein may be used in a variety of ways other than or as a combination of the above. The following is a compilation of exemplary methods that may be derived from the disclosure provided in this application.
Engineered immune cells and uses
In one embodiment, the cells of the present disclosure may be obtained by T cells obtained from a subject. T cells may be obtained, for example, from peripheral blood mononuclear cells, bone marrow, lymph node tissue, umbilical cord blood, thymus tissue, tissue from an infected site, ascites, pleural effusion, spleen tissue, tumors, or differentiated in vitro. In addition, T cells may be derived from one or more T cell lines available in the art. Various techniques known to the skilled artisan (such as FICOLL TM Isolation and/or apheresis) to obtain T cells from a blood unit collected from a subject. In some embodiments, cells collected by apheresis are washed to remove plasma fractions and placed in an appropriate buffer or medium for subsequent processing. In some embodiments, the cells are washed with PBS. It should be understood that washing steps, such as Such as by using a semi-automatic flow-through centrifuge, e.g. Cobetm 2991 cell processor, baxter CytoMate TM Etc. In some embodiments, the washed cells are resuspended in one or more biocompatible buffers or other saline solution with or without a buffer. In some embodiments, unwanted components of the blood apheresis sample are removed. Additional methods for isolating T cells for T cell therapy are disclosed in U.S. patent publication 2013/0287748, which is incorporated herein by reference in its entirety.
In some embodiments, the method comprises the steps of lysing the red blood cells and depleting the monocytes (e.g., by using a dye of the formula PERCOL TM Gradient centrifugation) to separate T cells from PBMCs. In some embodiments, specific subsets 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 a T cell population by negative selection can be accomplished using a combination of antibodies directed against a surface marker specific for the cells of the negative selection. In some embodiments, cell sorting and/or selection via negative magnetic immunoadhesion or flow cytometry may be used, using a mixture of monoclonal antibodies directed against cell surface markers present on negatively selected cells. For example, to enrich for cd4+ cells by negative selection, monoclonal antibody mixtures typically comprise antibodies directed against CD8, CD11b, CD14, CD16, CD20 and HLA-DR. In some embodiments, flow cytometry and cell sorting are used to isolate a population of cells of interest for use in the present disclosure.
In some embodiments, PBMCs are directly used for genetic modification of immune cells (such as CARs) using methods as described herein. In some embodiments, after isolating PBMCs, T lymphocytes are further isolated and cytotoxic and helper T lymphocytes are sorted into naive, memory and effector T cell subsets either before or after genetic modification and/or expansion.
In some embodiments, the 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 a phenotypic marker of central memory T cells includes expression of CCR7, CD3, CD28, CD45RO, CD62L, and CD127 and is negative for granzyme B. In some embodiments, the 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, the cd4+ T cells are further sorted into subpopulations. For example, cd4+ T helper cells can be sorted into naive, central memory, and effector cells by identifying a population of cells with cell surface antigens.
In some embodiments, immune cells (e.g., T cells) are genetically modified after isolation using known methods, or activated and expanded in vitro (or differentiated in the case of progenitor cells) prior to genetic modification of immune cells. In another embodiment, immune cells (e.g., T cells) are genetically modified (e.g., transduced with a viral vector comprising one or more nucleotide sequences encoding a CAR) with the chimeric antigen receptor described herein, and then activated and/or expanded in vitro. Methods for activating and expanding T cells are known in the art and are described, for example, in U.S. patent nos. 6,905,874, 6,867,041 and 6,797,514 and PCT publication No. WO 2012/079000, the contents of which are hereby incorporated by reference in their entirety. Generally, such methods involve contacting PBMCs or isolated T cells with stimulators and co-stimulators (such as anti-CD 3 and anti-CD 28 antibodies) that are typically attached to beads or other surfaces in a medium with an appropriate cytokine (such as IL-2). The anti-CD 3 antibody and the anti-CD 28 antibody attached to the same bead act as "surrogate" Antigen Presenting Cells (APCs). One example isThe system, a CD3/CD28 activator/stimulator system for physiological activation of human T cells. In other embodiments, use is made of materials such as those described in U.S. Pat. nos. 6,040,177 and 5,827,642 and PCT publication WO 2012/129514 (the contents of these patents are hereby incorporated by reference in their entirety) In this method, T cells are activated and stimulated to proliferate with feeder cells and appropriate antibodies and cytokines. In some embodiments, the T cells are obtained from a donor subject. In some embodiments, the donor subject is a human patient suffering from a cancer or tumor. In some embodiments, the donor subject is a human patient not suffering from cancer or tumor.
In one embodiment, the present disclosure provides a method of manufacturing an immunotherapeutic product with improved clinical efficacy and/or reduced toxicity. In some embodiments, the immunotherapeutic product comprises blood cells. In some embodiments, blood cells collected from the subject are washed, e.g., to remove plasma fractions and the cells are placed in an appropriate buffer or medium for subsequent processing steps. In some embodiments, the cells are washed with Phosphate Buffered Saline (PBS). In some embodiments, the wash solution is free of calcium and/or magnesium and/or many or all divalent cations. In some embodiments, the washing step is accomplished by a semi-automated "flow-through" centrifuge (e.g., cobe 2991 cell processor, baxter) according to manufacturer's instructions. In some embodiments, the washing step is accomplished by Tangential Flow Filtration (TFF) according to manufacturer's instructions. In some embodiments, cells are resuspended in various biocompatible buffers after washing, such as PBS without ca++ mg++. In certain embodiments, components of the blood cell sample are removed and the cells are resuspended directly in culture medium.
In some embodiments, these methods include density-based cell separation methods, such as the preparation of leukocytes from peripheral blood by lysing erythrocytes and by Percoll or Ficoll gradient centrifugation. In some embodiments, the methods comprise leukocyte isolation.
In some embodiments, at least a portion of the selecting step comprises incubating the cells with a selection reagent. Incubation with one or more selection reagents, for example, as part of a selection method, may be performed using one or more selection reagents for selecting one or more different cell types based on the expression or presence of one or more specific molecules (such as surface markers, e.g., surface proteins, intracellular markers, or nucleic acids) in or on the cell. In some embodiments, any known method employing one or more selection reagents for isolation based on such markers may be used. In some embodiments, the one or more selection reagents result in an isolation based on affinity or immunoaffinity separation. For example, selection in some embodiments includes incubation with one or more reagents for isolating cells and cell populations based on the expression or expression levels of cells of one or more markers (typically cell surface markers), e.g., by incubation with antibodies or binding partners that specifically bind these markers, followed by a washing step and separating cells that have bound to the antibodies or binding partners from cells that have not bound to the antibodies or binding partners.
In some embodiments of such methods, a volume of cells is mixed with a quantity of a desired affinity-based selection reagent. Immunoaffinity-based selection can be performed using any system or method that results in favorable energy interactions between the isolated cells and molecules that specifically bind to markers on the cells (e.g., antibodies or other binding partners on a solid surface, e.g., particles). In some embodiments, the method is performed using particles such as beads (e.g., magnetic beads) coated with a selective agent (e.g., an antibody) specific for a cellular marker. The particles (e.g., beads) can be incubated or mixed with the cells in a container (such as a tube or bag) while shaking or mixing, with a constant ratio of cell density to particles (e.g., beads) to help promote energetically favorable interactions. In other cases, these methods include selecting cells, wherein all or part of the selection is performed in the lumen of the chamber, e.g., under centrifugal rotation. In some embodiments, the incubation of the cells with a selection reagent (such as a selection reagent based on immunoaffinity) is performed in a chamber.
In some embodiments, by performing such a selection step or portion thereof (e.g., incubation with antibody-coated particles such as magnetic beads) in the cavity of a chamber, a user is able to control certain parameters, such as the volume of the various solutions, the addition of solutions during processing, and timing thereof, which may provide advantages over other available methods. For example, the ability to reduce the volume of liquid in the chamber during incubation can increase the concentration of particles (e.g., bead reagents) used in the selection and thus increase the chemical potential of the solution without affecting the total number of cells in the chamber. This in turn may enhance the pairwise interactions between the treated cells and the particles for selection. In some embodiments, performing the incubation step in the chamber allows the user to achieve agitation of the solution at a desired time during the incubation, such as when associated with systems, circuits, and controls as described herein, which may also improve interaction.
In some embodiments, at least a portion of the selecting step is performed in a chamber, which includes incubating the cells with a selection reagent. In some embodiments of such methods, a volume of cells is mixed with a quantity of the desired affinity-based selection reagent that is much less than the quantity typically used when similar selections are made in a tube or vessel for selecting the same number of cells and/or the same volume of cells according to manufacturer's instructions. In some embodiments, the amount of one or more selection reagents used does not exceed 5%, 10%, 15%, 20%, 25%, 50%, 60%, 70% or 80% of the amount of the same selection reagent used to select cells in a tube or vessel based incubation for the same number of cells and/or the same volume of cells according to manufacturer's instructions.
In some embodiments, for selection, e.g., cell selection based on immunoaffinity, the cells are incubated in a composition in a chamber that also contains a selection buffer and a selection reagent, such as a molecule that specifically binds to a surface marker on the cells that need to be enriched and/or depleted but not on other cells in the composition, such as an antibody, which is optionally coupled to a scaffold such as a polymer or a surface, e.g., a bead, e.g., a magnetic bead, such as a magnetic bead coupled to a monoclonal antibody specific for CD4 and CD 8. In some embodiments, as described, when selecting in a tube that is shaken or rotated, the selection reagent is added to the cells in the chamber in an amount that is substantially less (e.g., no more than 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, or 80%) than that necessary to achieve about the same or similar selection efficiency for the same number of cells or the same volume of cells that are typically used or are used. In some embodiments, the incubation is performed with the addition of a selection buffer to the cells and selection reagent to achieve a target volume for the reagent incubation, e.g., 10mL to 200mL, such as at least or about at least 10mL, 20mL, 30mL, 40mL, 50mL, 60mL, 70mL, 80mL, 90mL, 100mL, 150mL, or 200mL. In some embodiments, the selection buffer and the selection reagent are pre-mixed prior to addition to the cells. In some embodiments, the selection buffer and the selection reagent are added separately to the cells. In some embodiments, the selection incubation is performed under periodic gentle mixing conditions, which may help promote energetically favorable interactions, allowing for the use of less total selection reagent while achieving high selection efficiency. In some embodiments, the total duration of incubation with the selection reagent is about 5 minutes to 6 hours, such as 30 minutes to 3 hours, for example at least or about at least 30 minutes, 60 minutes, 120 minutes, or 180 minutes.
In some embodiments, the incubation is typically performed under mixing conditions, such as in the presence of rotation, typically at a relatively low force or speed, such as below the speed used to precipitate the cells, such as at or about 600rpm to 1700rpm (e.g., at or about or at least 600rpm, 1000rpm, or 1500rpm or 1700 rpm), such as at or about 80g to l00g (e.g., at or about or at least 80g, 85g, 90g, 95g, or 100 g) of RCF at the sample or wall of the chamber or other container. In some embodiments, rotation is used at repeated intervals of such low speed rotation followed by rest periods, such as rotation and/or rest for 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 seconds, such as rotation at about 1 or 2 seconds followed by rest for about 5, 6, 7, or 8 seconds.
In some embodiments, such a process is performed within a completely closed system integral to the chamber. In some embodiments, the process (and in some embodiments also one or more additional steps, such as a previous washing step to wash a cell-containing sample (such as a single sample)) is performed in an automated fashion such that cells, reagents, and other components are drawn into and out of the chamber at the appropriate time and centrifuged to complete the washing and binding steps in a single closed system using an automated procedure.
In some embodiments, after incubating and/or mixing the cells with one or more selection reagents, the incubated cells are isolated to select the cells based on the presence or absence of the one or more specific reagents. In some embodiments, the isolation is performed in the same closed system in which the incubation of the cells with the selection agent is performed. In some embodiments, after incubation with the selection agent, the incubated cells (including cells in which the selection agent has been bound) are transferred to a system for immunoaffinity-based cell separation. In some embodiments, the system for immunoaffinity-based separation is or comprises a magnetic separation column.
In some embodiments, the isolation method comprises isolating the different cell types based on the expression or presence in the cell of one or more specific molecules, such as a surface marker, e.g., a surface protein, an intracellular marker, or a nucleic acid. In some embodiments, any known isolation method based on such markers may be used. In some embodiments, the separation is an affinity or immunoaffinity based separation. For example, isolation in some embodiments includes isolating cells and cell populations based on cell expression or expression levels of one or more markers (typically cell surface markers), e.g., by incubation with antibodies or binding partners that specifically bind such markers, followed by a washing step typically and separating cells that have bound to the antibodies or binding partners from cells that have not bound to the antibodies or binding partners. Such isolation steps may be based on positive selection in which cells that have bound the reagent are retained for further use and/or negative selection in which cells that have not bound the antibody or binding partner are retained. In some examples, both portions are reserved for further use.
In some embodiments, negative selection may be particularly useful when no antibodies are available that specifically identify cell types in a heterogeneous population, such that the isolation is preferably based on markers expressed by cells other than the desired population.
Isolation need not result in 100% enrichment or removal of a particular cell population or cells expressing a particular marker. For example, positive selection or enrichment of a particular type of cell (such as a cell that expresses a marker) refers to increasing the number or percentage of such cells, but does not necessarily result in the complete absence of cells that do not express the marker. Likewise, positive selection, removal, or depletion of a particular type of cell (such as a cell expressing a marker) refers to a reduction in the number or percentage of such cells, but does not necessarily result in complete removal of all such cells.
In some examples, multiple rounds of separation steps are performed, wherein portions of positive or negative selections from one step are subjected to another separation step, such as subsequent positive or negative selections. In some examples, a single isolation step may consume cells that simultaneously express multiple markers, such as by incubating the cells with multiple antibodies or binding partners, each antibody or binding partner being specific for the marker targeted by negative selection. Likewise, multiple cell types can be positively selected simultaneously by incubating the cells with multiple antibodies or binding partners expressed on the multiple cell types.
For example, in some embodiments, a particular subpopulation of T cells, such as cells that are positive or express high levels of one or more surface markers, e.g., cd28+, cd62l+, ccr7+, cd27+, cd127+, cd4+, cd8+, cd45ra+ and/or cd45ro+ T cells, are isolated by positive or negative selection techniques. For example, anti-CD 3/anti-CD 28 conjugated magnetic beads may be used (e.g.,m-450CD3/CD28T cell expander) to positively select CD3+, CD28+ T cells. In some embodiments, the cell population is enriched for T cells having a naive phenotype (cd45ra+ccr7+).
In some embodiments, the isolation is performed by enriching a specific cell population by positive selection or depleting a specific cell population by negative selection. In some embodiments, positive or negative selection is accomplished by incubating the cells with one or more antibodies or other binding agents that specifically bind to one or more surface markers expressed on the positive or negative selected cells (labeled +) or at a relatively high level (labeled high), respectively.
In particular embodiments, a biological sample (e.g., a sample of PBMCs or other leukocytes) is subjected to selection of cd4+ T cells, wherein both negative and positive portions are retained. In certain embodiments, the cd8+ T cells are selected from the negative portion. In some embodiments, the biological sample is subjected to cd8+ T cell selection, wherein both the negative and positive portions are retained. In certain embodiments, the cd4+ T cells are selected from the negative part.
In some embodiments, T cells are isolated from a PBMC sample by negative selection for a marker expressed on non-T cells such as B cells, monocytes or other leukocytes such as CD 14. In some embodiments, cd4+ or cd8+ selection steps are used to isolate cd4+ helper T cells and cd8+ cytotoxic T cells. Such cd4+ and cd8+ populations may be further sorted into subpopulations by positive or negative selection of markers expressed on one or more naive, memory and/or effector T cell subpopulations or expressed to a relatively high degree.
In some embodiments, the cd8+ cells are further enriched or naive, central memory, effector memory and/or central memory stem cells are depleted, such as by positive or negative selection based on surface antigens associated with the respective subpopulations. In some embodiments, enrichment of central memory T (TCM) cells is performed to increase efficacy, such as improving long-term survival, expansion, and/or engraftment after administration, which in some embodiments is particularly robust in such subpopulations. In some embodiments, combining cd8+ T cells enriched for TcM with cd4+ T cells further enhances efficacy. In some embodiments, T cells with a naive phenotype (cd45ra+ccr7+) are enriched for enhanced efficacy. In embodiments, memory T cells are present in the cd62l+ and CD62L subsets of cd8+ peripheral blood lymphocytes. PBMCs may enrich or deplete cd62lcd8+ and/or cd62l+cd8+ fractions, such as with anti-CD 8 and anti-CD 62L antibodies.
In some embodiments, enrichment of central memory T (TCM) cells is based on positive or high surface expression of CD45RO, CD62L, CCR7, CD28, CD3 and/or CD 127; in some embodiments, it is based on negative selection of cells expressing or highly expressing CD45RA and/or granzyme B. In some embodiments, isolation of the cd8+ population enriched for TCM cells is performed by depleting cells expressing CD4, CD14, CD45RA and positively selecting or enriching for cells expressing CD 62L. In one embodiment, enrichment of central memory T (TCM) cells is performed starting from the negative portion of cells selected based on CD4 expression, which undergoes negative selection based on CD14 and CD45RA expression and positive selection based on CD 62L. Such selections are made simultaneously in some embodiments, and sequentially in either order in other embodiments. In some embodiments, the same CD4 expression-based selection step used to prepare the cd8+ cell population or subpopulation is also used to generate the cd4+ cell population or subpopulation such that both positive and negative portions from the CD 4-based isolation are retained and used in subsequent steps of these methods, optionally after one or more additional positive or negative selection steps. In a specific example, a PBMC sample or other leukocyte sample is subjected to cd4+ cell selection, wherein both negative and positive portions are retained. The negative portion is then negative selected based on the expression of CD14 and CD45RA or CD19, and positive selection is performed based on a marker characteristic of central memory T cells (such as CD62L or CCR 7), wherein the positive and negative selections are performed in either order.
Cd4+ T helper cells are sorted into naive, central memory and effector cells by identifying a population of cells with cell surface antigens. Cd4+ lymphocytes can be obtained by standard methods. In some embodiments, the naive cd4+ T lymphocytes are cd45ro, cd45ra+, cd62l+, cd4+ T cells. In some embodiments, the central memory cd4+ cells are cd62l+ and cd45ro+. In some embodiments, effector cd4+ cells are CD62L and CD45RO. In some embodiments, the T cells with a naive phenotype are cd45ra+ccr7+.
In one example, to enrich for cd4+ cells by negative selection, a monoclonal antibody mixture typically includes antibodies to CD14, CD20, CD11b, CD16, HLA-DR, and CD 8. In some embodiments, the antibody or binding partner is bound to a solid support or matrix, such as magnetic or paramagnetic beads, to allow separation of cells for positive and/or negative selection. For example, in some embodiments, immunomagnetic (or affinity magnetic) separation techniques are used to separate or segregate cells and cell populations. In some embodiments, a sample or composition of cells to be isolated is incubated with a small, magnetizable or magnetically responsive material, such as magnetically responsive particles or microparticles, such as paramagnetic beads (e.g., dynalbeads or MACS beads). The magnetically responsive material (e.g., particle) is typically directly or indirectly attached to a binding partner (e.g., antibody) that specifically binds to a molecule (e.g., a surface marker) present on one or more cells or cell populations that are desired to be isolated (e.g., negative or positive selection is desired).
In some embodiments, the magnetic particles or beads comprise magnetically responsive material bound to a specific binding member (such as an antibody or other binding partner). Many well known magnetically responsive materials are used in the magnetic separation process. Incubation is typically performed under conditions in which the antibody or binding partner or molecules (such as secondary antibodies or other reagents) that specifically bind to such antibodies or binding partners attached to the magnetic particles or beads specifically bind to cell surface molecules (if present on cells within the sample). In some embodiments, the sample is placed in a magnetic field and those cells with magnetically responsive or magnetizable particles attached will be attracted to the magnet and separated from unlabeled cells. For positive selection, cells attracted to the magnet are retained; for negative selection, cells that were not attracted (unlabeled cells) were retained. In some embodiments, a combination of positive and negative selections is performed during the same selection step, wherein the positive and negative fractions are retained and further processed or subjected to further separation steps. In some embodiments, the magnetically responsive particles are coated in a primary or other binding partner, secondary antibody, lectin, enzyme or streptavidin. In certain embodiments, the magnetic particles are attached to the cells via a coating of a primary antibody specific for one or more markers. In certain embodiments, cells (rather than beads) are labeled with a primary antibody or binding partner, and then magnetic particles coated with a cell type-specific secondary antibody or other binding partner (e.g., streptavidin) are added. In certain embodiments, streptavidin-coated magnetic particles are used in combination with a biotinylated primary or secondary antibody. In some embodiments, the magnetically responsive particles are attached to cells that are subsequently incubated, cultured, and/or engineered; in some embodiments, the particles remain attached to the cells for administration to a patient. In some embodiments, the magnetizable or magnetically responsive particles are removed from the cells. Methods for removing magnetizable particles from cells are known and include, for example, the use of competitive non-labeled antibodies, as well as magnetizable particles or antibodies conjugated with cleavable linkers. In some embodiments, the magnetizable particles are biodegradable.
In some embodiments, affinity-based selection is via Magnetically Activated Cell Sorting (MACS) (Miltenyi Biotec, auburn, CA). Magnetically Activated Cell Sorting (MACS) systems are capable of selecting cells with magnetized particles attached thereto in high purity. In certain embodiments, MACS operates in a mode in which non-target and target substances elute sequentially after application of an external magnetic field. That is, cells attached to the magnetized particles are held in place, while unattached material is eluted. Then, after this first elution step is completed, the substances that are trapped in the magnetic field and prevented from eluting are released in a way that they can be eluted and recovered. In certain embodiments, non-target cells are labeled and removed from the heterogeneous cell population.
In some embodiments, the separation or isolation is performed using a system, apparatus, or device that performs one or more of the separation, cell preparation, isolation, processing, incubation, culturing, and/or formulating steps of the methods. In some embodiments, the system is used to perform each of these steps in a closed or sterile environment, for example, to minimize errors, user handling, and/or contamination. In one example, the system is a system as described in International patent application publication No. W02009/072003 or US 20110003380 Al. In some embodiments, the system or apparatus performs one or more (e.g., all) of the separation, processing, engineering, and formulation steps in an integrated or stand-alone system and/or in an automated or programmable manner. In some embodiments, the system or apparatus includes a computer and/or computer program in communication with the system or apparatus that allows a user to program, control, evaluate, and/or adjust various embodiments of the processing, separation, engineering, and formulation steps. In some embodiments, the isolation and/or other steps are performed using a clinic macs system (Miltenyi Biotec), for example for automatically isolating cells at the clinical scale level in a closed and sterile system. The components may include an integrated microcomputer, a magnetic separation unit, a peristaltic pump, and various pinch valves. In some embodiments, the computer controls the components of the instrument and directs the system to perform the repeated procedure in a standardized order. In some embodiments, the magnetic separation unit comprises a movable permanent magnet and a holder for the selection column. Peristaltic pumps control the flow rate throughout the tubing set and, together with pinch valves, ensure controlled flow of buffer through the system and continuous suspension of cells.
In some embodiments, the CliniMACS system uses antibody-conjugated magnetizable particles provided in a sterile, pyrogen-free solution. In some embodiments, after labeling the cells with magnetic particles, the cells are washed to remove excess particles. The cell preparation bag is then connected to a tubing set which in turn is connected to a bag containing buffer and a cell collection bag. The tubing set consists of pre-assembled sterile tubing, including pre-columns and separation columns, and is provided for single use only. After the separation procedure is initiated, the system automatically applies the cell sample to the separation column. The labeled cells remain in the column, while unlabeled cells are removed by a series of washing steps. In some embodiments, the population of cells used in the methods described herein is not labeled and does not remain in the column. In some embodiments, the population of cells used in the methods described herein is labeled and retained in the column. In some embodiments, the population of cells used in the methods described herein elutes from the column after removal of the magnetic field and is collected in a cell collection bag.
In certain embodiments, the isolation and/or other steps are performed using a CliniMACS Prodigy system (Miltenyi Biotec). In some embodiments, the CliniMACS Prodigy system is equipped with a cell handling unit that allows for automatic washing and fractionation of cells by centrifugation. CliniMACS Prodigy system may also include an on-board camera and image recognition software to determine the optimal cell classification endpoint by discriminating the macroscopic layers of the source cell product. For example, peripheral blood is automatically separated into red blood cells, white blood cells, and plasma layers. CliniMACS Prodigy systems may also include integrated cell culture chambers that accomplish cell culture protocols such as cell differentiation and expansion, antigen loading, and long term cell culture. The input port may allow for sterile removal and replenishment of the culture medium, and an integrated microscope may be used to monitor the cells.
In some embodiments, the cell populations described herein are collected and enriched (or depleted) by flow cytometry, wherein cells stained for a plurality of cell surface markers are carried in a fluid stream. In some embodiments, the cell populations described herein are collected and enriched (or depleted) by preparative scale (FACS) sorting. In certain embodiments, the cell populations described herein are collected and enriched (or depleted) by using a combination of microelectromechanical systems (MEMS) chips and FACS-based detection systems (see, e.g., WO 2010/033140, cho et al (2010) Lab Chip 10,1567-1573; and Godin et al (2008) J Biophoton.l (5): 355-376). In both cases, the cells can be labeled with a variety of markers, allowing the isolation of well-defined T cell subsets in high purity.
In some embodiments, the antibody or binding partner is labeled with one or more detectable markers to facilitate isolation of positive and/or negative selections. For example, the separation may be based on binding to a fluorescently labeled antibody. In some examples, cell separation based on binding of antibodies or other binding partners specific for one or more cell surface markers is performed in a fluid stream, such as by Fluorescence Activated Cell Sorting (FACS), including preparation-scale (FACS) and/or microelectromechanical systems (MEMS) chips, e.g., in combination with a flow cytometry detection system. Such methods allow for positive and negative selection based on multiple markers simultaneously.
In some embodiments, the method of preparation includes the step of freezing (e.g., cryopreserving) the cells before or after isolation, incubation, and/or engineering. In some embodiments, the freezing and subsequent thawing steps remove granulocytes and, to some extent, monocytes from the cell population. In some embodiments, the cells are suspended in a frozen solution, for example after a washing step to remove plasma and platelets. In some embodiments, any of a variety of known freezing solutions and parameters may be used. One example involves the use of PBS or other suitable cell freezing medium containing 20% DMSO and 8% Human Serum Albumin (HSA). Then diluted 1:1 with medium so that the final concentrations of DMSO and HSA were 10% and 4%, respectively. The cells are then typically frozen to-80 ℃ at a rate of 1 ℃/min and stored in the gas phase of a liquid nitrogen storage tank.
In some embodiments, one or more import compositions that produce enriched T cells, e.g., cd3+ T cells, cd4+ T cells, and/or cd8+ T cells, are isolated and/or selected. In some embodiments, two or more separate input compositions are isolated, selected, enriched, or obtained from a single biological sample. In some embodiments, the isolated input composition is isolated, selected, enriched, and/or obtained from an isolated biological sample collected, obtained, and/or obtained from the same subject.
In certain embodiments, the one or more input compositions are or include enriched T cell compositions comprising at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99%, at least 99.5%, at least 99.9%, or equal to or about 100% cd3+ T cells. In one embodiment, the enriched T cell input composition consists essentially of cd3+ T cells.
In certain embodiments, the one or more import compositions are or include enriched cd4+ T cell compositions comprising at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99%, at least 99.5%, at least 99.9%, or equal to or about 100% cd4+ T cells. In certain embodiments, the input composition of cd4+ T cells comprises less than 40%, less than 35%, less than 30%, less than 25%, less than 20%, less than 15%, less than 10%, less than 5%, less than 1%, less than 0.1%, or less than 0.01% cd8+ T cells, and/or is free of cd8+ T cells, and/or is free or substantially free of cd8+ T cells. In some embodiments, the composition of enriched T cells consists essentially of cd4+ T cells.
In certain embodiments, the one or more compositions are or comprise cd8+ T cells, which are or comprise at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99%, at least 99.5%, at least 99.9%, or equal to or about 100% cd8+ T cells. In certain embodiments, the composition of cd8+ T cells contains less than 40%, less than 35%, less than 30%, less than 25%, less than 20%, less than 15%, less than 10%, less than 5%, less than 1%, less than 0.1%, or less than 0.01% of cd4+ T cells, and/or is free or substantially free of cd4+ T cells. In some embodiments, the composition of enriched T cells consists essentially of cd8+ T cells.
In some embodiments, the cells are incubated and/or cultured prior to or in combination with genetic engineering. The incubation step may include incubation, culturing, stimulation, activation, and/or propagation. Incubation and/or engineering may be performed in a culture vessel, such as a unit, chamber, well, column, tube set, valve, vial, petri dish, bag, or other vessel for culturing or culturing cells. In some embodiments, the composition or cell is incubated in the presence of a stimulating condition or a stimulating agent. Such conditions include those designed to induce proliferation, expansion, activation and/or survival of cells in the population, mimic antigen exposure and/or elicit genetic engineering of cells, such as for the introduction of recombinant antigen receptors. The conditions may include one or more of a particular medium, temperature, oxygen content, carbon dioxide content, time, agent (e.g., nutrient, amino acid, antibiotic, ion) and/or stimulating factor (such as a cytokine, chemokine, antigen, binding partner, fusion protein, recombinant soluble receptor) and any other agent designed to activate the cell.
In some embodiments, the stimulation conditions or agents include one or more agents, e.g., ligands, capable of stimulating or activating the intracellular signaling domain of the TCR complex. In some embodiments, the agent turns on or initiates a TCR/CD3 intracellular signaling cascade in the T cell. Such agents may include antibodies, such as those specific for TCRs, e.g., anti-CD 3. In some embodiments, the stimulation conditions include one or more agents, such as ligands, capable of stimulating a co-stimulatory receptor, such as anti-CD 28. In some embodiments, such reagents and/or ligands may be bound to a solid support such as a bead and/or one or more cytokines. Optionally, the amplification method may further comprise the step of adding anti-CD 3 and/or anti-CD 28 antibodies to the culture medium (e.g., at a concentration of at least about 0.5 ng/mL). In some embodiments, the stimulatory agent includes IL-2, IL-15 and/or IL-7. In some embodiments, the IL-2 concentration is at least about 10 units/mL. In some embodiments, according to U.S. patent No. 6,040,177, such as Riddell et al; klebaroff et al (2012) J Immunother.35 (9): 651-660; terakura et al (2012) blood.1:72-82; and/or Wang et al (2012) J Immunother35 (9): 689-701.
In some embodiments, T cells are expanded by: adding feeder cells, such as non-dividing Peripheral Blood Mononuclear Cells (PBMCs), to the culture starting composition (e.g., such that the resulting cell population contains at least about 5, 10, 20, or 40 or more PBMC feeder cells for each T lymphocyte in the initial population to be expanded); and incubating the culture (e.g., for a time sufficient to expand the number of T cells). In some embodiments, the non-dividing feeder cells may comprise gamma-irradiated PBMC feeder cells. In some embodiments, the PBMCs are irradiated with gamma rays in the range of about 3000 to 3600 rads to prevent cell division. In some embodiments, the feeder cells are added to the culture medium prior to the addition of the T cell population.
In some embodiments, the stimulation conditions include a temperature suitable for growth of human T lymphocytes, for example, at least about 25 degrees celsius, typically at least about 30 degrees celsius, and typically at or about 37 degrees celsius. Optionally, the incubating may further comprise adding non-dividing EBV-transformed Lymphoblastoid Cells (LCLs) as feeder cells. The LCL may be irradiated with gamma rays in the range of about 6000 to 10,000 rads. In some embodiments, the LCL feeder cells are provided in any suitable amount, such as a ratio of LCL feeder cells to naive T lymphocytes of at least about 10:1.
In embodiments, antigen-specific T cells, such as antigen-specific cd4+ and/or cd8+ T cells, are obtained by stimulating naive or antigen-specific T lymphocytes with an antigen. For example, antigen-specific T cell lines or clones directed against cytomegalovirus antigens can be generated by isolating T cells from an infected subject and stimulating the cells in vitro with the same antigen.
In some embodiments, at least a portion of the incubation in the presence of one or more stimulation conditions or agents is performed in the lumen of the centrifugal chamber, e.g., under centrifugal rotation, such as described in international publication No. WO 2016/073602. In some embodiments, at least a portion of the incubation performed in the centrifugal chamber comprises mixing with one or more agents to induce stimulation and/or activation. In some embodiments, cells (such as selected cells) are mixed with a stimulating condition or agent in a centrifugal chamber. In some embodiments of such methods, a volume of cells is mixed with an amount of one or more stimulation conditions or agents that is much less than the amount typically employed when performing similar stimulation in a cell culture plate or other system.
In some embodiments, when selected in the chamber without mixing (e.g., in a tube or bag with periodic shaking or rotation), the stimulus is added to the cells in the chamber in an amount substantially less than (e.g., no more than 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, or 80%) of the amount of stimulus necessary to achieve about the same or similar selection efficiency for the same number of cells or the same volume of cells as is commonly used or achieved. In some embodiments, the incubation is performed with the addition of incubation buffer to the cells and the stimulating agent to achieve a target volume of reagent incubation, e.g., 10mL to 200mL, such as at least or about or 10mL, 20mL, 30mL, 40mL, 50mL, 60mL, 70mL, 80mL, 90mL, 100mL, 150mL, or 200mL. In some embodiments, the incubation buffer and the stimulating agent are pre-mixed prior to addition to the cells. In some embodiments, the incubation buffer and the stimulating agent are added separately to the cells. In some embodiments, the stimulation incubation is performed under periodic gentle mixing conditions, which may help promote energetically favorable interactions, allowing for less total stimulator to be used while achieving stimulation and activation of the cells.
In some embodiments, the incubation is typically performed under mixing conditions, such as in the presence of rotation, typically at a relatively low force or speed, such as below the speed used to precipitate the cells, such as at or about 600rpm to 1700rpm (e.g., at or about or at least 600rpm, 1000rpm, or 1500rpm or 1700 rpm), such as at or about 80g to l00g (e.g., at or about or at least 80g, 85g, 90g, 95g, or 100 g) of RCF at the sample or wall of the chamber or other container. In some embodiments, rotation is used at repeated intervals of such low speed rotation followed by rest periods, such as rotation and/or rest for 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 seconds, such as rotation at about 1 or 2 seconds followed by rest for about 5, 6, 7, or 8 seconds.
In some embodiments, for example, the total duration of incubation with the stimulus is or is about 1 hour to 96 hours, 1 hour to 72 hours, 1 hour to 48 hours, 4 hours to 36 hours, 8 hours to 30 hours, or 12 hours to 24 hours, such as at least or about at least 6 hours, 12 hours, 18 hours, 24 hours, 36 hours, or 72 hours. In some embodiments, the further incubation is at or about 1 hour to 48 hours, 4 hours to 36 hours, 8 hours to 30 hours, or 12 hours to 24 hours (inclusive).
In some embodiments, the stimulation conditions include incubating, and/or culturing the composition of enriched T cells with and/or in the presence of one or more cytokines. In particular embodiments, the one or more cytokines are recombinant cytokines. In some embodiments, the one or more cytokines are human recombinant cytokines. In certain embodiments, one or more cytokines bind to and/or are capable of binding to receptors expressed by and/or endogenous to T cells. In particular embodiments, the one or more cytokines are or include members of the 4-alpha-helix bundle family of cytokines. In some embodiments, members of the 4- α -helix bundle family of cytokines include, but are not limited to, interleukin-2 (IL-2), interleukin-4 (IL-4), interleukin-7 (IL-7), interleukin-9 (IL-9), interleukin 12 (IL-12), interleukin 15 (IL-15), granulocyte colony-stimulating factor (G-CSF), and granulocyte-macrophage colony-stimulating factor (GM-CSF). In some embodiments, the stimulation results in activation and/or proliferation of the cells, e.g., prior to transduction.
In some embodiments, an engineered cell (such as a T cell) used in conjunction with the provided methods, uses, articles, or compositions is a cell that has been genetically engineered to express a recombinant receptor (e.g., a CAR or TCR described herein). In some embodiments, the cell is engineered by the introduction, delivery, or transfer of nucleic acid sequences encoding recombinant receptors and/or other molecules. In some embodiments, a method for producing an engineered cell comprises introducing a polynucleotide encoding a recombinant receptor (e.g., an anti-CD 19 CAR) into a cell, e.g., a stimulated or activated cell. In particular embodiments, the recombinant protein is a recombinant receptor, such as any described. Nucleic acid molecules encoding recombinant proteins, such as recombinant receptors, can be introduced into cells using any of a variety of known vectors. Such vectors include viral and nonviral systems, including lentiviral and gamma retroviral systems, as well as transposon-based systems, such as PiggyBac or sleep Beauy-based gene transfer systems. Exemplary methods include methods for transferring nucleic acids encoding a receptor, including by virus (e.g., retrovirus or lentivirus), transduction, transposon, and electroporation. In some embodiments, engineering results in one or more engineered enriched T cell compositions.
In certain embodiments, the one or more stimulated T cell compositions are or include two separate stimulated enriched T cell compositions. In some embodiments, two separate enriched T cell compositions, e.g., two separate enriched T cell compositions that have been selected, isolated, and/or enriched from the same biological sample, are engineered separately. In certain embodiments, the two separate compositions comprise enriched compositions of cd4+ T cells. In some embodiments, the two separate compositions comprise a composition of enriched cd8+ T cells. In some embodiments, the two separate compositions of enriched cd4+ T cells and enriched cd8+ T cells are genetically engineered separately. In some embodiments, the same composition is enriched for both cd4+ T cells and cd8+ T cells, and these cells are genetically engineered together.
In some embodiments, the composition comprising the engineered T-cells comprises a pharmaceutically acceptable carrier, diluent, solubilizer, emulsifier, preservative, and/or adjuvant. In some embodiments, the composition comprises an excipient. By "pharmaceutically acceptable carrier" is meant an ingredient of the pharmaceutical formulation that is non-toxic to the subject, other than the active ingredient. Pharmaceutically acceptable carriers include, but are not limited to, buffers, excipients, stabilizers, or preservatives.
In some embodiments, the composition is selected for parenteral delivery, for inhalation, or for delivery through the digestive tract, such as oral administration. The preparation of such pharmaceutically acceptable compositions is within the ability of those skilled in the art. In some embodiments, buffers are used to maintain the composition at physiological pH or slightly lower, typically in the 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 in a pharmaceutically acceptable vehicle, the aqueous solution comprising the composition described herein, with or without an additional therapeutic agent. In some embodiments, the vehicle for parenteral injection is sterile distilled water in which the compositions described herein are formulated as a sterile isotonic solution for suitable storage, with or without at least one additional therapeutic agent. In some embodiments, the preparation involves formulating the desired molecule with a polymeric compound (such as polylactic acid or polyglycolic acid), beads, or liposomes to provide controlled or sustained release of the product, which is then delivered by depot injection. In some embodiments, implantable drug delivery devices are used to introduce the desired molecules.
In some embodiments, the method of treating cancer in a subject in need thereof comprises T cell therapy. In some embodiments, the T cell therapies disclosed herein are engineered autologous cell therapies (eACT TM ). According to this embodiment, the method may comprise collecting blood cells from the patient. The isolated blood cells (e.g., T cells) can then be engineered to express the CARs disclosed herein. In particular embodiments, the CAR T cells are administered to a patient. In some embodiments, the CAR T cells treat a tumor or cancer in a patient. In some embodiments, the CAR T cells reduce the size of a tumor or cancer.
In some embodiments, donor T cells for T cell therapy (e.g., for autologous T cell therapy) are obtained from a patient. In other embodiments, donor T cells for T cell therapy are obtained from a subject other than a patient. In certain embodiments, the T cell is a Tumor Infiltrating Lymphocyte (TIL), an engineered autologous T cell (eACT TM ) An allogeneic T cell, a xenogeneic T cell, or any combination thereof.
In some embodiments, the engineered T cells are administered in a therapeutically effective amount. For example, the treatment is An effective amount of engineered T cells may be at least about 10 4 Individual cells, at least about 10 5 Individual cells, at least about 10 6 Individual cells, at least about 10 7 Individual cells, at least about 10 8 Individual cells, at least about 10 9 And/or at least about 10 10 And each. In another embodiment, the therapeutically effective amount of T cells is about 10 4 Individual cells, about 10 5 Individual cells, about 10 6 Individual cells, about 10 7 Individual cells or about 10 8 Individual cells. In some embodiments, the therapeutically effective amount of T cells is about 2 x 10 6 Individual cells/kg, about 3X 10 6 Individual cells/kg, about 4X 10 6 Individual cells/kg, about 5X 10 6 Individual cells/kg, about 6X 10 6 Individual cells/kg, about 7X 10 6 Individual cells/kg, about 8X 10 6 Individual cells/kg, about 9X 10 6 Individual cells/kg, about 1X 10 7 Individual cells/kg, about 2X 10 7 Individual cells/kg, about 3X 10 7 Individual cells/kg, about 4X 10 7 Individual cells/kg, about 5X 10 7 Individual cells/kg, about 6X 10 7 Individual cells/kg, about 7X 10 7 Individual cells/kg, about 8X 10 7 Individual cells/kg or about 9X 10 7 Individual cells/kg.
In some embodiments, a therapeutically effective amount of engineered live T cells is between about 1X 10 per kg body weight 6 And about 2X 10 6 Between engineered live T cells up to about 1X 10 8 Maximum dose of individual engineered live T cells. In some embodiments, the engineered T cell is an anti-CD 19 CART T cell. In some embodiments, the anti-CD 19 CAR T cell is an alemtuquor product. In some embodiments, the anti-CD 19 CAR T cell is a brexucabtagene autoleucel product, also known as KTE-X19.
Therapeutic method
The methods disclosed herein can be used to treat cancer in a subject, reduce the size of a tumor, kill tumor cells, prevent proliferation of tumor cells, prevent growth of a tumor, eliminate a tumor in a patient, prevent recurrence of a tumor, prevent metastasis of a tumor, induce remission in a patient, or any combination thereof. In some embodiments, the method induces a complete response. In other embodiments, the method induces a partial response.
Treatable cancers include non-vascularized, insufficiently vascularized or vascularized tumors. Cancers may also include solid or non-solid tumors. In some embodiments, the cancer is a hematologic cancer. In some embodiments, the cancer is a white blood cell cancer. In other embodiments, the cancer is a cancer of plasma cells. In some embodiments, the cancer is leukemia, lymphoma, or myeloma. In some embodiments, the cancer is Acute Lymphoblastic Leukemia (ALL) (including non-T cell ALL), acute Lymphoblastic Leukemia (ALL), and hemophagocytic lymphoproliferative disorder (HLH)), B cell prolymphocytic leukemia, B cell acute lymphoblastic leukemia ("BALL"), blast plasmacytoid dendritic cell tumor, burkitt lymphoma, chronic Lymphocytic Leukemia (CLL), chronic Myelogenous Leukemia (CML), chronic or acute granulomatosis, chronic or acute leukemia, 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 disorder, burkitt lymphoma, mantle Cell Lymphoma (MCL), marginal Zone Lymphoma (MZL), monoclonal promyelocytic leukemia (mgl), myelodysplastic disorder (MGUS), myelodysplasia (malhyperplasia), myelodysplasia (MDS), myelodysplasia (non-myelomas), myelomatosis (MDS), myelodysplastic Disease (MDS), myelosis (myelodysplastic disease (MDS), and myelosis (myelodysplastic disease (non-myelosis), asymptomatic myeloma (stasis type multiple myeloma or indolent myeloma), plasmablasts, plasmacytoid lymphomas, plasmacytoid dendritic cell neoplasms, plasmacytoma (e.g., plasmacytoid; isolated myeloma; isolated plasmacytoma; extramedullary plasmacytoma and multiple plasmacytoma), POEMS syndrome (Crow-Fukase syndrome; takatsuki disease; PEP syndrome), primary mediastinal large B-cell lymphoma (PMBC), small 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 macroglobulinemia, or combinations thereof.
In some embodiments, the cancer is myeloma. In some embodiments, the cancer is multiple myeloma. In some embodiments, the cancer is leukemia. In some embodiments, the cancer is acute myelogenous leukemia.
In some embodiments, the cancer is non-hodgkin's lymphoma. In some embodiments, the cancer is relapsed/refractory NHL. In some embodiments, the cancer is mantle cell lymphoma.
In some embodiments, the cancer is advanced indolent non-hodgkin's lymphoma (iNHL), including Follicular Lymphoma (FL) and Marginal Zone Lymphoma (MZL). In some embodiments, the patient has recurrent/refractory disease after ≡2 past line number therapies (including anti-CD 20 monoclonal antibodies and alkylating agents). In some embodiments, the patient may have received a PI3K inhibitor. In some embodiments, the patient may (also) have received autologous stem cell transplantation. In some embodiments, the patient undergoes leukocyte apheresis to obtain T cells for CAR T cell manufacturing, followed by administration of 500mg/m on day-5, day-4, and day-3 2 Cyclophosphamide per day and 30mg/m 2 Fludarabine/day for conditioning chemotherapy; on day 0, the patient may receive a target dose of 2X 10 6 Single intravenous infusion of CAR T cell therapy (e.g., alemtuzite) of individual CAR T cells/kg. In some embodiments, additional infusions may be administered at a later time. In some embodiments, if the patient develops post-response progression at the time of evaluation at month 3 following initial administration, the patient can receive a retreatment of CAR T cell therapy (e.g., alemtuzite). In some embodiments, the patient may receive bridging therapy. Examples of bridging therapies include dexamethasone, rituximab (Rituximab), etoposide, carboplatin, ifosfamide, bendamustine-Rituximab (Bendamustine-Rituximab), bendamustine, methylprednisolone, mitoxantrone, cyclophosphamide, fludarabine, ibrutinib (ibretinib). In some embodiments, the patient experiences CRS. In some implementations, CRS is managed using any of the protocols described in this application (including the examples). In some embodiments, CRS is managed with tolizumab, corticosteroids, and/or vasopressors.
In some embodiments, the cancer is a relapsed/refractory indolent non-hodgkin lymphoma and the method of treating a subject in need thereof comprises administering to the subject a therapeutically effective amount of CAR T cells as a retreatment, wherein the subject has previously received a first treatment with CAR T cells. In some embodiments, the first treatment with CAR T cells can be administered as a first line therapy or a second line therapy, optionally wherein the lymphoma is R/R Follicular Lymphoma (FL) or Marginal Zone Lymphoma (MZL), and optionally wherein the previous prior line number therapy comprises an anti-CD 20 monoclonal antibody in combination with an alkylating agent. In some embodiments, the opsonic therapy comprises 30mg/m of IV fludarabine on day-5, day-4 and day-3 2 And IV cyclophosphamide 500mg/m 2 . In some embodiments, the CAR T cell therapy comprises a single IV infusion of 2 x 10 on day 0 6 Individual CAR T cells/kg. In some embodiments, at least about 10 is administered 4 Individual cells, at least about 10 5 Individual cells, at least about 10 6 Individual cells, at least about 10 7 Individual cells, at least about 10 8 Individual cells, at least about 10 9 Or at least about 10 10 And (3) CAR T cells. In another embodiment, the therapeutically effective amount of T cells is about 10 4 Individual cells, about 10 5 Individual cells, about 10 6 Individual cells, about 10 7 Individual cells or about 10 8 Individual cells. In some embodiments, the therapeutically effective amount of T cells is about 2 x 10 6 Individual cells/kg, about 3X 10 6 Individual cells/kg, about 4X 10 6 Individual cells/kg, about 5X 10 6 Individual cells/kg, about 6X 10 6 Individual cells/kg, about 7X 10 6 Individual cells/kg, about 8X 10 6 Individual cells/kg, about 9X 10 6 Individual cells/kg, about 1X 10 7 Individual cells/kg, about 2X 10 7 Individual cells/kg, about 3X 10 7 Individual cells/kg, about 4X 10 7 Individual cells/kg, about 5X 10 7 Individual cells/kg, about 6X 10 7 Individual cells/kg, about 7X 10 7 Individual cells/kg, about 8X 10 7 Individual cells/kg or about 9X 10 7 Individual cells/kg. In some embodiments, the CAR T is an anti-CD 19 CAR T cell. In some embodiments, the CAR T cell is an alemtuzite CAR T cell. In some embodiments, the CAR T cell is brexucabtagene autoleucel/KTE-X19. In some embodiments, the eligibility criterion for retreatment includes response and subsequent progression of CR or PR at the time of the 3 rd month disease assessment; evidence of no CD19 loss in progressive biopsies by local examination; and/or no grade 4 CRS or neurological events or life-threatening toxicities were present at the first treatment with CAR T cells. In some embodiments, the method of treatment is a method following the CLINICAL TRIAL-2 clinical trial (NCT 02601313). In some embodiments, the method of treatment is a method following the CLINICAL TRIAL-5 clinical trial (NCT 03105336). In some embodiments, the method of treatment is a method following the clinical trial-1 (NCT 02348216) clinical trial, which has been described in detail in Neelapu, SS et al 2017,N Engl J Med 2017;377 (26) 2531-44) and Locke et al Lancet Oncol.2019; 20:31-42.
In some embodiments, the cancer is NHL and the CAR T cell therapy is administered as a first line therapy. In some embodiments, the cancer is LBCL. In some embodiments, the LBCL is a high risk/high level LBCL with MYC and BCL2 and/or BCL6 translocation or a DLBCL with IPI score ≡3 at any time prior to group entry. In some embodiments, the first line therapy comprises the combination of CAR T cell therapy with an anti-CD 20 monoclonal antibody and anthracycline-containing regimen. In some embodiments, CAR T cell therapy is administered first. In some embodiments, an anti-CD 20 monoclonal antibody/anthracycline-containing regimen is administered first. In some embodiments, the treatment is administered for at least 2 weeks, at least 4 weeks, at least 6 weeks, at least 1 month, at least 2 months, at least 3 months, at least 4 months, at least 5 months, less than one year apart, or the like. In some embodiments, the method further comprises a bridging therapy administered after the white blood cell apheresis and completed prior to the initiation of the conditioning chemotherapy. In some embodimentsIn the scheme, additional inclusion criteria include age.gtoreq.18 years and ECOGGS 0-1. In some embodiments, the opsonic therapy comprises 30mg/m of IV fludarabine on day-5, day-4 and day-3 2 And IV cyclophosphamide 500mg/m 2 . Other exemplary beneficial preconditioning treatment regimens are described in U.S. provisional patent applications 62/262,143 and 62/167,750, and U.S. patent nos. 9,855,298 and 10,322,146, the entire contents of which are hereby incorporated by reference. These provisional patent applications describe, for example, methods of modulating a patient in need of T cell therapy comprising administering to the patient a prescribed beneficial amount of cyclophosphamide (200 mg/m 2 Day and 2000mg/m 2 Between/day) and the prescribed dose of fludarabine (20 mg/m 2 Day and 900mg/m 2 Between days). One such dosage regimen involves treating a patient, including administering to the patient about 500mg/m daily for three days prior to administering to the patient a therapeutically effective amount of engineered T cells 2 Cyclophosphamide per day and about 60mg/m 2 Fludarabine per day. Another embodiment includes serum cyclophosphamide and fludarabine at-4, -3 and-2 days prior to T cell administration at a dose of 500mg/m during this period 2 Cyclophosphamide with body surface area per day and dose of 30mg/m 2 Fludarabine in body surface area per day. Another embodiment includes cyclophosphamide at day-2 and fludarabine at day-4, day-3 and day-2 prior to T cell administration at a dose of 900mg/m during this period 2 Cyclophosphamide with body surface area and dose of 25mg/m 2 Fludarabine in body surface area per day. In another embodiment, the modulation comprises serum cyclophosphamide and fludarabine at a dose of 500mg/m for a period of time of-5, 4 and 3 days prior to T cell administration 2 Cyclophosphamide with body surface area per day and dose of 30mg/m 2 Fludarabine in body surface area per day. Other preconditioning embodiments include 200-300mg/m 2 Cyclophosphamide with surface area per day and dosage of 20-50mg/m 2 Fludarabine in body surface area per day for three days. Clinical trial-1 (NCT 02348216) clinical trials have been described in detail in the examples and Neelapu, SS et al 2017,N Engl J Med 2017;377 (26):2531-44) and Locke et al Lancet Oncol.2019; 20:31-42.
In some embodiments, the method of treatment comprises administering immune cells in combination with other therapeutic agents or treatments (e.g., radiation, debulking). In some embodiments, additional therapeutic agents or treatments are included to manage adverse events. In some embodiments, additional therapeutic agents or treatments are included to improve the therapeutic efficacy of cellular therapies. In some embodiments, they achieve both. Examples of therapeutic agents that may be used with immune cells (before, after, and/or simultaneously) are provided below and elsewhere in the specification.
In some embodiments, the method further comprises administering a chemotherapeutic agent. In some embodiments, the chemotherapeutic agent selected is a lymphoablative (preconditioning) chemotherapeutic agent. Beneficial preconditioning treatment regimens are described in U.S. provisional patent applications 62/262,143 and 62/167,750 and U.S. patent nos. 9,855,298 and 10,322,146, which provisional patent applications are hereby incorporated by reference in their entirety, along with related beneficial biomarkers. These provisional patent applications describe, for example, methods of modulating a patient in need of T cell therapy comprising administering to the patient a prescribed beneficial amount of cyclophosphamide (200 mg/m 2 Day and 2000mg/m 2 Between/day) and the prescribed dose of fludarabine (20 mg/m 2 Day and 900mg/m 2 Between days). One such dosage regimen involves treating a patient, including administering to the patient about 500mg/m daily for three days prior to administering to the patient a therapeutically effective amount of engineered T cells 2 Cyclophosphamide per day and about 60mg/m 2 Fludarabine per day. Another conditioning embodiment includes cyclophosphamide 500mg/m at day-5, day-4 and day-3 2 Day/day and fludarabine 30mg/m 2 Day/for 3 days. Another embodiment includes serum cyclophosphamide and fludarabine at-4, -3 and-2 days prior to T cell administration at a dose of 500mg/m during this period 2 Cyclophosphamide with body surface area per day and dose of 30mg/m 2 Fludarabine in body surface area per day. Another embodiment includes cyclophosphamide on day-2 and cyclophosphamide on day-4 prior to T cell administration,Fludarabine at day-3 and day-2, at a dose of 900mg/m during this period 2 Cyclophosphamide with body surface area and dose of 25mg/m 2 Fludarabine in body surface area per day. In another embodiment, the modulation comprises serum cyclophosphamide and fludarabine at a dose of 500mg/m for a period of time of-5, 4 and 3 days prior to T cell administration 2 Cyclophosphamide with body surface area per day and dose of 30mg/m 2 Fludarabine in body surface area per day. Another embodiment includes 300mg/m per day 2 Cyclophosphamide for 3 days, and 30mg/m per day 2 Fludarabine for 3 days. Another embodiment comprises administering fludarabine (30 mg/m) 2 to 7 days prior to T cell administration for 3 days 2 ) And cyclophosphamide (300 mg/m) 2 ). Another embodiment includes 3000mg/m once on day-7 to day-2 2 Cyclophosphamide. Another embodiment includes 300mg/m per day 2 Cyclophosphamide 30mg/m daily for 3 days 2 Fludarabine last for 3 days, with the last day occurring on days-7 to-2.
In some embodiments, the antigen binding molecule, transduced (or otherwise engineered) cells (such as a CAR), and chemotherapeutic agent are each administered in an amount effective to treat the disease or disorder in the subject.
In some embodiments, a composition comprising an immune cell (e.g., CAR-expressing immune effector cell) disclosed herein can be administered in combination with any number of other chemotherapeutic agents and/or radiation (before, after, and/or simultaneously with administration of the immune cell). Examples of chemotherapeutic agents include alkylating agents such as thiotepa and Cyclophosphamide (CYTOXAN) TM ) The method comprises the steps of carrying out a first treatment on the surface of the Alkyl sulfonates such as busulfan, imperoshu and piposhu; aziridines such as benzotepa, carboquinone, rituximab and uratepa; ethyleneimine and methyl melamines, including altretamine, triethylenemelamine, triethylenephosphoramide, triethylenethiophosphamide, and trimethylol melamine; nitrogen mustards such as chlorambucil, napthalene mustards, cholesteryl phosphoramide, estramustine, ifosfamide, dichloromethyl diethylamine, mechlorethamine hydrochloride, melphalan, chlorambucil cholesterol, prednisomusA statin, trefosfamine, uracil mustard; nitrosoureas such as carmustine, chlorourea, fotemustine, lomustine, nimustine, and ramustine; antibiotics such as aclacinomycin, actinomycin, anthracycline, diazoserine, bleomycin, actinomycin C, calicheamicin, carminomycin, amphotericin, chromomycin, actinomycin D, daunorubicin, ditubicin, 6-diazo-5-oxo-L-norleucine, doxorubicin, epirubicin, idarubicin, doxycycline, mitomycin, mycophenolic acid, nolamycin, olivomycin, pelomycin, pofeomycin, puromycin, tri-iron doxorubicin, rodobicubicin, streptozocin, tubercidin, ubenimex, jistatin, zorubicin; antimetabolites such as methotrexate and 5-fluorouracil (5-FU); folic acid analogs such as, for example, dimethyl folic acid, methotrexate, pterin, trimetrexate; purine analogs such as fludarabine, 6-mercaptopurine, thioxanthine, thioguanine; pyrimidine analogs such as ambcitabine, azacytidine, 6-azauridine, carmofur, cytarabine, dideoxyuridine, deoxyfluorouridine, enocitabine, fluorouridine, 5-FU; androgens such as carbosterone, drotasone propionate, cyclothioandrostanol, emaandran, testosterone; anti-epinephrine such as aminoglutethimide, mitotane, trilostane; folic acid supplements such as folinic acid; acetoglucurolactone; aldehyde phosphoramide glycosides; aminolevulinic acid; amsacrine; a method of treating a patient with a tumor; a specific group; eda traxas; a phosphoramide; colchicine; deaquinone; erlotinib (elformithin); ammonium elegance; eggshell robust; gallium nitrate; hydroxyurea; lentinan; lonidamine; mitoguazone; mitoxantrone; mo Pai dar alcohol; diamine nitroacridine; prastatin; egg ammonia nitrogen mustard; pirarubicin; podophylloic acid; 2-ethyl hydrazide; procarbazine; polysaccharide K (PSK); carrying out a process of preparing the raw materials; a sirzopyran; germanium spiroamine; tenuazonic acid; triiminoquinone; 2,2',2 "-trichlorotriethylamine; uratam; vindesine; dacarbazine; mannitol; dibromomannitol; dibromodulcitol; pipobromine; gacetin (gacytosine); arabinoside ("Ara-C"); cyclophosphamide; thiotepa; taxus chinensis Alkanes, e.g. TAXOL (TAXOL) TM Bristol-Myers Squibb) and docetaxel (doxetaxel)Rhone-Poulenc Rorer); chlorambucil; gemcitabine; 6-thioguanine; mercaptopurine; methotrexate; platinum analogs such as cisplatin and carboplatin; vinblastine; platinum; etoposide (VP-16); ifosfamide; mitomycin C; mitoxantrone; vincristine; vinorelbine; north vitamin; norxiaoling; teniposide; daunomycin; aminopterin; hilded; ibandronate; CPT-11; topoisomerase inhibitor RFS2000; difluoromethyl ornithine (DMFO); retinoic acid derivatives such as tartretin TM (Besalutin), panretin TM (aliskiric acid); ONTAK (optical network Unit) TM (diniinterleukin); esperamicin Mi Mei; capecitabine; and pharmaceutically acceptable salts, acids or derivatives of any of the above. In some embodiments, compositions comprising the CAR-expressing immune effector cells disclosed herein can be administered in combination with an anti-hormonal agent, such as an anti-estrogen, for example, including tamoxifen, raloxifene, aromatase-inhibiting 4 (5) -imidazole, 4-hydroxy tamoxifen, trawoxifene, raloxifene hydrochloride, LY117018, onapristone, and toremifene (faropenem), for modulating or inhibiting hormonal effects on tumors; and antiandrogens such as flutamide, nilutamide, bicalutamide, leuprorelin, and goserelin; and pharmaceutically acceptable salts, acids or derivatives of any of the above. Combinations of chemotherapeutic agents, including but not limited to CHOP, i.e., cyclophosphamide +. >Doxorubicin (hydroxy doxorubicin), vincristine +.>And prednisone, R-CHOP (CHOP plus rituximab), and G-CHOP (CHOP plus oxtuzumab).
In some embodiments, the chemotherapeutic agent is administered concurrently with or within one week of administration of the engineered immune cell. In other embodiments, the chemotherapeutic agent is administered 1 to 4 weeks or 1 week to 1 month, 1 week to 2 months, 1 week to 3 months, 1 week to 6 months, 1 week to 9 months or 1 week to 12 months after the engineered cell or nucleic acid is administered. In some embodiments, the chemotherapeutic agent is administered at least 1 month prior to administration of the cell or nucleic acid. In some embodiments, the method further comprises administering two or more chemotherapeutic agents.
A variety of additional therapeutic agents may be used in combination with the compositions described herein (before, after, and/or simultaneously with T cell administration). For example, potentially useful additional therapeutic agents include PD-1 inhibitors, such as nivolumabParbolizumab +>Cimetidine Li Shan antibody (Libtayo), pilates bead mab (CureTech) and actelib mab (roche), as well as PD-L1 inhibitors such as actelib mab, dulcit You Shan antibody and avermectin.
Additional therapeutic agents suitable for use in combination with the compositions and methods disclosed herein (before, after, and/or concurrently with T cell administration) include, but are not limited to ibrutinib AofatuzumabRituximab->Bevacizumab->Trastuzumab->Enmetrastuzumab +.>ImatinibCetuximab->Panitumumab->Katuxostat, temozolomide, ofatuzumab, tositumomab, bentuximab, alemtuzumab, gemtuzumab, erlotinib, gefitinib, vandetanib, afatinib, lapatinib, sunitinib, sorafenib, torantinib, letatinib, axitinib, ceritinib, ceridinib, lenvatinib, cerivatinib, panoratinib, regrinanib, simansamib, sorafenib, sunitinib, tivalanib, tulathanib, vandaltetinib, entriptinib, vande, emtrictinib cabotinib, imatinib, dasatinib, nilotinib, ponatinib, radatinib, bosutinib, letatinib, lu Suoti, panatinib, cobratinib, sematinib, tramatinib, bimatinib, ai Leti, ceritinib, crizotinib, aflibercept, arabino peptide (adiotoide), diltiazem, mTOR inhibitors such as everolimus and temsirolimus, hedgehog inhibitors such as sonidegiand valimod gedy, CDK inhibitors such as CDK inhibitors (palbociclib), GM-CSF, CSF1, GM-CSFR or inhibitors of CSF1R, anti-thymus cytoglobulin, lorentz mab and mefloumab.
In one embodiment, the GM-CSF inhibitor is selected from the group consisting of lorentzumab; nalmefene mab (AMG 203); GSK3196165/MOR 103/octreotide Li Shan antibody (GSK/MorphoSys); KB002 and KB003 (KaloBios); MT203 (Micromet and Nycomed); MORAb-022/bemycetin (morphek); or any of them; E21R; and small molecules. In one embodiment, the CSF1 inhibitor is selected from RG7155, PD-0360324, MCS 110/rituximab or a bioimitated pharmaceutical form of any of them; and small molecules. In one embodiment, the GM-CSFR inhibitor and the CSF1R inhibitor are selected from the group consisting of MAFreimumab (previously known as CAM-3001; medImmune, inc.); calicheamicin (Five Prime Therapeutics); LY3022855 (IMC-CS 4) (Gift corporation), event-Mi Tuozhu mab, also known as RG7155 or RO5509554; FPA008 (Five Prime/BMS); AMG820 (security company); ARRY-382 (Array Biopharma); MCS110 (nowa); PLX3397 (Plexxikon); ELB041/AFS98/TG3003 (ElsalLys Bio, transgene), SNDX-6352 (Syndax); a bioimitated pharmaceutical form of any of them; and small molecules.
In some embodiments, the agent is administered by injection, such as intravenous or subcutaneous injection, intraocular injection, periocular injection, subretinal injection, intravitreal injection, transseptal injection, subscleral injection, intracoronary injection, intracameral injection, subconjunctival injection (subconjectval injection, subconjuntival injection), sub-tenon's capsule injection, retrobulbar injection, peribulbar injection, or posterior juxtascleral delivery. In some embodiments, they are administered parenterally, intrapulmonary, and intranasally, and intralesionally, if local therapy is desired. Parenteral infusion includes intramuscular, intravenous, intraarterial, intraperitoneal or subcutaneous administration.
In some embodiments, the treatment further comprises a bridging therapy, which is a therapy that modulates between the compositions disclosed herein or is administered after apheresis of the white blood cells and completed prior to initiation of conditioning chemotherapy. In some embodiments, bridging therapies include CHOP, G-CHOP, R-CHOP (rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisolone), corticosteroids, bendamustine, platinum compounds, anthracyclines, and/or phosphoinositide 3-kinase (PI 3K) inhibitors. In some embodiments, the PI3K inhibitor is selected from Du Weili sibirib (duvelisib), ai Deli sibirib (idelalisib), vinetogram (venteclax), pityriasisb (piciliib) (GDC 0941), ke Pan Lixi cloth (copanlisib), PX 866, buparlixib (buparliib) (BKM 120), pi Lala luxib (XL 147), GNE 317, alpeliib Li Xibu (alpeliib) (BYL 719), INK1117, GSK2636771, AZD8186, SAR260301, and tamponade Li Xibu (taseliib) (GDC 0032). In some embodiments, the AKT inhibitor is pirifaxin, MK-2206. In one embodiment, the mTOR inhibitor is selected from everolimus, sirolimus, temsirolimus, li Dafu limus. In some embodiments, the dual PI3K/mTOR inhibitor is selected from BEZ235, XL765, and GDC-0980. In some embodiments, the PI3K inhibitor is selected from Du Weili sibirib (duvelisib), ai Deli sibirib (idelalisib), vinetogram (venteclax), pityriasisb (piciliib) (GDC 0941), ke Pan Lixi cloth (copanlisib), PX 866, buparlixib (buparliib) (BKM 120), pi Lala luxib (XL 147), GNE 317, alpeliib Li Xibu (alpeliib) (BYL 719), INK1117, GSK2636771, AZD8186, SAR260301, and tamponade Li Xibu (taseliib) (GDC 0032).
In some embodiments, the bridging therapy comprises acartinib, bentuximab Shan Kangrui statin, coomassie Pan Lixi brix, nilamide, bemisat, bendamustine hydrochloride, carmustine, bleomycin sulfate, bortezomib, zebutinib, carmustine, chlorambucil, coo Pan Lixi brix hydrochloride, deniinterleukin, dexamethasone, doxorubicin hydrochloride, du Weili sibutramine, prasugrel, omuzumab, temozolomide, ibrutinib, ai Deli siban, recombinant interferon alpha-2 b, romidepsin, lenalidomide, nitrogen mustard hydrochloride, methotrexate, mo Jiamu bead monoclonal antibody-kpc, prasugrel, nimustine-Shan Kangrui bead, mo Jiamu bead monoclonal antibody-kpc, prednisolone, rituximab, eprofuzumab, eprofuzol-52, pervomica, and combinations thereof.
In some embodiments, the cellular immunotherapy is administered in combination with an debulking therapy for the purpose of reducing tumor burden. In one embodiment, the debulking therapy is administered after the white blood cell apheresis and prior to the administration of the conditioning chemotherapy or cell infusion. Examples of subtractive therapies include the following:
Abbreviations: AUC, area under curve
a Other subtractive treatment options may be used and discussed with the medical monitor. Support treatments such as hydration, antiemetic, mesna, growth factor support, and tumor lysis prevention can be used according to local standards. Allowing for more than 1 cycle.
b At least 1 target lesion should remain outside the radiation field to allow tumor measurement
In some embodiments, the composition comprising immune cells (e.g., engineered CAR T cells) is administered with an anti-inflammatory agent (before, after, and/or simultaneously with immune cell administration). Anti-inflammatory agents or agents include, but are not limited to, steroids and glucocorticoids (including betamethasone, budesonide, dexamethasone, hydrocortisone acetate, hydrocortisone, methylprednisolone, prednisolone, prednisone, triamcinolone); non-steroidal anti-inflammatory drugs (NSAIDS) including aspirin, ibuprofen, naproxen, methotrexate, sulfasalazine, leflunomide, anti-TNF drugs, cyclophosphamide and mycophenolate mofetil. Exemplary NSAIDs include ibuprofen, naproxen sodium, cox-2 inhibitors, and sialylates (sialylates). Exemplary analgesics include acetaminophen, oxycodone, tramadol, or propoxyphene hydrochloride. Exemplary glucocorticoids include cortisone, dexamethasone, hydrocortisone, methylprednisolone, prednisolone, or prednisone. Exemplary biological response modifiers include agents directed against cell surface markers (e.g., CD4, CD5, etc.) Molecules, cytokine inhibitors such as TNF antagonists (e.g., etanercept)Adalimumab->And infliximab->Chemokine inhibitors and adhesion molecule inhibitors. Biological response modifiers include monoclonal antibodies and recombinant forms of the molecules. Exemplary DMARDs include azathioprine, cyclophosphamide, cyclosporine, methotrexate, penicillamine, leflunomide, sulfasalazine, hydroxychloroquine, gold formulations (oral (auranofin) and intramuscular), and minocycline.
In some embodiments, the compositions described herein are administered in combination with a cytokine (before, after, or simultaneously with T cell administration). Examples of cytokines are lymphokines, monokines and traditional polypeptide hormones. Cytokines include growth hormone such as human growth hormone, N-methionyl human growth hormone and bovine growth hormone; parathyroid hormone; thyroxine; insulin; proinsulin; relaxin; a relaxin source; glycoprotein hormones such as Follicle Stimulating Hormone (FSH), thyroid Stimulating Hormone (TSH) and Luteinizing Hormone (LH); hepatocyte Growth Factor (HGF); fibroblast Growth Factor (FGF); prolactin; placental lactogen; a mullerian duct inhibiting substance; a mouse gonadotrophin-associated peptide; inhibin; an activin; vascular endothelial growth factor; an integrin; thrombopoietin (TPO); nerve Growth Factor (NGF), such as NGF- β; platelet growth factors; transforming Growth Factors (TGFs), such as TGF- α and TGF- β; insulin-like growth factors-I and-II; erythropoietin (EPO), ) The method comprises the steps of carrying out a first treatment on the surface of the An osteoinductive factor; interferons such as interferons α, β, and γ; colony Stimulating Factors (CSF), such as macrophage-CSF (M-CSF); granulocyte-macrophage-CSF(GM-CSF); and granulocyte-CSF (G-CSF); interleukins (IL), such as IL-1, IL-1α, 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 such as TNF- α or TNF- β; 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 cultures, as well as biologically active equivalents of the native sequence cytokines.
In some embodiments, the administration of the cells and the administration of the additional therapeutic agent are performed on the same day, no more than 36 hours apart, no more than 24 hours apart, no more than 12 hours apart, no more than 6 hours apart, no more than 4 hours apart, no more than 2 hours apart, or no more than 1 hour apart or no more than 30 minutes apart. In some embodiments, the administration of the cell and the administration of the additional therapeutic agent are performed at the following times: between or about 0 and or about 48 hours, between or about 0 and or about 36 hours, between or about 0 and or about 24 hours, between or about 0 and or about 12 hours, between or about 0 and or about 6 hours, between or about 0 and or about 2 hours, between or about 0 and or about 1 hour, between or about 0 and about 1 minute, between or about 0 and about 30 minutes and one or about 48 hours, between or about 30 minutes and one or about 36 hours, between or about 30 minutes and one or about 24 hours, between or about 30 minutes and one or about 12 hours, between or about 30 minutes and one or about 6 hours, between or about 30 minutes and one or about 4 hours, between or about 30 minutes and one or about 2 hours, between or about 30 minutes, between or about 1 hour, about 24 hours, between or about 30 minutes and one or about 2 hours, between or about 30 minutes and one or about 1 hour, between or about 2 hours, between or about 30 minutes and one or about 1 hour, between or about 24 hours, between or about 2 hours, between or about 30 minutes and one or about 4 hours, between or about 24 hours, between or about 2 hours, between or about 30 and one or about 24 hours, between or about 24 and one or about 24 hours, between or about 30 minutes, between or about 30 and one or about 24 hours Between or about 4 hours and or about 6 hours, between or about 6 hours and or about 48 hours, between or about 6 hours and or about 36 hours, between or about 6 hours and or about 24 hours, between or about 6 hours and or about 12 hours, between or about 12 hours and or about 48 hours, between or about 12 hours and or about 36 hours, between or about 12 hours and or about 24 hours, between or about 24 hours and or about 48 hours, between or about 24 hours and or about 36 hours, between or about 24 hours and or between or about 36 hours and or about 48 hours. In some embodiments, the cell and the additional therapeutic agent are administered simultaneously.
In some embodiments, the agent is administered at a dose of about 30mg to 5000mg, such as 50mg to 1000, 50mg to 500, 50mg to 200, 50mg to 100, 100mg to 1000, 100mg to 500, 100mg to 200, 200mg to 1000, 200mg to 500mg, or 500mg to 1000 mg. In some embodiments, the medicament is administered at a dose of 0.5mg/kg to 100mg/kg, 1mg/kg to 50mg/kg, 1mg/kg to 25mg/kg, 1mg/kg to 10mg/kg, 1mg/kg to 5mg/kg, 5mg/kg to 100mg/kg, 5mg/kg to 50mg/kg, 5mg/kg to 25mg/kg, 5mg/kg to 10mg/kg, 10mg/kg to 100mg/kg, 10mg/kg to 50mg/kg, 10mg/kg to 25mg/kg, 25mg/kg to 100mg/kg, 25mg/kg to 50mg/kg to 100 mg/kg. In some embodiments, the agent is administered at a dose of 1mg/kg to 10mg/kg, 2mg/kg to 8mg/kg, 2mg/kg to 6mg/kg, 2mg/kg to 4mg/kg, or 6mg/kg to 8mg/kg, respectively. In some aspects, the agent is administered at a dose of at least 1mg/kg, 2mg/kg, 4mg/kg, 6mg/kg, 8mg/kg, 10mg/kg, or more.
Measuring response and efficacy
In some embodiments, the methods described herein can provide clinical benefit to a subject. In some embodiments, at least 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, or 95% of the patients obtain a clinical benefit. In some embodiments, about 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 0%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90% or 95% and any non-enumerated% of the patients therebetween obtain a clinical benefit. In some embodiments, the reaction rate is 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 9.5%, 10.5%, 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%, 25%, 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% or 100% or some other non-enumerated percentage, and ranges between 1% and 100%. In some embodiments, the reaction rate is between 0% -10%, 10% -20%, 20% -30%, 30% -40%, 40% -50%, 50% -60%, 60% -70%, 70% -80%, 80% -90% or 90% -100%. In some embodiments, the reaction rate is between 0% -1%, 1% -1.5%, 1.5% -2%, 2% -3%, 3% -4%, 4% -5%, 5% -6%, 6% -7%, 7% -8%, 8% -9%, 9% -10%, 10% -15%, 15% -20%, 20-25%, 25% -30%, 35-40%, etc., and so on, to 95% -100%.
The clinical benefit may be an objective response or a persistent clinical response, defined as a sustained response at a median follow-up time of 1 year. In some embodiments, the response, the level of CAR T cells in the blood, or immune-related factors are determined by follow-up about 1 day, about 2 days, about 3 days, about 4 days, about 5 days, about 6 days, or about 7 days after administration of the engineered CAR T cells. In some embodiments, the response, the level of CAR T cells in the blood, or immune-related factors are determined by follow-up about 1 week, about 2 weeks, about 3 weeks, or about 4 weeks after administration of the engineered CAR T cells. In some embodiments, the level of CAR T cells in the reaction, blood, and/or immune-related factor is determined by a follow-up of about 1 month, about 2 months, about 3 months, about 4 months, about 5 months, about 6 months, about 7 months, about 8 months, about 9 months, about 10 months, about 11 months, about 12 months, about 13 months, about 14 months, about 15 months, about 16 months, about 17 months, about 18 months, about 19 months, about 20 months, about 21 months, about 22 months, about 23 months, or about 24 months after administration of the engineered CAR T cells. In some embodiments, the response, the level of CAR T cells in the blood, and/or the immune-related factor is determined by a follow-up about 1 year, about 1.5 years, about 2 years, about 2.5 years, about 3 years, about 4 years, or about 5 years after administration of the engineered CAR T cells.
Monitoring
In some embodiments, administration of the chimeric receptor T cell immunotherapy is performed at a certified medical facility.
In some embodiments, the methods disclosed herein comprise monitoring the patient for CRS signs and symptoms and neurotoxicity and other adverse effects of CAR T cell therapy at least daily at a certified medical facility for 7 days after infusion. In some embodiments, the symptom of neurotoxicity is selected from brain disease, headache, tremor, dizziness, aphasia, delirium, insomnia, and anxiety. In some embodiments, the symptoms of the adverse reaction are selected from the group consisting of fever, hypotension, tachycardia, hypoxia, and cold, including arrhythmias (including atrial fibrillation and ventricular tachycardia), cardiac arrest, heart failure, renal insufficiency, capillary leak syndrome, hypotension, hypoxia, organ toxicity, hemophagocytic lymphoproliferative disorder/macrophage activation syndrome (HLH/MAS), epilepsy, encephalopathy, headache, tremor, dizziness, aphasia, delirium, insomnia anxiety, anaphylaxis, febrile neutropenia, thrombocytopenia, neutropenia, and anemia. In some embodiments, the patient is instructed to remain in the vicinity of the certified medical facility for at least 4 weeks after infusion.
Clinical results
In some embodiments, the clinical outcome is a complete response. In some embodiments, the clinical outcome is a persistent response. In some embodiments, the clinical outcome is a complete response. In some embodiments, the clinical outcome is non-response. In some embodiments, the clinical outcome is a partial response. In some embodiments, the clinical outcome is an objective response. In some embodiments, the clinical outcome is survival. In some embodiments, the clinical outcome is recurrence.
In some embodiments, the Objective Response (OR) is determined according to the revised IWG response criteria for malignant lymphoma (Cheson, 2007) and by the IWG response criteria for malignant lymphoma (Cheson et al Journal of Clinical Oncology 32, 27 (2014, 9 month) 3059-3067). Duration of response (DOR) is defined only for subjects experiencing an objective response and is the time from the first objective response to disease progression (Cheson et al, 2014) or disease-related death (based on prior occurrence). Progression Free Survival (PFS) was assessed by a researcher assessment according to the rugasan response classification criteria.
Prevention and management of severe adverse reactions
In some embodiments, the present disclosure provides methods of preventing the development of or reducing the severity of an adverse reaction based on the level of one or more attributes. In some embodiments, the cell therapy is administered with one or more agents that prevent, delay the onset of, reduce the symptoms of, treat, or otherwise treat an adverse event, such that the adverse event includes cytokine release syndrome and neurotoxicity. In one embodiment, the agent has been described above. In other embodiments, the agent is described below. In some embodiments, the agent is administered before, after, or simultaneously with the administration of the cells by one of the methods and dosages described elsewhere in this specification. In one embodiment, the agent is administered to a subject who may be susceptible to the disease but has not yet been diagnosed with the disease.
In this regard, the disclosed methods may include administering a "prophylactically effective amount" of tolizumab, corticosteroid therapy, and/or antiepileptic drugs for toxicity prevention. In some embodiments, the method comprises administering GM-CSF, CSF1, an inhibitor of GM-CSFR or CSF1R, renzerumab, marvelimumab, a cytokine, and/or an anti-inflammatory agent. The pharmacological and/or physiological effect may be prophylactic, i.e., the effect is wholly or partially prophylactic for a disease or a symptom thereof. A "prophylactically effective amount" may refer to an amount effective (both in dosages and for periods of time necessary) to achieve a desired prophylactic result (e.g., prevent the onset of an adverse effect).
In some embodiments, the method comprises management of adverse reactions in any subject. In some embodiments, the adverse reaction is selected from the group consisting of Cytokine Release Syndrome (CRS), neurotoxicity, hypersensitivity, severe infection, cytopenia, 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 cryogenicity, including cardiac arrhythmias (including atrial fibrillation and ventricular tachycardia), cardiac arrest, heart failure, renal insufficiency, capillary leak syndrome, hypotension, hypoxia, organ toxicity, hemophagocytic lymphoproliferative disorder/macrophage activation syndrome (HLH/MAS), epilepsy, encephalopathy, headache, tremors, dizziness, aphasia, delirium, insomnia anxiety, anaphylaxis, febrile neutropenia, thrombocytopenia, neutropenia, and anemia.
In some embodiments, the patient is identified and selected based on one or more of the biomarkers described herein. In some embodiments, patients have been identified and selected simply by clinical presentation (e.g., presence and level of toxic symptoms).
Cytokine release synthesisSign (CRS)
In some embodiments, the method comprises preventing or reducing the severity of CRS in a chimeric receptor therapy. In some embodiments, the engineered immune cells (e.g., CAR T cells) are inactivated after administration to the patient.
In some embodiments, the method comprises identifying CRS based on clinical manifestations. In some embodiments, the method comprises assessing and treating other causes of fever, hypoxia, and hypotension. Patients with grade 2 CRS (e.g., hypotension, non-response to replacement fluid, or hypoxia requiring replacement of oxygen) should be monitored using continuous cardiac telemetry and pulse oximetry. In some embodiments, for patients with severe CRS, performing echocardiography is considered to assess cardiac function. For severe or life threatening CRS, intensive care support therapy may be considered.
In some embodiments, the method comprises monitoring the patient for CRS signs and symptoms at the certified medical facility at least daily for 7 days after infusion. In some embodiments, the method comprises monitoring the patient for signs or symptoms of CRS for 4 weeks after infusion. In some embodiments, the method comprises suggesting that the patient seek immediate medical attention if signs or symptoms of CRS occur at any time. In some embodiments, the treatment is with supportive care, tobulab or tobulab and a corticosteroid at the time of exhibiting the first sign of CRS.
Neurotoxicity (NT)
In some embodiments, the method comprises monitoring the patient for signs and symptoms of neurotoxicity. In some embodiments, the method includes excluding other etiologies of the neurological symptom. Patients with grade 2 or greater neurotoxicity should be monitored using continuous cardiac telemetry and pulse oximetry. Providing intensive care support therapy for severe or life threatening neurotoxicity. In some embodiments, the symptom of neurotoxicity is selected from brain disease, headache, tremor, dizziness, aphasia, delirium, insomnia, and anxiety.
Management of adverse events
In some embodiments, the cellular therapy is administered before, during, and/or after administration of one or more agents (e.g., steroids) or treatments (e.g., atherectomy) that treat and/or prevent one or more symptoms of (prophylactic) adverse events. A prophylactically effective amount refers to an amount effective to achieve the desired prophylactic result at a dosage and for a period of time necessary. In one embodiment, a prophylactically effective amount is administered to the subject prior to or at an earlier stage of the disease. In one embodiment, the prophylactically effective amount will be less than the therapeutically effective amount. In some embodiments, the patient is selected to manage an adverse event based on the expression of one or more markers described herein in the present specification. In one embodiment, the adverse event treatment or prevention is administered to any patient who will receive, symptomatically receive, or has received cell therapy.
In some embodiments, the method of managing adverse events includes monitoring the patient for neurotoxic signs and symptoms at a certified medical facility at least daily for 7 days after infusion. In some embodiments, the method comprises monitoring the patient for neurotoxicity and/or signs or symptoms of CRS for 4 weeks after infusion.
In some embodiments, the present disclosure provides two methods of managing adverse events in a subject receiving CAR T cell therapy with a steroid and an anti-IL 6/anti-IL 6R antibody. In one embodiment, these methods are depicted in fig. 46. In one embodiment, the present disclosure shows that early steroid intervention in queue 4 is associated with a lower incidence of severe CRS and neurological events than observed in queue 1+2. In one embodiment, the present disclosure shows that early use of the steroid in cohort 4 correlates with about 15% of the median cumulative cortisone equivalent dose in cohort 1+2, indicating that early steroid use may allow for a reduction in overall steroid exposure. Thus, in one embodiment, the present disclosure provides a method of managing adverse events, wherein corticosteroid therapy is initiated for managing all cases of class 1 CRS if there is no improvement after 3 days and for all class 1 neurological events. In one embodiment, tobrazumab is initiated for all cases of class 1 CRS if there is no improvement after 3 days and for all grade 2 neurological events. In one embodiment, the present disclosure provides a method of reducing overall steroid exposure in a patient receiving adverse event management following CAR T cell administration, the method comprising initiating corticosteroid therapy for managing all cases of grade 1 CRS if not improved after 3 days and for all grade 1 neurological events, and/or initiating tobulimib for all cases of grade 1 CRS if not improved after 3 days and for all grade 2 neurological events. In one embodiment, the corticosteroid and tolizumab are administered in a regimen selected from those exemplified in table 12. In one embodiment, the disclosure demonstrates that earlier steroid use is not associated with increased risk of severe infection, reduced CAR T cell expansion, or reduced tumor response.
In one embodiment, the present disclosure supports the safety of levetiracetam prevention in CAR T cell cancer treatment. In one embodiment, the cancer is NHL. In one embodiment, the cancer is R/R LBCL and the patient receives aliskiren. Accordingly, in one embodiment, the present disclosure provides a method of managing an adverse event in a patient treated with immune cells (e.g., CAR T cells), the method comprising administering to the patient a prophylactic dose of an antiepileptic drug. In some embodiments, if a neurological event occurs after discontinuing prophylactic levetiracetam, the patient begins on day 0 of CAR T cell therapy (post-modulation) and also receives levetiracetam at the onset of ≡2 neurotoxicity (e.g., 750mg administered orally or intravenously twice daily). In one embodiment, levetiracetam is tapered and discontinued as clinically indicated if the patient does not experience any grade 2 neurotoxicity. In one embodiment, levetiracetam prevention is combined with any other adverse event management regimen.
In one embodiment, the present disclosure shows that CAR T cell levels in patients receiving poor management regimens for cohort 4 are comparable to those of cohorts 1+2. In one embodiment, the present disclosure shows that the numerical levels of inflammatory cytokines (e.g., ifnγ, IL-2, and GM-CSF) associated with CAR-related inflammatory events are lower in queue 4 than in queue 1+2. Accordingly, the present disclosure provides a method of reducing CAR T cell therapy-related inflammatory events without affecting CAR T cell levels, the method comprising administering to a patient an adverse event management regimen of cohort 4. The present disclosure also provides a method of reducing cytokine production by immune cells following CAR T cell therapy, the method comprising administering to a patient an adverse event management regimen of cohort 4. In one embodiment, this effect is achieved without affecting CAR T cell expansion and response rate. In one embodiment, the patient has R/R LBCL. In one embodiment, the CAR T cell therapy is anti-CD 19 CAR T cell therapy. In one embodiment, the CAR T cell therapy comprises aliskiren.
In one embodiment, the present disclosure demonstrates that early or prophylactic use of tolizumab for adverse event management after alemtuquor reduces grade 3 cytokine release syndrome but increases grade 3 neurological events. Accordingly, the present disclosure provides a method for adverse event management in CAR T cell therapy, as described in fig. 56. In one embodiment, the patient starts receiving levetiracetam on day 0 (750 mg given orally or intravenously twice daily). At the onset of a grade 2 or more neurological event, the levetiracetam dose was increased to 1000mg twice daily. In one embodiment, levetiracetam is gradually decreased and stopped as clinically indicated if the patient does not experience any grade 2 neurological events. Patients also received tolizumab on day 2 (8 mg/kg [ no more than 800mg ] IV administration over 1 hour). Further tolizumab (±corticosteroid) may be recommended at grade 2 CRS onset in patients with co-morbid or older age or in case of grade 3 CRS. Tozumazumab is initiated for patients experiencing a grade-2 neurological event, and corticosteroids are added for patients with co-morbid or aged years, or if there is any occurrence of a grade-3 neurological event, despite symptomatic deterioration with Tozumazumab.
In one embodiment, the present disclosure shows that prophylactic steroid use appears to reduce the rate of severe CRS and NE to a similar extent as early steroid use in infusion following administration of aliskiren. Thus, the present disclosure provides methods for managing adverse events in CAR T cell therapy, wherein the patient receives 10mg of PO-administered dexamethasone on day 0 (prior to infusion of aliskiren), day 1, and day 2. Steroid administration was also started from grade 1 NE and grade 1 CRS when no improvement was observed after 3 days supportive care. Touzumab was also administered for management of grade 1 CRS if no improvement was observed after 24 hours of supportive care.
In one embodiment, the present disclosure demonstrates that adverse event management with CAR T cell therapies that neutralize and/or deplete antibodies to GM-CSF prevents or reduces treatment-associated CRS and/or NE in the treated patient. In one embodiment, the antibody is lorentzumab.
In some embodiments, adverse events are managed by administering one or more agents that are antagonists or inhibitors of IL-6 or IL-6 receptor (IL-6R). In some embodiments, the agent is an antibody that neutralizes IL-6 activity, such as an antibody or antigen binding fragment that binds IL-6 or IL-6R. For example, in some embodiments, the agent is or includes tolizumab (atizumab) or Sha Lim mab, an anti-IL-6R antibody. 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 that targets IL-6 is an anti-TL-6 antibody, such as, for example, cetuximab, ai Ximo mab, ALD518/BMS-945429, western Lu Kashan antibody (CNTO 136), CPSI-2634, ARGX 109, FE301, FM101, or olobulab (CDP 6038), and combinations thereof. In some embodiments, the agent can neutralize IL-6 activity by inhibiting ligand-receptor interactions. In some embodiments, the IL-6/IL-6R antagonist or inhibitor is an IL-6 mutein, such as the IL-6 mutein described in U.S. Pat. No. 5591827. In some embodiments, the agent that is an IL-6/IL-6R antagonist or inhibitor is a small molecule, protein, or peptide, or nucleic acid.
In some embodiments, other agents useful for managing adverse effects and symptoms thereof include antagonists or inhibitors of cytokine receptors or cytokines. In some embodiments, the cytokine or receptor is IL-10, TL-6 receptor, IFNy, IFNGR, IL-2, IL-2R/CD25, MCP-1, CCR2, CCR4, MIP13, CCR5, TNFα, TNFR1 such as TL-6 receptor (IL-6R), IL-2 receptor (IL-2R/CD 25), MCP-1 (CCL 2) receptor (CCR 2 or CCR 4), TGF- β receptor (TGF- β I, II or III), IFN- γ receptor (IFNGR), MIP1P receptor (e.g., CCR 5), TNFα receptor (e.g., TNFR 1), IL-1 receptor (IL-1 Ra/IL-1 RP) or IL-10 receptor (IL-10R), IL-1 and IL-1Rα/IL-1 β. In some embodiments, the agent comprises cetuximab, sha Lim mab, olobulizumab (CDP 6038), ai Ximo mab, ALD518/BMS-945429, west Lu Kashan antibody (CNTO 136), CPSI-2634, ARGX 109, FE301, or FM101. In some embodiments, the agent is an antagonist or inhibitor of a cytokine, such as transforming growth factor beta (TGF-beta), interleukin 6 (TL-6), interleukin 10 (IL-10), IL-2, MIP13 (CCL 4), TNF alpha, IL-1, interferon gamma (IFN-gamma), or monocyte chemotactic protein-I (MCP-1). In some embodiments, the agent is an agent that targets a cytokine receptor (e.g., inhibits or is an antagonist of a cytokine receptor), such as a TL-6 receptor (IL-6R), an IL-2 receptor (IL-2R/CD 25), a MCP-1 (CCL 2) receptor (CCR 2 or CCR 4), a TGF-beta receptor (TGF-beta I, II or III), an IFN-gamma receptor (IFNGR), a MIP1P receptor (e.g., CCR 5), a TNFα receptor (e.g., TNFR 1), an IL-1 receptor (IL 1-Ra/IL-1 RP), or an IL-10 receptor (IL-10R), and combinations thereof. In some embodiments, the agent is administered before, after, or simultaneously with the administration of the cells by one of the methods and dosages described elsewhere in this specification.
In some embodiments, the agent is administered at a dose of about 1mg/kg to 10mg/kg, 2mg/kg to 8mg/kg, 2mg/kg to 6mg/kg, 2mg/kg to 4mg/kg, or 6mg/kg to 8mg/kg (all inclusive), or at least about 2mg/kg, 4mg/kg, 6mg/kg, or 8 mg/kg. In some embodiments, the administration is at a dose of about 1mg/kg to 12mg/kg (such as at or about 10 mg/kg). In some embodiments, the agent is administered by intravenous infusion. In one embodiment, the agent is tolizumab. In some embodiments, the agent (e.g., specific tolizumab) is administered prior to, after, or simultaneously with administration of the cells by one of the methods and dosages described elsewhere in this specification.
In some embodiments, the method comprises identifying CRS based on clinical manifestations. In some embodiments, the method comprises assessing and treating other causes of fever, hypoxia, and hypotension. If CRS is observed or suspected, CRS may be managed according to the recommendation in scheme a, which may also be used in conjunction with other treatments of the present disclosure (including neutralization or reduction of CSF/CSFR1 axis). Patients with grade 2 CRS (e.g., hypotension, non-response to replacement fluid, or hypoxia requiring replacement of oxygen) should be monitored using continuous cardiac telemetry and pulse oximetry. In some embodiments, for patients with severe CRS, performing echocardiography is considered to assess cardiac function. For severe or life threatening CRS, intensive care support therapy may be considered. In some embodiments, in the methods disclosed herein, a biomimetic or equivalent of tobrazumab can be used in place of tobrazumab. In other embodiments, another anti-IL 6R may be used instead of tolizumab.
In some embodiments, adverse events are managed according to the following protocol (protocol a):
(a) Lee DW et al, (2014) Current concepts in the diagnosis and management of cytokine release syndrome. Blood.2014, 7 months, 10 days; 124 (2):188-195.
(b) For management of neurotoxicity, refer to table 2.
(c) For detailed information, please refer toPrescription information (tobulab), https:// www.gene.com/download/pdf/actemra_prescription. Pdf (last visit time: 10 month 18 of 2017). The time of the first approval in the united states was indicated as 2010.
Neurotoxicity (neurotoxicity)
In some embodiments, the method comprises monitoring the patient for signs and symptoms of neurotoxicity. In some embodiments, the method includes excluding other etiologies of the neurological symptom. Patients with grade 2 or greater neurotoxicity should be monitored using continuous cardiac telemetry and pulse oximetry. Providing intensive care support therapy for severe or life threatening neurotoxicity. For any grade-2 neurotoxicity, non-sedating antiepileptic drugs (e.g., levetiracetam) are contemplated for epileptic prevention. The following treatments may be used in combination with other treatments of the present disclosure, including neutralization or reduction of the CSF/CSFR1 axis.
In some embodiments, adverse events are managed according to the following protocol (protocol B):
additional security management policies with corticosteroids
Administration of the corticosteroid and/or tolizumab at grade 1 may be considered prophylactic. Support care may be provided in all scenarios at all CRS and NE severity levels.
In one embodiment of a regimen for managing CRS-related adverse events, tolizumab and/or corticosteroid are administered as follows: class 1 CRS: no tobulimib; no corticosteroid; 2-stage CRS: tobrazumab (only in cases of co-morbidities or older ages); and/or corticosteroids (only in cases of co-morbidities or older ages); 3-stage CRS: tobulimib; and/or a corticosteroid; 4-stage CRS: tobulimib; and/or corticosteroids. In another embodiment of a regimen for managing CRS-related adverse events, tolizumab and/or corticosteroid are administered as follows: class 1 CRS: tobrazumab (if no improvement after 3 days); and/or corticosteroid (if no improvement after 3 days); 2-stage CRS: tobulimib; and/or a corticosteroid; 3-stage CRS: tobulimib; and/or a corticosteroid; 4-stage CRS: tobulimib; and/or corticosteroids, high doses.
In one embodiment of the regimen for managing adverse events associated with NE, tolizumab and/or corticosteroid is administered as follows: stage 1 NE: no tobulimib; no corticosteroid; stage 2 NE: no tobulimib; no corticosteroid; stage 3 NE: tobulimib; and/or corticosteroids (standard doses only used when tolizumab does not improve); stage 4 NE: tobulimib; and/or corticosteroids.
In another embodiment of the regimen for managing adverse events associated with NE, tolizumab and/or corticosteroid is administered as follows: stage 1 NE: no tobulimib; and/or a corticosteroid; stage 2 NE: tobulimib; and/or a corticosteroid; stage 3 NE: tobulimib; and/or corticosteroids, high doses; stage 4 NE: tobulimib; and/or corticosteroids, high doses.
In one embodiment, corticosteroid treatment is initiated at CRS grade No. 2 and toboggan treatment is initiated at CRS grade No. 2. In one embodiment, corticosteroid treatment is initiated when CRS grade is greater than or equal to 1 and toboggan treatment is initiated when CRS grade is greater than or equal to 1. In one embodiment, corticosteroid treatment is initiated at NE levels > 3 and toboggan treatment is initiated at CRS levels > 3. In one embodiment, corticosteroid treatment is initiated at CRS grade No. 1 and toboggan treatment is initiated at CRS grade No. 2. In some embodiments, prophylactic use of tolizumab administered on day 2 may reduce the rate of grade 3 CRS.
In one embodiment, adverse events can be managed by a method comprising scheme C:
a gradually reducing the treatment as symptoms improve at the discretion of the researcher; b no more than 800mg; AE, adverse events; CRS, cytokine release syndrome; IV, intravenous; N/A, inapplicable; NE, neural event
Any corticosteroid may be appropriate for this use. In one embodiment, the corticosteroid is dexamethasone. In some embodiments, the corticosteroid is methylprednisolone. In some embodiments, the two substances are administered in combination. In some embodiments, the glucocorticoids include synthetic and non-synthetic glucocorticoids. Exemplary glucocorticoids include, but are not limited to: beclomethasone, alcrogestone, 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), clobetasol, chlorocortolone (e.g., chlorocortolone pivalate), methylprednisole, corticosterone, cortisone, and hydrocortisone (e.g., hydrocortisone acetate), cocoa valazol, deflazacort, dexamethasone (e.g., 21-dexamethasone phosphate, dexamethasone acetate, dexamethasone sodium phosphate), diflorasone (e.g., diflorasone diacetate), difluprednate, glycyrrhetinic acid, fluzacort, fluclonide, fludrocortisone (e.g., fludrocortisone acetate), fluorometsone (e.g., flumidsone pivalate), flunisolide, fluocinolone (e.g., fluocinolone acetate), fluocinolone acetonide, flucortisone, fluorometholone (e.g., fluometlone acetate), haloperidol (e.g., fluopelone acetate), fluprednisodine, fluprednisolone, fludropinlide, fluticasone (e.g., fluticasone propionate), formosastat, halcinonide, halobetasol, halometasone, haloprednisone, hydrocortisone (e.g., hydrocortisone 21-butyrate, hydrocortisone propyl acetate, hydrocortisone acetate, propyl Ding Qinghua hydrocortisone, hydrocortisone butyrate, hydrocortisone cyclopentanepropionate, hydrocortisone hemisuccinate, hydrocortisone propiolate, hydrocortisone sodium phosphate, hydrocortisone sodium succinate, hydrocortisone valerate), loteprednol etabonate, maprenone, mevalonate, methylprednisone, methylprednisolone (methylprednisolone acetate, methylprednisolone hemisuccinate, methylprednisolone sodium succinate), mometasone (e.g., mometasone furoate), perasone (e.g., praethasone acetate), prednisoester, prednisolone (e.g., prednisolone 25-diethylaminoacetate, prednisolone sodium phosphate, prednisolone 21-hemisuccinate, prednisolone acetate); prednisolone farnesoate, prednisolone hemisuccinate, prednisolone-21 (. Beta. -D-glucuronide), prednisolone metasulphobenzoate, stavudine, prednisolone butoxide, prednisolone tetrahydrophthalate), prednisone, prednisolone valerate, prednisodine, rimexolone, tike, triamcinolone (e.g., triamcinolone acetonide, hexatriamcinolone acetonide, triamcinolone acetonide 21 palmitate, triamcinolone diacetate). These glucocorticoids and their salts are discussed in detail in, for example, the following documents: remington's Pharmaceutical Sciences, A.osol, mack Pub.Co., easton, pa. (16 th edition 1980) and Remington: the Science and Practice of Pharmacy, 22 nd edition Lippincott Williams & Wilkins, philadelphia, pa. (2013) and any other versions, which are hereby incorporated by reference. In some embodiments, the glucocorticoid is selected from the group consisting of cortisone, dexamethasone, hydrocortisone, methylprednisolone, prednisolone, 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.
The one or more corticosteroids may be administered at any dosage and frequency of administration that may be adapted to the severity/level of adverse events (e.g., CRS and NE). Tables 1 and 2 provide examples of dosing regimens for managing CRS and NEs, respectively. In another embodiment, corticosteroid administration comprises oral or IV administration of 10mg dexamethasone 1 to 4 times per day. Another embodiment (sometimes referred to as a "high dose" corticosteroid) includes IV administration of 1g of methylprednisone per day alone or in combination with dexamethasone. In some embodiments, the one or more corticosteroids are administered at a dose of 1 to 2mg/kg per day.
The corticosteroid may be administered in any amount effective to ameliorate one or more symptoms associated with an adverse event, such as CRS or neurotoxicity. A corticosteroid (e.g., glucocorticoid) may be administered to a 70kg adult subject, e.g., in an amount of between or about 0.1 and 100mg, 0.1 to 80mg, 0.1 to 60mg, 0.1 to 40mg, 0.1 to 30mg, 0.1 to 20mg, 0.1 to 15mg, 0.1 to 10mg, 0.1 to 5mg, 0.2 to 40mg, 0.2 to 30mg, 0.2 to 20mg, 0.2 to 10mg, 0.2 to 5mg, 0.4 to 40mg, 0.4 to 30mg, 0.4 to 20mg, 0.4 to 15mg, 0.4 to 10mg, 0.4 to 5mg, 0.4 to 4mg, 1 to 20mg, 1 to 15mg, or 1 to 10mg per dose. Typically, a corticosteroid (such as a glucocorticoid) is administered to a normal adult subject in an amount of between or about 0.4 and 20mg per dose, for example at or about 0.4mg, 0.5mg, 0.6mg, 0.7mg, 0.75mg, 0.8mg, 0.9mg, 1mg, 2mg, 3mg, 4mg, 5mg, 6mg, 7mg, 8mg, 9mg, 10mg, 11mg, 12mg, 13mg, 14mg, 15mg, 16mg, 17mg, 18mg, 19mg, or 20 mg.
In some embodiments of the present invention, in some embodiments, for example, at or about 0.001mg/kg (subject), 0.002mg/kg, 0.003mg/kg, 0.004mg/kg, 0.005mg/kg, 0.006mg/kg, 0.007mg/kg, 0.008mg/kg, 0.009mg/kg, 0.01mg/kg, 0.015mg/kg, 0.02mg/kg, 0.025mg/kg, 0.03mg/kg, 0.035mg/kg, 0.04mg/kg, 0.045mg/kg, 0.05mg/kg, 0.055mg/kg, 0.06mg/kg, 0.065mg/kg, 0.07mg/kg, 0.075mg/kg, 0.08mg/kg, 0.085mg/kg, 0.09mg/kg A dosage of 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, 1.1mg/kg, 1.15mg/kg, 1.20mg/kg, 1.25mg/kg, 1.3mg/kg, 1.35mg/kg or 1.4mg/kg, the corticosteroid is administered to a normal adult subject, which typically weighs about 70kg to 75 kg.
Generally, the dose of corticosteroid administered depends on the particular corticosteroid, as there is a potency difference between different corticosteroids. It will be generally understood that the potency of a drug varies, and thus the dosage will vary to achieve an equivalent effect. The equivalence of various glucocorticoids and the efficacy of the route of administration is well known. Information relating to the administration of equivalent steroids (in a non-time therapeutic manner) can be found in the national formulary of England (British National Formulary, BNF) 37, 3 nd 1999.
In some embodiments, adverse events are managed by the following scheme: patients received levetiracetam (750 mg orally or intravenously, twice daily) beginning on day 0 of administration of T cell therapy; increasing the levetiracetam dose to 1000mg twice a day at the onset of a grade 2 or more neurological event; in one embodiment, if the patient does not experience any grade-2 neurological event, levetiracetam is gradually decreased and stopped as indicated clinically; patients also received tolizumab on day 2 (IV administration of 8mg/kg [ no more than 800mg ] over 1 hour); further tobrazumab (±corticosteroid) may be recommended at grade 2 CRS onset in patients with co-morbid or elderly patients or in the case of grade 3 CRS; tozumazumab is initiated for patients experiencing a grade-2 neurological event, and corticosteroids are added for patients with co-morbid or aged years, or if there is any occurrence of a grade-3 neurological event, despite symptomatic deterioration with Tozumazumab. In some embodiments, levetiracetam is administered for prophylaxis at the onset of ≡2 neurotoxicity if a neurological event occurs after cessation of prophylactic levetiracetam, and/or gradually decreases and stops levetiracetam if the patient does not experience any ≡2 neurotoxicity.
In some embodiments, adverse events are managed by the following scheme: patients received 10mg PO of dexamethasone on day 0 (prior to T cell therapy infusion), day 1 and day 2; steroids were also administered starting from grade 1 NE, and no improvement was observed after 3 days of supportive treatment for grade 1 CRS; touzumab was also administered for management of grade 1 CRS if no improvement was observed after 24 hours of supportive care.
Secondary malignant tumor
In some embodiments, a patient receiving immune cells (e.g., CAR T cells) (e.g., CD 19-directed) or other genetically modified autologous T cell immunotherapy treatment may suffer from a secondary malignancy. In certain embodiments, a patient receiving CAR T cells (e.g., CD 19-directed) or other genetically modified allogeneic T cell immunotherapy treatment may suffer from a secondary malignancy. In some embodiments, the method comprises lifelong monitoring for secondary malignancy.
Examples
Example 1
This example provides the results of the analysis of clinical trial-2. Clinical trial-2 the study design of the trial is summarized in figure 1. Provided herein are reports of safety and efficacy outcomes and pharmacokinetic profiles for patients in clinical trial-2 with and without disease progression (POD 24) within 24 months of diagnosis. Disease progression (POD 24) within 24 months after the initial diagnosis is an indicator of poor outcome in Mantle Cell Lymphoma (MCL) patients. (Visco C et al Br J Haemato.2019; 185:940-944). In a retrospective analysis of patients with MCL, median Overall Survival (OS) from time of progression was 12 months for those patients with POD24 compared to those patients without POD 24. KTE-X19, an autologous anti-CD 19 Chimeric Antigen Receptor (CAR) T cell therapy, is approved in the united states and the european union for the treatment of recurrent/refractory (R/R) MCL. ( (brexucabtagene autoleucel) prescription information Kite Pharma, inc;2021;Summary of product characteristics (autologous anti-CD 19 transduced cd3+ cells) Kite Pharma EU b.v.; 2021). Critical phase 2 clinical trial-2 studies KTE-X19 was evaluated in patients with MCL who were R/R for 1 to 5 past therapies, including Bruton's Tyrosine Kinase Inhibitor (BTKi). (Wang M et al N Engl J Med.2020; 382:1331-1342). After 17.5 months of median follow-up (n=60) in clinical trial-2, the Objective Response Rate (ORR) was 92%, completeThe reaction (CR) rate was 67%. (Wang M et al blood.2020;136 (journal 1): 20-22). At the time of data cutoff, 48% of all patients and 70% of CR patients continued to respond. Cytokine Release Syndrome (CRS) and Neurological Events (NE) were mostly reversible (n=68 treated patients). 15% of patients show grade 3 CRS and 31% have grade 3 NE. Two patients showed grade 5 Adverse Events (AEs), only one of which was KTE-X19-related. No new security signal was observed in the case of long-term follow-up. (Wang M et al N Engl J Med.2020;382:1331-1342; wang M et al blood.2020;136 (journal 1): 20-22).
A eligible patient was 18 years old or older, had pathologically confirmed Mantle Cell Lymphoma (MCL), recorded cyclin D1 overexpression or the presence of t (11; 14), and was relapsed/refractory to 1 to 5 past MCL regimens. Previous therapies included chemotherapy with anthracyclines or bendamustine, anti-CD 20 monoclonal antibodies, and ibrutinib or acartinib. All patients received prior BTKi. Although patients must have past BTKi therapies, they do not need to be the last line of therapy before study entry, and patients do not need to be refractory to BTKi therapies. Patients eligible have an absolute lymphocyte count of > 100/. Mu.L. Patients experiencing autologous SCT or having past CD19 targeted therapies or allogeneic SCT within 6 weeks of CD19 CAR-T cell infusion were excluded. All patients underwent leukapheresis to obtain cells for CD19 CAR-T cell therapy manufacturing. The patient received an optional bridging therapy comprising dexamethasone (20 mg to 40mg or equivalent), ibrutinib (560 mg administered via the oral cavity (PO) per day), or acartinib (100 mg administered twice daily PO). The method of manufacture of KTE-X19 was modified with respect to the method of manufacture of alemtujopsis to enrich CD4 positively + /CD8 + Cells remove circulating lymphoma cells. This product is referred to herein as a "CAR T cell". Single intravenous infusion 2 x 10 on day 0 6 CD19 CAR-T cells of individual CAR T cells/kg, fludarabine was administered on day-5, day-4 and day-3 (30 mg/m per day 2 ) And cyclophosphamide (500 mg/m per day) 2 ) Is a conditioning chemotherapy.
In this example, the safety results, pharmacological profile and product attributes of all 68 patients treated with KTE-X19 are reported. Efficacy results for 60 treated patients with a 1 year or more follow-up (median 17.5 months) are reported. The data is presented at a data expiration date of 2019, 12, 31.
The baseline characteristics of the patients are summarized in table 1. High risk disease features are common in patients with and without POD 24. Patients with POD24 have higher tumor burden and Lactate Dehydrogenase (LDH) levels, and more patients have a maternal cell pattern MCL, suggesting that these patients may not be as suitable as those without POD 24. Patients with POD24 are more likely to have high risk disease characteristics (high tumor burden, high LDH levels, and blast-like MCL) than patients without POD 24.
TABLE 1 baseline characteristics per POD24 status 。
The overall response rates (ORR; CR and partial response) were assessed by the independent radiological examination committee according to the Rugos classification. Cheson BD et al Journal of clinical oncology 2014;32:3059-68.ORR is similar in patients with and without POD24, with slightly higher CR rates in patients without POD24 (fig. 2). KTE-X19 provided high CR rates in patients with and without POD24 following a median 17.5 month follow-up. Minimal residual disease (MRD; 10) -5 Sensitivity) was assessed by next generation sequencing as previously reported. Wang M et al New Engl J Med.2020;382:1331-1342. MRD was assessed in patients on week 4 with available samples. Similar MRD negative rates were also observed in patients with (75%; n=9/12) and without (79%; n=15/19) POD 24.
Secondary endpoints associated with efficacy include duration of response (DOR), progression Free Survival (PFS), and Overall Survival (OS). (FIG. 3). In patients with POD24, median progression-free survival (PFS) was 11.3 months (95% ci,6.0 ne). The median PFS appears to be shorter in patients with POD24 compared to patients without POD 24. The reaction Duration (DOR) and the median of OS were not reached in either group (fig. 3).
The safety profile of patients with and without POD24 is generally similar. The incidence of grade 3 adverse events was generally similar in patients with and without POD24 (Table 2). The incidence of thrombocytopenia and neutropenia is higher in patients with POD24 than in patients without POD 24. Patients without POD24 present a higher incidence of infection than patients with POD 24. There were no cases of grade 5 cytokine release syndrome, KTE-X19 associated secondary malignancy, or replication competent retrovirus in either group.
TABLE 2 summary of adverse events in patients with and without POD24。
a CRS according to Lee DW et al blood.2014; 124:188-195. CRS symptoms and all other AEs were ranked according to the american national cancer institute adverse events common terminology standard version 4.03. AE, adverse events; CRS, cytokine release syndrome; with POD24, disease progression <24 months after initial diagnosis; without POD24, disease progression was > 24 months after initial diagnosis.
Product attributes and CAR T cell levels in blood were analyzed using the methods previously described. Locke FL et al, mol Ther.2017;25:285-295. KTE-X19 product characteristics were similar in patients with and without POD24 (table 3). Patients with POD24 appear to have lower CAR T cell expansion than patients without POD 24. In patients with POD24, median peak CAR T cell levels and median area under the curve (AUC) were 53.4 cells/. Mu.L (range 0.2-2566) and 583.4 cells/. Mu.L X days (range 1.8-27,743.6; FIG. 4). Patients without POD24 had median peak CAR T cell levels and median AUC of 112.4 cells/μl (range, 0.2-2589) and 1588.3 cells/μl×days (range, 3.8-27,238.7).
TABLE 3 KTE-X19 product characteristics according to POD24 status。
Among patients with evaluable efficacy with available data, B cells were detectable in 8/11 (73%) of patients with POD24 and 7/15 (47%) of patients without POD24 at 12 months (fig. 5). Early intervention with CAR T cell therapy for CD19 may benefit MCL patients with known high risk factors (such as POD 24). (Visco C et al Br J Haemato.2019; 185:940-944).
Example 2
This example provides the results of a clinical trial-5 (a phase 2 multicenter single-group study) analysis of aliskiren in patients with recurrent/refractory (R/R) indolent non-hodgkin's lymphoma (R/R inHI; NCT 03105336). Clinical trial-5 the study design of the trial is summarized in figure 6. Alkylrensaine (axi-cel) is an autologous anti-CD 19 Chimeric Antigen Receptor (CAR) T-cell therapy that has been approved in the United States (US) for treatment of adults with relapsed/refractory (R/R) FL after > 2 lines of systemic therapy, and in the United states and European Union for treatment of adults with R/R large B-cell lymphomas (LBCL) after > 2 lines of systemic therapy. (Kite Pharma, inc.; 2021;(Alkylenzepine) [ characterization of product ] ].Amsterdam,The Netherlands:Kite Pharma EU B.V.；2018)。
Progression within 24 months from the first receipt of anti-CD 20 containing chemotherapy (POD 24) is a risk factor for adverse survival in patients with indolent non-hodgkin's lymphoma (iNHL). (Casulo C and Barr P.blood.2019;133 (14): 1540-154; casulo C et al J Clin Oncol.2015;33 (23): 2516-2522). About 20% of patients with Follicular Lymphoma (FL) have POD24. In the observation analysis from National LymphoCare Study, patients with FL with early progression had a lower 5-year Overall Survival (OS) (50%) than those without early progression (90%). This is a report on safety and efficacy results with longer follow-up and pharmacokinetic/pharmacodynamic profiles in patients in clinical trial-5 with and without POD24.
Adults with Follicular Lymphoma (FL) (grade 1-3 a) or marginal zone lymphoma (MZL; lymph nodes or extranodal) suffer from R/R disease after > 2 line number therapies (including anti-CD 20 mAb plus alkylating agent) and ECOG 0-1. Patients underwent leukapheresis followed by conditioning therapy (intravenous fludarabine at day-5, day-4 and day-3 (30 mg/m) 2 Body surface area) and cyclophosphamide (500 mg/m 2 Body surface area)), and at day 0 at 2×10 6 Single infusion of each CAR T cell/kg of alopecie. The primary endpoint was the Objective Response Rate (ORR) of the center review (complete response (CR) +partial response (PR)) (according to the Rugoson classification; cheson BD et al J Clin Oncol.2014;32 (27): 3059-3068.Doi: 10.1200/JCO.2013.54.8800). Secondary endpoints include Complete Response (CR) rate (according to the rugo classification; cheson et al J Clin oncol.2014), duration of response (DOR) defined only for subjects undergoing objective response, and are the time from the first objective response to disease progression (Cheson et al 2014) or death associated with disease (whichever occurs earlier), progression Free Survival (PFS) (PFS defined as the time from day of infusion of alemtujose to day of disease progression (Cheson et al 2014) or death due to any cause), overall Survival (OS) (OS defined as the time from infusion of alemtujose to day of death), incidence of Adverse Events (AE), and levels of CAR T cells and cytokines in serum. Major efficacy analysis was performed when 80 or more FL patients receiving treatment were followed for 12 or more months. Median follow-up in the preliminary analysis was 17.5 months (Jacobson et al ASH 2020.# 700), 92% of patientsReaction (76% complete reaction [ CR) ]Rate). Patients with FL have a lower rate of ≡3 neural events (NE; 15%) than patients with MZL (41%). In the preliminary analysis, the Overall Response Rate (ORR) after the 17.5 month medium follow-up was similarly high in patients with and without POD24 (93% versus 92%).
Updated efficacy analysis was performed when 80 or more FL patients receiving treatment were followed for 18 or more months. By the date of data expiration, patients with evaluable efficacy included 80 FL patients receiving treatment, who were followed by No. 18 months after infusion of aliskiren, and MZL patients receiving treatment, who were followed by No. 4 weeks (14 days 9 months in 2020) after infusion of aliskiren. The analyzed POD24 included data of progression after treatment with anti-CD 20 mAb + alkylating agent in either FL or MZL patients treated with aliskiren (n=129).
The baseline characteristics were generally similar in patients with and without POD24 (table 4). In evaluable patients with FL, median tumor burden calculated by product diameter Sum (SPD) was similar in value in patients with and without POD24 (2303 mm 2 Compared with 2839mm 2 ). In evaluable patients without MZL, the median SPD appears to be higher in those with POD24 than in those without POD24 (2028 mm 2 Compared with 954mm 2 )。
TABLE 4 Baseline disease signatures。
a disease burden, as defined by the genf standard: involving more than or equal to 3 node sites (each diameter more than or equal to 3 cm); any nodule or extra-nodule tumor mass with a diameter of greater than or equal to 7 cm; symptoms B; splenomegaly; pleural effusion or peritoneal ascites; cytopenia; or leukemia. b Before modification of the regimen requiring more than or equal to 2 past line counts of therapy, the clinical trial-5 was followed3 FL patients were grouped with 1 past line number of therapies. c Patients with iNHL, who progressed within 6 months of completion of the last past treatment. FL, follicular lymphoma; FLIPI, international prognosis index for follicular lymphoma; GELF, follicular lymphoma group (group' Etude des Lymphomes Folliculaires); MZL, marginal zone lymphoma; PI3Ki, phosphoinositide 3-kinase inhibitors; POD24, disease progression from first receiving anti-CD 20 containing chemotherapy<24 months; SCT, stem cell transplantation.
Secondary endpoints associated with efficacy include duration of response (DOR), progression Free Survival (PFS), and Overall Survival (OS). The overall response rates (ORR; CR and partial response) were assessed by the independent radiological examination committee according to the Rugos classification. Cheson BD et al Journal of clinical oncology 2014;32:3059-68. ORR was similar in patients with and without efficacy assessments of POD24 (fig. 7; table 5). Patients with and without POD24 did not reach the estimated median duration of response (DOR) following median follow-up for 17.1 months and 17.5 months, respectively (fig. 8A, 8B, 8C; table 5). At the time of data cutoff, 52% of patients with efficacy assessments of POD24 and 70% of patients without POD24 responded. The 18 month DOR rates for patients with and without POD24 were 60% and 78%, respectively. Median progression-free survival (PFS) and median OS were not achieved in patients with and without POD24 (FIG. 8B, FIG. 8C; table 5). The 18 month PFS rates for patients with and without POD24 were 55% and 84%, respectively. The OS rate for 18 months was 85% and 94%, respectively.
TABLE 5 efficacy results in patients with FL and MZL by POD24 status。
CR, complete reaction; DOR, duration of reaction; FL, follicular lymphoma; mo, month; MZL, marginal zone lymphoma; ND, incomplete/undefined; NE, not estimated; NR, not reached;
OS, overall survival; PFS, progression free survival; POD24, disease progression <24 months from first receiving anti-CD 20 containing chemotherapy; PR, partial reaction.
The incidence of grade 3 adverse events was generally similar in patients with and without POD24 (Table 6). Grade 5 events occurred in 3 patients with POD24, including 1 event in the case of Cytokine Release Syndrome (CRS); no grade 5 event occurred in patients without POD 24. Class 4 CRS occurred in 1 patient with POD 24. Grade 4 neurological events occurred in 2 patients with POD 24. In patients without POD24, no grade 4 CRS or neurological event occurred.
The CAR T cell levels in blood, cytokine levels in serum and product attributes and their association with clinical outcome were analyzed by using the methods described previously. Locke FL et al, mol Ther.2017;25:285-295.
In patients with FL where efficacy was assessed, median peak CAR T cell levels were similar in patients with and without POD24 (36.9 cells/. Mu.L and 34.5 cells/. Mu.L, respectively; FIG. 9A). The median AUC in patients with and without POD24 was also similar (422.5 cells/. Mu.L. Times.day and 407.6 cells/. Mu.L. Times.day; FIG. 9B), respectively. Pre-treatment levels of CCL17 (TARC) and CCL22 (MDC) were higher in patients with POD24 than in patients without POD 24. The peak levels of biomarkers associated with the toxicity of aliskiren appear to be generally similar in all treated patients with and without POD24 (table 7). Pharmacokinetic/pharmacodynamic findings between groups were similar in patients with MZL.
TABLE 7 peak cytokine levels in patients with FL by POD24 status。
P-values were calculated using Wilcoxon rank sum test. a Lower limit of quantification in the assay used. b The upper limit of quantification in the assay used. c For patients with at peakData for 2 patients of FL are not available. CCL, chemokine (C-C motif) ligand; CRP, C-reactive protein; CXCL, C-X-C motif chemokine ligands; FL, follicular lymphoma; GM-CSF, granulocyte-macrophage colony stimulating factor; IFN, interferon; IL, interleukin; MCP-1, monocyte chemotactic protein 1; POD24, disease progression from first receiving anti-CD 20 containing chemotherapy<24 months; RA, receptor agonist; SAA, serum amyloid A; TNF, tumor necrosis factor;
out of 14 patients (13 FL;1 MZL) in the broad clinical trial-5 population with data available at recurrence after alemtuquor, 100% had detectable CD19. Detectable CD19 was confirmed in all evaluable biopsies from patients with and without POD 24. The product attributes of the alemtuquor are generally similar in patients with and without POD24 (table 8).
TABLE 8 characteristics of Alkylrensaine products in patients with FL by POD24 status 。
a Based on the available data: having POD24, n=57; without POD24, n=36 (ccr7+cd45ra+ cells) and n=35 (CD 4/CD8 ratio). Axi-cel, alemtujopsis; FL, follicular lymphoma; IFN, interferon; POD24, disease progression from first receiving anti-CD 20 containing chemotherapy<For 24 months.
The alopecie showed a high rate of persistent response in patients with POD24 iNHL. Although the median PFS was not reached in either group, the estimated PFS rate at 18 months appeared to be lower in patients with POD24 than in patients without POD 24. In patients with FL, higher pre-treatment levels of analytes previously associated with relapse (CC 17 TARC and CCL22 MDC ] (Plaks V et al AACR 2021.#ct 036)) were observed in patients with POD24 compared to patients without POD24, potentially contributing to a difference in PFS rate of 18 months. The safety profile is similarly manageable in patients with and without the POD 24. In patients with FL, the pharmacokinetic and pharmacodynamic profile after treatment appears to be substantially comparable in patients with and without POD 24. Alkylrensai is a promising option for patients with POD24 inHI (populations with particularly high risk of disease).
Example 3
This example characterizes two anti-CD 19 CAR T therapies, KTE-X19 and alemtuzite. The method of manufacture of KTE-X19 was modified with respect to the method of manufacture of alemtujopsis to enrich CD4 positively + /CD8 + Cells remove circulating lymphoma cells. Cells were labeled with fluorescent conjugated antibodies to CD3 (pan T cell marker), CD14, CD19 (B cell marker), CD45 (pan white cell marker) and CD56 (activation and NK markers) and evaluated by flow cytometry. Cell viability was assessed using negative staining with a viability dye (SYTOX near IR). The lower limit of quantification (LLOQ) was determined to be 0.2% and 5% for NK cells and monocytes. Determination of the percentage of NK cells (NK cells are CD 45) + 、CD14 - 、CD3 - And CD56 + The method comprises the steps of carrying out a first treatment on the surface of the T cell is CD45 + 、CD14 - And CD3 - ). The median percentages of NK cells from 23 batches of Alkylene and 97 batches of KTE-X19 were 1.9% (range 0.8% -3.2%) and 0.1% (range 0.0% -2.8%), respectively. CD3 from the same batch of Alkylrensaine and KTE-X19 - The median percentages of cellular impurities are 2.4% (range 0.9% -4.6%) and 0.5% (range 0.3% -3.9%), respectively. The results of KTE-X19 (brexucabtagene autoleucel, TECARTUS) and Alkerese (YESCARTA) in terms of cell viability were not less than 72% and not less than 80%, respectively; the results in terms of anti-CD 19 CAR expression were 24% or more and 15% or more, respectively; results in terms of IFN-gamma production are 190pg/mL or more and 520pg/mL or more, respectively; in CD3 + The results in terms of percentage of cells were not less than 90% and not less than 85%, respectively. Brexucabtagene autoleucel consists mainly of CD3+ T cells (99.3% + -0.8%), which can be further depicted as CD4+ (37.9% + -16.5%) and CD8+ (59.3% + -16.5%) subsets. The presence of circulating lymphoma cells may play a role in the depletion of anti-CD 19 CAR T cells during manufacturing failure and ex vivo.
Example 4
This example relates to the 3-year results of clinical trial-4 (NCT 02625480), a 1/2-stage multicenter study evaluating the safety and efficacy of KTE-X19, an autologous anti-CD 19 Chimeric Antigen Receptor (CAR) T cell therapy, as described above, in pediatric/adolescent patients with relapsed/refractory (R/R) B-cell acute lymphoblastic leukemia (B-ALL; median follow-up time for ALL patients receiving treatment: 36.1 months). Exploring to receive lower doses 1 x 10 in phase 1 6 Two formulations of Chimeric Antigen Receptor (CAR) T cells/kg patient (one with a total volume of 40mL and the other with a volume of 68 mL). The 40mL formulation is intended to maintain cell density and cell viability during the freeze/thaw process. For body weight>100kg of patients, administered 2X 10 8 Or 1X 10 8 Maximum flat dose of individual anti-CD 19 CAR T cells. The primary endpoint was the incidence of Dose Limiting Toxicity (DLT). Of the 31 patients in the group, KTE-X19 was administered to 24 (median age 13.5 years, range 3-20), with a median follow-up of 36.1 months. No DLT was observed. All treated patients had grade 3 or more adverse events, typically hypotension (50%) and anemia (42%). The ratio of the level 3 cytokine release syndrome was 2×10 in all treatments 6 、1×10 6 (68 mL of formulation) and 1X 10 6 33%, 75%, 27% and 22% of CAR T cells/kg group in each (40 mL formulation); 21%, 25%, 27% and 11% of patients experience > 3 grade neurological events, respectively. Total Complete Remission (CR) rate (CR and CR with incomplete hematological recovery) at all treatments, 2 x 10 6 、1×10 6 (68 mL) and 1X 10 6 67%, 75%, 64% and 67% of the (40 mL) CAR T cells/kg group, respectively. The overall MRD negative rate was 100% among the responders. At 1X 10 6 In the (40 mL) group (recommended phase 2 dose), median duration of remission and overall survival were not achieved. Pediatric/adolescent patients with R/R B-ALL achieved high rates of MRD negative remission and controllable safety profiles after a single dose of KTE-X19. Phase 2 at 1X 10 6 Individual CAR T cells/kg (40 mL) doses were taken.
In clinical trial-4In phase 1, a eligible patient is 21 years old or less, weighs 6kg or more and has only R/R B-ALL, defined as refractory to first line therapy, R/R after 2 lines of systemic therapy or R/R after alloSCT if the patient is 100 days or more from alloSCT at time of group entry and stops immunosuppressive drugs for 4 weeks or more before group entry. Pre-treatment with bolamitraz is allowed. Dose Limiting Toxicity (DLT) is defined as follows: grade 4 hematologic toxicity, if not due to underlying disease, persists for more than 30 days (except for lymphopenia); all KTE-X19-associated grade 3 non-hematological toxicity persists >7 days; and all KTE-X19 associated grade 4 non-hematological toxicities were independent of duration, with the exception shown in table 9. Specific bridging chemotherapies are allowed after leukapheresis and at least 7 days or 5 half-lives are completed before the onset of conditioning chemotherapies consisting of 25mg/m of Intravenous (IV) fludarabine on day-4, day-3 and day-2, whichever is shorter 2 Day and single dose of IV cyclophosphamide 900mg/m on day-2 2 Composition is prepared. At day 0 at 2X 10 6 Or 1X 10 6 A single IV infusion of KTE-X19 was administered at a target dose of CAR T cells/kg. Hospitalization is required for a minimum of 7 days after infusion. All patients who completed the 3 rd month visit were subjected to a long follow-up period of survival and disease status lasting up to 15 years every 3 months up to 18 months, every 6 months between 24 th month and 60 th month, and once a year. If the patient withdraws consent for further follow-up, no follow-up or death, it can be removed from the study. Allogeneic stem cell transplantation (alloSCT) is not a requirement of the protocol, but is allowed by researchers as appropriate.
Additional phase 1 inclusion criteria
Morphological diseases with >5% myeloid blast:
1. lansky or Karnoofsky behavior state is more than or equal to 80 percent during screening
2. Patients with philadelphia chromosome positive disease are eligible if they are intolerant to tyrosine kinase inhibitor therapy or they have R/R disease despite treatment with at least two different Tyrosine Kinase Inhibitors (TKIs).
3. In patients previously treated with Bonauzumab, leukemia blasts with CD19 expression > 90% were required
4. Absolute Neutrophil Count (ANC). Gtoreq.500/. Mu.L unless the major investigator believes that cytopenia is caused by underlying leukemia and that leukemia treatment is likely to be reversed
5. Platelet count ≡50,000/. Mu.L unless the first investigator thought that cytopenia was caused by potential leukemia and leukemia treatment was likely to be reversed
6. Absolute lymphocyte count is greater than or equal to 100/. Mu.L
7. Adequate kidney, liver, lung and heart functions are defined as:
a. creatinine clearance (estimated according to Cockcroft Gault or Schwartz) of 60cc/min or more
b. Serum alanine aminotransferase and aspartate aminotransferase are less than or equal to 5 times the Upper Limit of Normal (ULN)
c. Total bilirubin is less than or equal to 1.5 XULN, except for patients with Gilbert's syndrome
d. The left ventricular shortening fraction is more than or equal to 30 percent or the left ventricular ejection fraction is more than or equal to 50 percent, and the ultrasonic cardiogram has no signs of pericardial effusion and no clinical obvious arrhythmia
e. No clinically significant pleural effusion
f. Baseline oxygen saturation in indoor air >92%
8. The serum or urine pregnancy test of women with fertility (defined as first menses) must be negative
Additional phase 1 exclusion criteria
1. Diagnosis of Burkitt leukemia (Burkitt Leukemia)/lymphoma or chronic myelogenous leukemia lymphoid blast crisis according to the world health organization classification
2. History of malignancy other than non-melanoma skin cancer or carcinoma in situ unless no disease is present for 3 or more years
3. History of severe hypersensitivity to aminoglycosides or any agent used in the study
4. History or presence of any Central Nervous System (CNS) disorder, such as seizure disorders, cerebrovascular ischemia/hemorrhage, dementia, cerebellar disorders, any autoimmune disease with CNS involvement, reversible posterior encephalopathy syndrome, or cerebral edema
a. The < 5/mm 3 leukocytes with or without a neurological change (CNS-2) detectable in the cerebrospinal fluid sample, or the > 5/mm 3 leukocytes with or without a neurological change (CNS-3) detectable in the cerebrospinal fluid sample, are also excluded.
5. A history of concomitant genetic syndromes associated with bone marrow failure, such as Fanconi anemia (Fanconi anemia), coleman syndrome (Kostmann syndrome) or shwa schmann-Dai Mengde syndrome (Shwachman-Diamond syndrome)
6. A history of myocardial infarction, cardiovascular angioplasty or stent implantation, unstable angina or other clinically significant heart disease within 12 months of the group;
7. history of symptomatic deep vein thrombosis or pulmonary embolism within 6 months of the group entry
8. Primary immunodeficiency
9. Infection with HIV, hepatitis b or hepatitis c virus is known. If viral load cannot be detected based on quantitative Polymerase Chain Reaction (PCR) and/or nucleic acid testing, a history of hepatitis B or hepatitis C is allowed
10. The presence of fungi, bacteria, viruses or other infections is uncontrolled or requires IV antimicrobial administration. Allowing simple urinary tract infections and uncomplicated bacterial pharyngitis if responsive to active treatment and after consultation with a kit medical monitor
11. Past drug treatment
a. Salvage systemic therapy (including chemotherapy, TKI for ph+all, and boscalid) within 1 week or 5 half-lives (whichever is shorter) prior to group entry
b. Past CD19 directed therapies other than bordetention
c. Standard history of common adverse event terminology (grade 4 neurological events or grade 4 CRS (according to Lee et al 2014) in the case of past CD19 targeted therapies)
d. Albizumab was used within 6 months prior to administration, clofarabine or cladribine was used within 3 months prior to administration, and PEG-asparaginase was used within 3 months prior to administration
e. Donor lymphocyte infusions within 28 days prior to group entry
f. Any drug used for Graft Versus Host Disease (GVHD) within 4 weeks prior to group entry (e.g., calcineurin inhibitor, methotrexate, mycophenolate mofetil, rapamycin (rapamycin), thalidomide) or immunosuppressive antibodies (e.g., anti-CD 20, anti-tumor necrosis factor, anti-interleukin 6 or anti-interleukin 6 receptor) used within 4 weeks prior to group entry.
g. At least 3 half-lives (e.g., ipilimumab, nivolumab, pembrolizumab, atuzumab, OX40 agonist, 4-1BB agonist) must elapse before entry into the group, starting with any previous systemic inhibitory/stimulatory immune checkpoint molecular therapy;
h. corticosteroid treatment (0.7 mg/kg/day or equivalent doses of hydrocortisone or corticosteroid) and other immunosuppressive drugs at pharmacological doses must be avoided 7 days prior to group entry;
12. there are any built-in wires or tubes (e.g., percutaneous nephrostomy tubes, built-in Foley catheters, bile delivery tubes, or spleen/peritoneal/pericardial catheters). Ommaya reservoir and a dedicated central venous access catheter, such as Port-a-Cath or Hickman catheters, are permissible
13. Acute or chronic GVHD requiring systemic treatment within 4 weeks prior to group entry according to Gluckberg criteria or grade II-IV GVHD severity B-D by International marrow transplantation registry index (International Bone Marrow Transplant Registry index)
14. Live vaccine less than or equal to 4 weeks before group entry
15. Pregnant or lactating women with fertility potential due to the potentially dangerous effects of preparative chemotherapy on fetuses or infants
16. Patients of two sexes with fertility potential who are reluctant to practice fertility control from the time of consent to 6 months after completion of KTE-X19
17. At the discretion of the researcher, the subject is unlikely to complete the study visit or procedure (including follow-up visit) required for all protocols or to follow the participation requirements of the study;
18. history of autoimmune disease (e.g., crohn's disease, rheumatoid arthritis, systemic lupus), leading to end organ damage or the need for systemic immunosuppression/systemic disease modulation over the last 2 years.
Study design and treatment were as follows: phase 1 targets were to assess the safety of KTE-X19 and determine the recommended phase 2 dose (RP 2D) of KTE-X19 based on the incidence of Dose Limiting Toxicity (DLT) and overall safety profile. DLT is defined as: grade 4 hematologic toxicity (except lymphopenia) persisting for more than 30 days without being attributed to underlying disease all KTE-X19-associated grade 3 non-hematologic toxicities persist>7 days; and all KTE-X19 associated grade 4 non-hematological toxicities were independent of duration, with the exception shown in table 9. Specific bridging chemotherapies are allowed after leukapheresis and at least 7 days or 5 half-lives are completed before the onset of conditioning chemotherapies consisting of 25mg/m of Intravenous (IV) fludarabine on day-4, day-3 and day-2, whichever is shorter 2 Day/day, single dose IV cyclophosphamide 900mg/m2 at day-2. At day 0 at 2X 10 6 Or 1X 10 6 A single IV infusion of KTE-X19 was administered at a target dose of CAR T cells/kg. Hospitalization is required for a minimum of 7 days after infusion. All patients who completed the 3 rd month visit were subjected to a long follow-up period of survival and disease status lasting up to 15 years every 3 months up to 18 months, every 6 months between 24 th month and 60 th month, and once a year. If the patient withdraws consent for further follow-up, no follow-up or death, it can be removed from the study. Allogeneic stem cell transplantation (alloSCT) is not a requirement of the protocol, but is allowed by researchers as appropriate.
TABLE 9 Standard for dose limiting toxicity
CRS, cytokine release syndrome; DLT, dose limiting toxicity; TLS, tumor lysis syndrome.
At 2X 10 6 DLT was assessed in the first 3 patients treated with an initial dose of CAR T cells/kg. Another patient was enrolled in a group to receive 2 x 10 6 Individual CAR T cells/kg. After 28 days following the infusion, the safety data was evaluated by a safety panel, with subsequent patients receiving 1×10 6 CAR T cells/kg to assess the potential to mitigate CRS and NE risk to improve risk: benefit ratio. (Shah BD et al, J Clin Oncol 37:abstr 7006,2019;Shah BD et al, blood In Press, 2021). To further optimize the risk to benefit ratio, for patients in the second cohort, 1X 10 6 The individual CAR T cells/kg modified the dosing formulation from 68mL to 40mL. Given the expected patient weight for this pediatric study population, 40mL of the formulation was intended to provide a higher cell density than 68mL of the formulation in order to mitigate the potential risk of lower end product volumes. Patients underwent leukocyte apheresis to obtain cells for CAR T cell manufacturing, followed by subsequent opsonizing chemotherapy; fresh leukocyte apheresis material was used for CAR T cell manufacturing. Specific bridging chemotherapies were allowed between leukocyte apheresis and opsonic chemotherapy (table 10).
TABLE 10 bridging chemotherapy
* For patients with ph+ ALL and Ph-like ALL, TKI is allowed in combination with any of the above regimens;for patients who are intolerant to vincristine, another alkaloid may be used.Simultaneous treatment with intrathecal methotrexate should be avoided during FLAG chemotherapy administration. ALL, acute lymphoblastic leukemia; ANC, neutrophil count; FLAG, fludarabine, highA dose of cytarabine and G-CSF; G-CSF, granulocyte colony-stimulating factor; IV, intravenous; ph, philadelphia chromosome; PO, oral administration; SC, subcutaneous; TKI, tyrosine kinase inhibitors; VAD, vincristine, doxorubicin and dexamethasone.
At day 0 at 2X 10 6 Or 1X 10 6 KTE-X19 (68 mL or 40mL formulation) was administered at the corresponding target dose of each CAR T cell/kg. A minimum of 7 days of hospitalization is required after infusion, followed by response assessment at a predetermined time point. All patients who completed the 3 rd month visit were subjected to a long follow-up period of survival and disease status lasting up to 15 years every 3 months up to 18 months, every 6 months between 24 th month and 60 th month, and once a year. If the patient withdraws consent for further follow-up, no follow-up or death, it can be removed from the study. Allogeneic stem cell transplantation (alloSCT) is not a requirement of the protocol, but is allowed by researchers as appropriate.
Accept 1X 10 6 Patients with individual CAR T cells/kg (40 mL) were treated under revised toxicity management guidelines: if in the case of Cytokine Release Syndrome (CRS), tolizumab is administered only for Neurological Events (NEs) and the steroid is initiated for level 2 NEs and for 1×10 NEs 6 Grade 3 NE-initiated steroids in the initial toxicity management guidelines for the individual CAR T cells/kg (68 mL) cohort (table 11).
TABLE 11 original and revised guidelines for neurotoxicity management。
* The prophylactic levetiracetam is applicable to all classes. The initiation of replacement therapy should be discussed with medical monitors including, but not limited to, anakinra, cetuximab, lu Suoti n, cyclophosphamide, intravenous immunoglobulins and anti-thymocyte globulin.
BID, twice daily; CRS, cytokine release syndrome; CSF, cerebrospinal fluid; EEG, electroencephalogram; ICU, intensive care unit; IV, intravenous; MRI, magnetic resonance imaging;
results and assessment
The primary endpoint of stage 1 was the incidence of DLT in the DLT evaluable group, which included the use of KTE-X19 at 2X 10 6 The first 3 patients treated with each CAR T cell/kg dose. Secondary endpoints included safety, overall CR rate (cr+cri), duration of remission (DOR), MRD negative rate, alloSCT rate, OS, and Relapse Free Survival (RFS). Adverse events including individual symptoms of Cytokine Release Syndrome (CRS) and Neural Events (NE) were graded according to NCI adverse event common terminology standard (AE) version 4.03. CRS is according to Lee et al, blood.2014;124 And (2) modified standard grading of 188-195. The overall response was determined by the investigator after bone marrow and peripheral blood assessment, as detailed in the table below. (Cheson BD et al, J Clin Oncol.2007;25 (5): 579-586). Bone marrow evaluation and response evaluation were performed on day 28 and on month 2 and 3. In patients receiving bridging chemotherapy, additional bone marrow aspirate is required between day-4 (+/-2 days) at the end of bridging chemotherapy. For patients with extramedullary disease, the response is assessed according to the response criteria for extramedullary and CNS disease in the revised International working group malignant lymphoma criteria, as detailed in the following table. (Cheson BD et al, J Clin Oncol.2007;25 (5): 579-586). Minimal Residual Disease (MRD) was tested by flow cytometry (Neogenomics) with a sensitivity of 0.01% using the following markers: CD3, CD9, CD10, CD13/CD33, CD19, CD20, CD34, CD38, CD45, CD58 and CD71. (Gupta S et al, leukemia.2018;32 (6): 1370-1379; borowitz MJ et al, blood.2015;126 (8): 964-971; bruggemann M et al, hematology Am Soc Hematol Educ program.2017;2017 (1): 13-21). MRD negative assay Defined as evaluating according to a standard, MRD<10 -4 . The MRD of a portion of the bone marrow aspirate taken on day 28 and month 2 and month 3 was analyzed. The products, blood and tumor samples were subjected to translational analysis to evaluate the pharmacokinetic and pharmacodynamic profile of KTE-X19 in pediatric R/R B-ALL as an exploratory endpoint. Pharmacokinetic and pharmacodynamic evaluations have been previously described as well as associations with clinical outcomes. (Locke FL et al Mol Ther.2017;25 (1): 285-295). The overall disease response is as described in table 12.
TABLE 12 general disease response Classification。
* The units of Plt and ANC are per μl. ANC and Plt values should be evaluated each time a BM evaluation is performed. If not, the ANC and Plt values used for the reaction evaluation may be anywhere from 7 days before the BM results to anywhere after the BM results. § In patients evaluating non-CNS EMD, imaging and bone marrow outcome for assessing overall disease response must be within 30 days of each other.Morphological mother cell in BM. # For patients who achieved CR, the earlier was on day 28 or when the reaction was first assumed. ** Non-circulating leukemia refers to morphologically circulating blast cells<1%. The circulating leukemia is morphological circulating blast cell more than or equal to 1%. If morphologically the blast is 1% or more, and there is no evidence of other leukemias, a flow or molecular study should be performed to confirm that the blast is leukemic. ANC, absolute neutrophil count; BM, bone marrow; CNS, central nervous system; CR, complete remission; CRh, completely relieving the concomitant partial hematological recovery; CRi, complete remission response with incomplete hematological recovery; EMD, marrow External diseases; PD, progressive disease; plt, platelets; PR, partial reaction.
Results
Patient(s)
Between day 2016, 2, 17 and day 2018, 8, 1, 31 patients were enrolled and a leukapheresis procedure was performed. The median time from leukocyte isolation to release of KTE-X19 product was 14.0 days (range, 9.0-20.0) for all treated patients, 16.5 days (range, 12.0-23.0) from leukocyte isolation to delivery to the study center, and 27.0 days (range, 18.0-41.0) from leukocyte isolation to infusion. The data cut-off value is 9 months and 9 days in 2020. Of the 31 patients in the group, 24 (77%) received conditioning chemotherapy and were subsequently administered. Seven patients were not dosed for the following reasons: adverse events (AE; n=1), product unavailable (n=3), failed due to AE (n=1), product unavailable and failed (n=1), and death (n=1). Twenty four patients received conditioning chemotherapy followed by KTE-X19;4 patients received 2X 10 6 11 patients received 1X 10 dose of CAR T cells/kg 6 Several CAR T cells/kg (68 mL) dose formulation, and 9 patients received 1X 10 6 Individual CAR T cells/kg (40 mL) dose formulation. The median follow-up for all treated patients was 36.1 months (range, 24.0-53.9). The median age of the treated patient was 13.5 years (range, 3-20); 42% of patients have received more than or equal to 3 past line counts of therapy; 29% suffer from primary refractory disease; 25% R/R after alloSCT; and median myeloblasts at screening were 44% (range, 6-99; table 13). Prior to the group, 6 (25%) patients experienced prior alloSCT,8 (33%) patients received prior boswellizumab, including 3 (13%) patients who received boswellizumab as the last prior therapy, and 1 (4%) had extramedullary disease. Of the 31 patients in the group, 30 (97%) received each regimen of bridging therapy, with new baseline disease assessments performed just prior to lymphoproliferative chemotherapy. Safety and efficacy analyses were performed on all 24 patients dosed.
TABLE 13 patient characteristics
Safety of
At the time of receiving 2X 10 6 Of the 3 DLT-evaluable patients with individual CAR T cells/kg, no DLT was observed. All treated patients (n=24) experienced at least one grade-3 AE, most commonly hypotension (50%) and anemia (42%; table 14). Any level of severe AE occurred in 71% of patients. Grade 3 infection occurred in 42% of patients.
TABLE 14 adverse events
* The table includes any level of adverse events that occur in ≡20% of all patients
Among all treated patients (n=24), CRS was reported in 21 patients (88%), of which 8 patients (33%) underwent grade 3 CRS according to the modified Lee grading standard (table 15). Lee DW et al Blood 124:188-95,2014. No class 4 or class 5 CRS events occur. The most common grade 3 CRS symptoms are hypotension (50%) and fever (25%). Any grade and ≡3 grade hypoxia was observed in 13% and 8% of patients, respectively. The median time to CRS onset and duration after KTE-X19 infusion were 5 days (range, 1-14) and 7 days, respectively, with all events resolved.
TABLE 15 cytokine release syndrome and neurological events
* Including CRS symptoms and neurological events occurring in ≡10% of all patients. *** The cytokine release syndrome is classified according to a modified classification system proposed by Lee et al Blood, 2014.
Of all treated patients, any grade of NE (67%) was reported in 16 patients, and grade 3 event (21%) occurred in 5 patients, with encephalopathy (13%) being the most common grade 3 event (Table 15). A grade 4 fully reversible NE (cerebral edema) occurred in patients receiving a dose of 1×106 CAR T cells/kg (68 mL); for this event, patients were treated with dexamethasone, mannitol, sodium chloride, and tolizumab. There is no level 5 NE. Overall, the median time to NE onset after injection was 9.5 days (range, 3-60), and the median duration of NE was 8 days. NE resolved in 14 of the 16 patients (88%). NE from the remaining two patients is ongoing at death due to AE (n=1) or progressive disease (n=1).
Of all treated patients, 42% received steroid, 63% received tolizumab, and 46% received vasopressor (table 15). Relative to the use of 2X 10 under the original guidelines 6 4 patients treated with individual CAR T cells/kg and with 1X 10 6 11 patients treated with CAR T cells/kg (68 mL) were treated with 1X 10 under revised toxicity management 6 Improved overall safety was observed in 9 patients treated with a dose of CAR T cells/kg (40 mL). And accept 1X 10 6 27% and 22% of patients receiving 2X 10 CAR T cells/kg (68 mL and 40mL, respectively) compared to each other 6 Of the patients with CAR T cells/kg, 75% experienced grade 3 CRS. At 25% receive 2X 10 6 Patients with individual CAR T cells/kg and 27% of recipients 1X 10 6 Grade 3 NE or more was observed in patients with individual CAR T cells/kg (68 mL),but at receiving 1X 10 6 The lowest (11%) of the patients with individual CAR T cells/kg (40 mL). In addition, with 2X 10 6 The median time to onset of NE and CRS was 1X 10 compared to the individual CAR T cells/kg dose cohort 6 Delays appear in individual CAR T cells/kg dose cohorts.
Of the 8 patients (33%) who died in the study, 6 died from progressive disease (median 190.5 days after KTE-X19 infusion), and 2 died from AEs considered to be independent of KTE-X19 (except grade 5B-ALL), including disseminated mucormycosis (n=1, day 15 after KTE-X19 infusion) and Escherichia (Escherichia) sepsis (n=1, day 409 after KTE-X19 infusion). Of the dead patients, 3 received 2×10 patients 6 CAR T cells/kg, 4 patients received 1X 10 6 CAR T cells/kg (68 mL), and 1 patient received 1X 10 6 Each CAR T cell/kg (40 mL). At any time, no patient tested positive for replication competent retrovirus or anti-CD 19 CAR antibodies.
Efficacy of
Median follow-up time was 36.1 months (ranging from 24.0-53.9), and efficacy assessment was performed for all patients receiving treatment (n=24). The total remission rate assessed by the investigator was 67%,29% of the patients (n=7) achieved CR, and 38% of the patients (n=9; table 16) achieved CRi. At 2X 10 6 、1×10 6 (68 mL) and 1X 10 6 The CR+CRi rates were 75%, 64% and 67%, respectively, in the (40 mL) CAR T cell/kg dose group. The median time from infusion to cr+cri across dose levels was 30 days (range, 26 days to 113 days). The total MRD negative rate was 100% among 16 cr+cri patients. A total of sixteen patients (67%) received alloSCT as a subsequent consolidation therapy, including at 2 x 10 each 6 、1×10 6 (68 mL) and 1X 10 6 (40 mL) 2, 8 and 6 patients in the CAR T cell/kg dosing group. Fourteen (88%) out of 16 patients who achieved cr+cri (at 2×10 6 、1×10 6 [68mL]And 1X 10 6 [40mL ]]CAR T cells/kg dosing groups were 2, 7 and 5), respectively, subjected to subsequent alloSCT. Two of these patients relapsed before subsequent alloSCT; all accept before proceeding with alloSCT Consolidated chemotherapy is provided. Of 2 patients who achieved cr+cri but did not receive subsequent alloSCT, 1 died due to progressive disease, 1 failed visit. The 2 patients who did not obtain any response were subjected to subsequent alloSCT and CR was obtained as their response to alloSCT. After KTE-X19, the median time to transplantation was 2.3 months (range 1.4-24.9) for all patients receiving treatment.
TABLE 16 remission rate and minimal residual disease state
* MRD negatives were assessed by flow cytometry at day 28 and month 2 and month 3 with a sensitivity of 0.01%. MRD results after allogeneic stem cell transplantation or new anti-cancer therapies are excluded. MRD, minimal residual disease.
Median DOR of 16 patients achieving CR+CRi after KTE-X19 was 7.2 months (95% CI, 4.1-14.2) after examination of subsequent alloSCT, and at 2X 10 6 、1×10 6 (68 mL) and 1X 10 6 The CAR T cells/kg dosing group (40 mL) were 4.1 months, 10.7 months, respectively, and were not reached. Median DOR was 14.2 months (95% CI,3.9 NE) without review of subsequent alloSCT and recovery of tyrosine kinase inhibitors. Median DOR in 14 patients with cr+cri who received subsequent alloSCT after KTE-X19 was 10.7 months (95% CI, 7.2-14.2). Median RFS for all treated patients (n=24) was 5.2 months (95% CI, 0.0-17.8). Not up to 1X 10 6 Median RFS of individual CAR T cells/kg (40 mL) group, and at 2×10 6 Sum is 1×10 6 The (68 mL) cell/kg cohorts were 5.2 months (95% CI, 0.0-5.2) and 9.1 months (95% CI, 0.0-17.8), respectively. Median RFS in 16 patients with subsequent alloSCT was 9.1 months (95% CI, 9.1-17.8). In all treated patients and in 1X 10 6 Median OS was not reached in the individual CAR T cells/kg dose group, and for 2X 10 6 The CAR T cell/kg dose group was 8.0 months. 1X 10 6 24 months OS Rate at a dose of individual cells/kg (40 mL) for 1X 10 6 Individual cells/kg(40 mL) dose was 87.5% (95% CI, 38.7-98.1), and for 1X 10 6 The individual cells/kg (68 mL) dose was 72.7% (95% CI, 37.1-90.3). In general, 33% of the treated patients (8/24) died by the date of data expiration, 1 patient stopped treatment due to complete consent withdrawal, and 1 patient was out of visit. The remaining 58% of patients (14/24) remain alive and continue to follow up at the time of data collection, all of which received subsequent alloSCT after KTE-X19. Based on safety and efficacy data analysis, RP2D is 1×10 6 KTE-X19 cells/kg (40 mL formulation) and toxicity management was modified.
Translation analysis
CAR T-cell proliferation in peripheral blood, measured by Polymerase Chain Reaction (PCR) and expressed as CAR gene copy number in blood/μg DNA, was observed in each dosing group, with peak CAR T-cell levels reached on day 14, followed by CAR T-cell shrinkage to baseline (table 17). Median CAR T cell levels in all dose groups were undetectable in blood by PCR 3 months after KTE-X19 infusion (table 17). Median peak CAR copy number/. Mu.g DNA blood at 1X 10 6 The CAR T cell/kg dose cohorts were similar but at 2X 10 6 Higher in the individual CAR T cell/kg queue (fig. 10A). Patients who achieved cr+cri tended to have higher peak blood CAR gene copy number per μg DNA in blood than non-responders, as does MRD-negative patients relative to MRD-positive patients (fig. 10C). The CAR gene copy number/g DNA was higher in blood of grade 3 NE patients than grade 2 NE patients (FIG. 10D), whereas there was no significant difference in peak CAR gene copy number/g DNA in blood of high or low grade CRS patients in this limited sample size. Median peak CAR gene copy number/. Mu.g DNA in blood was 5.16X10 in 16 patients without prior Bonauzumab 4 (range, 0-2.40X10) 5 ) And 6.15X10 of 8 patients with the prior Bonus mab 3 (range, 0-2.49X10) 5 )。
TABLE 17 CAR Gene copy number in blood over time
Peak levels of various key serum cytokines, chemokines and pro-inflammatory biomarkers occurred on day 7. Commensurate with peak CAR amplification, 1×10 6 Several serum analytes at 2X 10 compared to CAR T cells/kg 6 Tends to be higher in the patients administered (interleukin [ IL ]]-2, IL-5, IL-6, IL-8, IL-10, IL-15, IL-16, ferritin, granzyme B, intercellular adhesion molecule 1[ ICAM-1 ] ]Interferon gamma [ IFN-gamma ]]And tumor necrosis factor alpha [ TNF-alpha ]](FIG. 11; table 18).
TABLE 18 peak selection of serum biomarkers at baseline and post-infusion
Peak serum levels of analytes VCAM-1 and IL-16 correlated with grade 3 CRS. This association was not observed in subjects with ≡3NE, probably due to the small number of patients with ≡3NE (table 19).
TABLE 19 correlation of serum biomarkers with cytokine release syndrome and neurological events。
* The value represents the lower limit of quantification of the assay used.The value represents the upper limit of quantification of the assay used. CRP, C-reactive protein; CXCL, C-X-C motif chemokine ligands; GM-CSF, granulocyte-macrophage colony stimulating factor; IFN-gamma, interferon gamma; ICAM, intercellular adhesion molecules; IL, interleukin; IP, interferon gamma-inducible protein; MCP, monocyte attractant protein; rα, receptor α; RA, receptor antagonists; SAA, serum amyloid A; TNF- α, tumor necrosis factor α; VCAM, vascular cell adhesion molecule.
The product characteristics were similar at different dose levels (table 20). The levels of lower differentiated CCR7+ T cells in the product were higher in patients with CR+ CRi and tended to be higher in MRD-negative patients (Table 21).
TABLE 20 characteristics of dosed products。
* Co-culture experiments were performed using Toledo cells mixed with KTE-X19 product cells at a 1:1 ratio. IFN-. Gamma.was measured in cell culture medium 24 hours after incubation using a qualified ELISA. ELISA, enzyme-linked immunosorbent assay; IFN-gamma, interferon gamma;
this product profile also showed a trend to have higher levels of neurotoxicity, but was independent of CRS. The ratio of CD4 to CD 8T cells is independent of response or toxicity.
TABLE 21 characterization of products by response, MRD status, cytokine release syndrome and nervous system events。
CR, complete remission; CRi, complete relief with incomplete hematological recovery; CRS, cytokine release syndrome; MRD, minimal residual disease; NE, neural event.
In phase 1 of clinical trial-4, no DLT of KTE-X19 was observed in pediatric or adolescent patients with a DLT of R/R B-ALL that could be assessed. Although at 2X 10 6 No DLT was observed at the initial dose of individual CAR T cells/kg, but 1 x 10 with 68mL formulation was explored in the second cohort of patients 6 Lower doses of individual CAR T cells/kg in an attempt to further improve risk to benefit ratio, and in the third cohort with 1 x 10 with 40mL formulation 6 Individual CAR T cells/kg and modified toxicity management further optimize dose and toxicity management. This results in a 1×10 for 6 More optimal risk to benefit ratio for individual CAR T cell/kg (40 mL) dose levels, with significant improvement for CRS and NE. In addition, although the MRD negative rate of all the preparations was 73% or more, 1X 10 was accepted 6 Among patients with individual CAR T cells/kg (40 mL), MRD negative rate and CR rate alone were highest. After 36.1 months of follow-up in all treated patients, the patients were followed at 1X 10 6 Median DOR, RFS and OS were still not reached in the individual CAR T cell/kg (40 mL) cohort, and the 24 month OS ratio was 87.5%, potentially indicating a meaningful persistence response with optimized dose/formulation of KTE-X19 in pediatric/adolescent patients with R/R B-ALL.
In pediatric/adolescent patients with R/R B-ALL, anti-CD 19 CAR T-cell therapyThe role of post alloSCT is still not quite clear; studies in the adult population provide somewhat conflicting results. (Park JH et al, N Engl J Med 378:449-459,2018; hay KA et al, blood133:1652-1663, 2019). In this study, following a follow-up at a middle of 36.1 months, the study was followed up at 1X 10 6 Median DOR and OS examined at the time of subsequent alloSCT (median of all treated patients) was not reached in RP2D treated patients of individual CAR T cells/kg (40 mL). Fourteen (88%) (including 5 patients treated with RP 2D) of 16 patients who achieved cr+cri received alloSCT as follow-up therapy. alloSCT is not a protocol requirement, but is allowed by researchers as appropriate. Although clinical trial-4 was not designed to evaluate the results after follow-up therapy, DOR evaluation without review of follow-up therapy including alloSCT showed a favorable median of 14.2 months given that most of the responding patients underwent alloSCT following KTE-X19. In addition, the median RFS for examination of subsequent alloscts was 5.2 months, but the median RFS for no examination was 9.1 months, indicating the potentially beneficial effect of alloSCT after KTE-X19. Pediatric and young adults with R/R CD19+ ALL who have no history of alloSCT but who receive consolidated alloSCT following anti-CD 19 CAR T-cell therapy have previously been reported to have a tendency to have no increased leukemia survival in a follow-up of > 1 year. (Summers C et al, blood 132:967-967,2018.). In the recently disclosed phase I study of anti-CD 19 CAR T-cell therapy in children and young adults with R/R B-ALL in which 75% of MRD negative response patients continued with alloSCT, the median OS at the 4.8 year follow-up was 70.2 months after alloSCT, 5 year event-free survival after alloSCT was 61.9%, and the cumulative incidence of relapse after alloSCT was only 9.5%. (Shah NN. et al Journal of Clinical Oncology0: JCO.20.02262). These data indicate that subsequent alloSCT may be important for maintaining relief after CAR T cell therapy in pediatric R/R B-ALL. Retrospective reviews of pediatric and young adult patients have found that CD34 selected T-cell depleted alloSCT following CAR T-cell therapy can result in improved graft-related mortality and OS compared to unmodified alloSCT. (Fabrizio VA et al Bone Marrow Transplant 55:2160-2169,2020). Median blood CAR T cell levels in clinical trial-4 all doses 3 months after infusion None of the amounts could be detected, with a median time of alloSCT of 2.3 months, which should take into account the relationship between CAR T cell persistence and response persistence, as the overall patient number is lower and the subsequent alloSCT ratio is higher. In the investigation of tisaganlecteics, and contrary to our investigation, subsequent alloSCT was performed in a few reactive patients (12% to 13%), with a short median follow-up of 13.1 months, whereas about 40% of reactive patients receiving tisaganlecteics had relapsed, mostly CD19 negative leukemia, despite the presence of sustained CAR T cells. (Maude SL et al, N Engl J Med 378:439-448,2018;Grupp SA et al, blood 132:895-895,2018;Pasquini MC et al, blood Adv 4:5414-5424,2020.).
While differences in trial design and patient population precluded direct trial-trial comparisons, recent studies with bolafirt mab (which also targets CD 19) demonstrated a median OS of only 7.7 months in pediatric R/R B-ALL, similar to the results in adult ALL. (Kantarjian H et al, N Engl J Med 376:836-847,2017). Furthermore, for bolamitraz, combination with subsequent alloSCT has presented improved results (12 month RFS rates with and without subsequent alloSCT: 70% and 30%, respectively). (Locatelli F et al, blood 136:24-25,2020). In addition, the remission rate of boscalid was higher in pediatric patients with lower baseline tumor burden (< 50% blast; 56% CR) than those with higher tumor burden (> 50% blast; 33% CR) at baseline. (von Stackelberg A et al, J Clin Oncol34:4381-4389, 2016). In clinical trial-4, the clear correlation between remission rate and baseline bone marrow blast is not apparent, as the CR rates for patients with >5 to ≡25%, >25 to ≡50%, >50 to ≡75% and >75 to 100% blast cells were 89%, 25%, 100% and 50% respectively at baseline tumor burden. However, the interpretation was limited by the small number of patients in each quartile and the relatively high median tumor burden at baseline (fig. 12). This is consistent with another pediatric and young adult study using CAR T-cell therapy for CD19, with no differences in response rate based on disease burden. (Gardner RA et al Blood 129:3322-3331,2017). The data from clinical trial-4 demonstrate that KTE-X19 has the potential to provide more beneficial efficacy in patients with high disease burden than reported with boletlizumab. In clinical trial-4, a trend toward lower cr+cri rates appears to be observed in patients with prior bolafuximab.
The AE profile in clinical trial-4 was consistent with previous studies of anti-CD 19 CAR T-cell therapy. For 24 patients receiving KTE-X19, the median time from leukocyte isolation to delivery to the study center was 16.5 days. In contrast, tisagenlect leucophore had a median throughput time of 23 days from the time of receiving the leukocyte isolation product to shipping to the study center. (Tyagarajan S et al Mol Ther Methods Clin Dev 16:136-144,2020). The rapid turnaround time of patients treated in clinical trial-4 supported the feasibility of setting up rapidly proliferating ALL. With the establishment of RP2D, clinical trial-4 has transitioned to the phase 2 portion of the study.
The unmet medical need in R/R pediatric ALL is highest for patients with early relapse or primary refractory disease, with 5 year OS rates of 21% to 28%. (Sun W et al, leukemia 32:2316-2325,2018;Crotta A et al, curr Med Res Opin 34:435-440,2018;Nguyen K et al, leukemia 22:2142-50,2008;Rheingold SR et al, journal of Clinical Oncology 37:10008-10008,2019;Oskarsson T et al, haemato logica 101:68-76,2016;Schrappe M et al, N Engl J Med 366:1371-81,2012). Furthermore, the risk of morbidity and mortality associated with treatment is 3-to 5-fold in patients with MRD-positive disease at the end of the initial and later treatment lines than in patients with undetectable MRD. 3 to address this ongoing unmet medical need, clinical trial-4 was further modified to expand the eligibility criteria to include patients with MRD-positive disease as well as patients with early first relapse (. Ltoreq.18 months). In addition, a second cohort was opened for pediatric patients with R/R NHL (diffuse large B-cell lymphoma, burkitt's lymphoma, and primary mediastinal B-cell lymphoma).
Example 5
An open-label, global, multicenter phase 3 study was conducted in adult patients with recurrent or refractory diffuse large B-cell lymphoma (DLBCL) to assess the safety and efficacy of current standard of care for alemtuzite and second line therapies (a combination platinum-based rescue chemotherapy regimen followed by high-dose therapy and autologous stem cell transplantation in patients who respond to rescue chemotherapy). In this study 359 patients were randomized (1:1) to receive current standard of care for single infusion of aliskiren or two-line therapy. The primary endpoint is Event Free Survival (EFS), defined as the time from random grouping to 20 th day of 9 months according to the Lukeno classification (see Cheson et al, J Clin Oncol.2014; 32 (27): 3059-68) at which disease progression occurs earliest, new lymphoma treatment begins, or death is caused by any cause. Key secondary endpoints include Objective Response Rate (ORR) and Overall Survival (OS). Other secondary endpoints include modified event-free survival, progression-free survival (PFS), and duration of response (DOR). Patients participating in this study range in age from 22 to 81 years, with 30% of patients exceeding 65 years of age. The study described in this example evaluates the single infusion of cell therapy, alopecie, in contrast to the second line standard of care (SOC) in adult patients with recurrent or refractory LBCL. The study of the SOC group is a 2-step process: after the initial relapse, the immune chemotherapy is reintroduced, and if the patient is responsive and can tolerate further treatment, the high dose chemotherapy plus stem cell transplantation is continued.
Critical inclusion criteria：
1. Histologically demonstrated large B cell lymphomas, including the following types defined by WHO 2016 (see Swerdlow et al blood.2016, month 5, 19; 127 (20): 2375-90.Doi:10.1182/blood-2016-01-643569.2016, month 3, 15 electronic publications. Reviews).
DLBCL (ABC/GCB) without additional designation
HGBL with or without MYC and BCL2 and/or BCL6 rearrangements
DLBCL caused by FL
T cell/tissue cell enriched large B cell lymphomas
DLBCL associated with chronic inflammation
Primary skin DLBCL, leg type
Epstein-Barr Virus (EBV) +DLBCL
2. Recurrent or refractory diseases after first-line chemotherapy
Refractory disease is defined as first line therapy not fully alleviated; individuals intolerant to first-line therapy are excluded.
Progressive Disease (PD) is the best response to first-line therapy
Disease Stabilization (SD) is an optimal response after at least 4 cycles of first line therapy (e.g., 4 cycles of R-CHOP)
Partial Response (PR) is a therapy with optimal response after at least 6 cycles and biopsy-demonstrated residual disease or disease progression of 12 months or less
Recurrent disease is defined as first line therapy with complete remission followed by biopsy-confirmed recurrence for less than or equal to 12 months
3. The individual must have received sufficient first line therapy, including at least:
anti-CD 20 monoclonal antibodies unless the investigator determines that the tumor is CD20 negative, an
Anthracycline-containing chemotherapy regimens
4. No lymphomas have a known history or suspicion of involvement of the central nervous system
5. Eastern tumor collaboration group (ECOG) physical stamina of 0 or 1 in the united states
6. Sufficient bone marrow function as demonstrated below:
absolute Neutrophil Count (ANC) greater than or equal to 1000/uL
Platelets not less than 75,000/uL
Absolute lymphocyte count is greater than or equal to 100/uL
7. Sufficient kidney function, liver function, heart function and lung function as demonstrated below:
creatinine clearance (Cockcroft Gault) is not less than 60mL/min
Serum alanine aminotransferase/aspartate aminotransferase (ALT/AST) is less than or equal to 2.5 upper normal limit (ULN)
Total bilirubin is less than or equal to 1.5mg/dl
The cardiac ejection fraction is greater than or equal to 50%, there is no evidence of pericardial effusion, as determined by Echocardiography (ECHO), and there is no clinically significant Electrocardiogram (ECG) results
No clinically significant pleural effusion
Baseline oxygen saturation in indoor air >92%
The key exclusion criteria were:
1. history of malignancy other than non-melanoma skin cancer or carcinoma in situ (e.g., cervical, bladder, breast) unless disease is absent for at least 3 years
2. More than one line of therapy is accepted for DLBCL
3. History of autologous or allogeneic stem cell transplantation
4. The presence of fungi, bacteria, viruses or other infections is uncontrolled or requires intravenous antimicrobial administration.
5. Has a known history of infection with Human Immunodeficiency Virus (HIV) or hepatitis b (HBsAg positive) or hepatitis c virus (anti-HCV positive). If there is a positive history of treating hepatitis B or C, the viral load must be undetectable by quantitative Polymerase Chain Reaction (PCR) and/or nucleic acid testing.
6. An individual having a detectable cerebrospinal fluid malignant cell or a known brain metastasis or a medical history of a cerebrospinal malignant cell or brain metastasis.
7. Non-malignant Central Nervous System (CNS) disorders such as epilepsy, cerebral vascular ischemia/hemorrhage, dementia, cerebellar disease, or the history or presence of any autoimmune disease involving the CNS
8. There are any indwelling wires or tubes. A dedicated central venous access catheter, such as a Port-a-Cath or Hickman catheter, is permissible.
9. Myocardial infarction, cardiac angioplasty or stenting, unstable angina, new York Heart Association grade II or higher congestive heart failure, or other clinically significant history of heart disease within 12 months of the group
10. History of symptomatic deep vein thrombosis or pulmonary embolism within 6 months of the group entry
11. History of autoimmune disease, systemic immunosuppression and/or systemic disease modulators were needed over the last 2 years
12. History of anti-CD 19 or CAR-T therapy or history of previous random groupings
The main analysis of this study shows that in secondary recurrent or refractory large B-cell lymphomas (LBCL), aliskiren has superiority compared to the standard of care (SOC). The study reached a primary endpoint for event-free survival (EFS; risk ratio 0.398, p < 0.0001), and a critical secondary endpoint for Objective Response Rate (ORR). Mid-term analysis of Overall Survival (OS) showed a trend toward favorable alemtujopsis, but the data was still immature and may require additional analysis and/or study.
The safety results of this study are consistent with the known safety profile of alopecie for the treatment of LBCL in a three-wire environment. 6% of patients experience CRS of grade 3 or higher, and 21% experience neurological events of grade 3 or higher. No new security issues are found in this two-wire environment.
T cell phenotypes were assessed in the alzem product from this study, using validated flow cytometry methods and analyzing the known surface markers CCR7 and CD45RA for T cell differentiation. Metrics were analyzed by the grade of Cytokine Release Syndrome (CRS) and neural events. P-values were obtained by the Kruskal-Wallis test.
Table 22 shows the total number of infused central memory T cells (ccr7+cd45ra-) associated with the occurrence of > =2 class CRS events (compared to class 1 and no events), while table 23 shows the total number of infused effector memory T cells and effector T cells (CCR 7-) associated with the occurrence of > =3 class neurological events (compared to class 2, class 1 and no events). These data reveal that different T cell subsets in alemtuquor are associated with differences in toxicity.
Table 22: and (3) with>Central memory T-thin for infusions correlated with the occurrence of class 2 CRS event (compared to class 1 and no event) 6 Total number of cells (CCR7+CD45RA-) (cell count. Times.10)
Table 23: infusion effect records associated with the occurrence of ≡3 neurological events (compared to grade 2, grade 1 and no events) 6 Total number of memory and effector T cells (CCR 7-) (cell count. Times.10)
Example 6
This example provides additional and subsequent results from example 4 above.
Two dosage levels and formulations were studied in phase 1; the primary endpoint was the incidence of Dose Limiting Toxicity (DLT). Of the 31 patients in the group, KTE-X19 was administered to 24 patients (median age 13.5 years, range 3-20; median follow-up 36.1 months). No DLT was observed. All treated patients had grade 3 or more adverse events, typically hypotension (50%) and anemia (42%). The level 3 cytokine release syndrome ratio was 2×10 in all treatments 6 、1×10 6 (68 mL of formulation) and 1X 10 6 33%, 75%, 27% and 22% in the individual (40 mL formulation) CAR T cells/kg group; 21%, 25%, 27% and 11% of patients experience > 3 grade neurological events, respectively. In all treatments, 2X 10 6 、1×10 6 (68 mL) and 1X 10 6 Total Complete Remission (CR) rates (including CR with incomplete hematological recovery) were 67%, 75%, 64% and 67%, respectively, in the (40 mL) CAR T cell/kg group. Overall Minimal Residual Disease (MRD) -negative rate was 100% among the responders; 88% of the responders underwent subsequent allogeneic stem cell transplantation (alloSCT). At 1X 10 6 In the (40 mL) group (recommended phase 2 dose), median duration of remission was checked at alloSCT and median overall survival was not achieved. Pediatric/adolescent patients with R/R B-ALL achieve high MRD-negative remission rates after a single dose of KTE-X19 with manageable safety. Phase 2 at 1X 10 6 Individual CAR T cells/kg (40 mL) doses were taken.
Results
Patient(s)
Between day 2016, 2, 17 and day 2018, 8, 1, 31 patients were enrolled and a leukapheresis procedure was performed. For all treated patients, the median time from leukocyte isolation to release of KTE-X19 product was 14.0 days (range 9.0-20.0), the time from leukocyte isolation to delivery to the study center was 16.5 days (range 12.0-23.0), and the time from leukocyte isolation to infusion was 27.0 days (range 18.0-41.0). Of the 31 patients in the group, 24 (77%) received conditioning chemotherapy and were subsequently administered. Seven patients were not dosed for the following reasons: adverse events (AE; n=1), unsuccessful product manufacture (n=3), failed due to AE (n=1), unsuccessful product manufacture and failed (n=1), and death (n=1). Twenty-four patients received conditioning chemotherapy followed by KTE-X19;4 patients received 2X 10 6 11 patients received 1X 10 dose of CAR T cells/kg 6 Several CAR T cells/kg (68 mL) dose formulation, and 9 patients received 1X 10 6 Individual CAR T cells/kg (40 mL) dose formulation. The median follow-up for all treated patients was 36.1 months (range, 24.0-53.9). The median age of the treated patient was 13.5 years (range, 3-20); 42% of patients have received more than or equal to 3 past line counts of therapy; 29% suffer from primary refractory disease; 25% R/R after alloSCT; and median myeloblasts at screening were 44% (range, 6-99). Prior to the group, 6 (25%) patients underwent past alloSCT,8 (33%) patients received past boltzumab, including 3 (13%) patients who received boltzumab as the last past therapy, and 1 (4%) had extramedullary disease. Of the 31 patients in the group, 30 (97%) received each regimen of bridging therapy, with new baseline disease assessments performed just prior to lymphoproliferative chemotherapy.
Safety of
At the time of receiving 2X 10 6 Of the 3 DLT-evaluable patients with individual CAR T cells/kg, no DLT was observed. All treated patients (n=24) experienced at least one grade-3 AE, most commonly hypotension (50%) and anemia (42%). Any level of severe AE occurred in 71% of patients. Grade 3 infection occurred in 42% of patients.
CRS was reported in 21 of 24 treated patients (88%), 8 of which (33%) underwent grade 3 CRS according to the modified Lee grading standard. 33 does not occur a class 4 or class 5 CRS event. The most common grade 3 CRS symptoms are hypotension (50%) and fever (25%). Any grade and ≡3 grade hypoxia was observed in 13% and 8% of patients, respectively. The median time to CRS onset and duration after KTE-X19 infusion were 5 days (range, 1-14) and 7 days, respectively, with all events resolved.
Of all treated patients, any grade of NE (67%) was reported in 16 patients, and grade 3 event (21%) occurred in 5 patients, with encephalopathy (13%) being the most common grade 3 event. At receiving 1X 10 6 One grade 4 fully reversible NE (cerebral edema) occurred in patients with CAR T cells/kg (68 mL); to manage this event, patients were treated with dexamethasone, mannitol, sodium chloride, and tolizumab. There is no level 5 NE. Overall, the median time to NE onset was 9.5 days after injection (range, 3-60), and the median time from first CRS to first NE onset was 4 days (range, -3 to 52[ first CRS resolved after first NE onset in 4 patients ]) And the median duration of NE was 8 days. NE resolved in 14 of the 16 patients (88%). Due to AE (n=1) or progressive disease (n=1)1) NE in the remaining two patients is ongoing at death. Ten of the 16 patients experiencing NE (63%) had concurrent CRS.
In all treated patients, 42% received steroid, 63% received tolizumab, and 46% received vasopressor. Relative to the use of 2X 10 under the original guidelines 6 4 patients treated with individual CAR T cells/kg and with 1X 10 6 11 patients treated with CAR T cells/kg (68 mL) were treated with 1X 10 under revised toxicity management 6 Improved overall safety was observed in 9 patients treated with a dose of CAR T cells/kg (40 mL). And accept 1X 10 6 27% and 22% of patients receiving 2X 10 CAR T cells/kg (68 mL and 40mL, respectively) compared to each other 6 Of the patients with CAR T cells/kg, 75% experienced grade 3 CRS. At 25% receive 2X 10 6 Patients with individual CAR T cells/kg and 27% of recipients 1X 10 6 Grade 3 NE was observed in patients receiving CAR T cells/kg (68 mL), but at 1X 10 6 The lowest (11%) of the patients with individual CAR T cells/kg (40 mL). In addition, with 2X 10 6 The median time to onset of NE and CRS was 1X 10 compared to the individual CAR T cells/kg dose cohort 6 Delays appear in individual CAR T cells/kg dose cohorts.
Of the 8 patients (33%) who died in the study, 6 died from progressive disease (median 190.5 days after KTE-X19 infusion), and 2 died from AEs considered to be independent of KTE-X19 (except grade 5B-ALL), including disseminated mucormycosis (n=1, day 15 after KTE-X19 infusion) and escherichia sepsis (n=1, day 409 after KTE-X19 infusion). Of the dead patients, 3 received 2×10 patients 6 CAR T cells/kg, 4 patients received 1X 10 6 CAR T cells/kg (68 mL), and 1 patient received 1X 10 6 Each CAR T cell/kg (40 mL). At any time, no patient tested positive for replication competent retrovirus or anti-CD 19CAR antibodies.
Efficacy of
Median follow-up time was 36.1 months (range, 24.0-53.9), and efficacy assessment was performed for all treated patients (n=24). The total remission rate evaluated by the investigator was 67In%, 29% of patients (n=7) achieved CR, and 38% achieved CRi (n=9). At 2X 10 6 、1×10 6 (68 mL) and 1X 10 6 The CR+CRi rates were 75%, 64% and 67%, respectively, in the (40 mL) CAR T cell/kg dose group. The median time from infusion to cr+cri for each dose level was 30 days (range, 26-113 days). The total MRD negative rate was 100% among 16 cr+cri patients. A total of sixteen patients (67%) received alloSCT as a subsequent consolidation therapy, including at 2 x 10 each 6 、1×10 6 (68 mL) and 1X 10 6 (40 mL) 2, 8 and 6 patients in the CAR T cell/kg dosing group. Fourteen (88%) out of 16 patients who achieved cr+cri (at 2×10 6 、1×10 6 [68mL]And 1X 10 6 [40mL ]]CAR T cells/kg dosing groups were 2, 7 and 5), respectively, subjected to subsequent alloSCT. In addition, patients with CR (n=1) and bone marrow hypoplasia/bone marrow aplasia without blast cells (n=1) that achieved partial hematological recovery all proceeded to alloSCT; they then implement CR. After KTE-X19, the median time to transplantation was 2.3 months (range, 1.4-24.9) for all treated patients. Of 2 patients who achieved cr+cri but did not receive subsequent alloSCT, 1 died due to progressive disease, 1 failed visit.
Median DOR of 16 patients achieving CR+CRi after KTE-X19 was 7.2 months (95% CI, 4.1-14.2) after examination of subsequent alloSCT, and at 2X 10 6 、1×10 6 (68 mL) and 1X 10 6 The CAR T cells/kg dosing group (40 mL) were 4.1 months, 10.7 months, respectively, and were not reached. Median DOR was 14.2 months (95% CI, 3.9-unpredictable) without review of subsequent alloSCT and recovery of tyrosine kinase inhibitors. Median DOR in 14 patients with cr+cri who received subsequent alloSCT after KTE-X19 was 10.7 months (95% CI, 7.2-14.2). Median RFS for all treated patients (n=24) was 5.2 months (95% CI, 0.0-17.8). 1X 10 6 Median RFS in the individual CAR T cells/kg (40 mL) group was not reached and was found to be 2 x 10 6 And 1X 10 6 The (68 mL) cells/kg queue were 5.2 months (95% CI, 0.0-5.2) and 9.1 months (95% CI, 0.0-17.8), respectively. Median RFS was 9.1 months (95% CI, 9.1-17.8) in 16 patients followed by alloSCT. At the position ofIn all treated patients and in 1X 10 6 Median OS was not reached in each CAR T cell/kg dose group, and was found to be 2X 10 6 The CAR T cell/kg dose group was 8.0 months. For 1X 10 6 A dose of individual cells/kg (40 mL) with an OS ratio of 87.5% (95% CI, 38.7-98.1) for 24 months, and for 1X 10 6 The individual cells/kg (68 mL) dose was 72.7% (95% CI, 37.1-90.3). In summary, 8 out of 24 treated patients (33%) died by the date of data expiration, 1 discontinued due to consent withdrawal, and 1 missed visit. The remaining 14 patients (58%) remained alive and continued follow-up at the time of data collection, all patients receiving subsequent alloSCT after KTE-X19.
Based on safety and efficacy data analysis, RP2D is 1×10 6 KTE-X19 cells/kg (40 mL formulation) and toxicity management was modified.
Translation analysis
CAR T-cell proliferation in peripheral blood, measured by Polymerase Chain Reaction (PCR) and expressed as CAR copy number in blood/μg DNA, was observed in each dosing group, with peak CAR T-cell levels reached on day 14, followed by CAR T-cell shrinkage to baseline. Median CAR T cell levels in blood were not detectable by PCR in all dose groups 3 months after KTE-X19 infusion. Median peak CAR copy number/. Mu.g DNA blood at 1X 10 6 The CAR T cell/kg dose cohorts were similar but at 2X 10 6 Higher in each CAR T cell/kg queue. Patients who achieved cr+cri tended to have higher peak blood CAR gene copy number per μg DNA in blood than non-responders, as do MRD negative and MRD positive patients. The CAR gene copy number/g DNA in blood of grade 3 NE patients is higher than grade 2 NE patients, whereas there is no significant difference in peak value of CAR gene copy number/g DNA in blood of high or low grade CRS patients in this limited sample size. Median peak CAR gene copy number in blood/. Mu.g DNA was 5.16X10 in 16 patients who had not previously had bostemmaab 4 (range, 0-2.40X10) 5 ) And 6.15X10 among 8 patients previously having Bonauzumab 3 (range, 0-2.49X10) 5 )。
Peak levels of various key serum cytokines, chemokines and pro-inflammatory biomarkers occurred on day 7. Commensurate with peak CAR amplification, 1×10 6 Several serum analytes at 2X 10 compared to CAR T cells/kg 6 The tendency in the administered patients is higher (interleukin [ IL]-2, IL-5, IL-6, IL-8, IL-10, IL-15, IL-16, ferritin, granzyme B, intercellular adhesion molecule 1[ ICAM-1 ] ]Interferon gamma [ IFN-gamma ]]And tumor necrosis factor alpha [ TNF-alpha ]]。
Peak serum levels of analytes VCAM-1 and IL-16 correlated with grade 3 CRS. Such association was not observed in subjects with ≡3NE, probably due to the small number of patients with ≡3 NE. Product characteristics are similar at the dose level. The level of less differentiated CCR7+ T cells in the product is higher in patients with CR+CRi and tends to be higher in MRD-negative patients. This product profile also showed a trend to have higher levels of neurotoxicity, but was independent of CRS. The ratio of CD4 to CD 8T cells is independent of response or toxicity.
Discussion of the invention
In phase 1 of clinical trial-4, no DLT of KTE-X19 was observed in pediatric or adolescent patients with a DLT of R/R B-ALL that could be assessed. Although at 2X 10 6 No DLT was observed at the initial dose of individual CAR T cells/kg, but 1 x 10 with 68mL formulation was explored in the second cohort of patients 6 Lower doses of individual CAR T cells/kg in an attempt to further improve risk to benefit ratio, and in the third cohort with 1 x 10 with 40mL formulation 6 Individual CAR T cells/kg and modified toxicity management further optimize dose and toxicity management. This results in a 1×10 for 6 More optimal risk to benefit ratio for individual CAR T cell/kg (40 mL) dose levels, with significant improvement for CRS and NE. In addition, although the MRD negative rate of all the preparations was 73% or more, 1X 10 was accepted 6 Among patients with individual CAR T cells/kg (40 mL), MRD negative rate and CR rate alone were highest. Importantly, at 1X 10 6 Median DOR, RFS and OS was not achieved in 9 patients in the individual CAR T cells/kg (40 mL) cohort, with most responders (5/6 [83 ]]) Continuing with the subsequentalloSCT. The limitation of a small queue was recognized, however the 24 month OS ratio in this group was 87.5%. These results potentially demonstrate a meaningful response persistence using optimized dosages/formulations of KTE-X19 and subsequent alloSCT in pediatric/adolescent patients with R/R B-ALL.
In pediatric/adolescent patients with R/R B-ALL, the role of alloSCT following anti-CD 19 CAR T-cell therapy remains undefined; studies in the adult population provide somewhat conflicting results. In this study, at 1X 10 6 Median DOR and OS examined at the time of subsequent alloSCT was not reached in RP2D treated patients of individual CAR T cells/kg (40 mL). Fourteen (88%) (including 5 patients treated with RP 2D) of 16 patients who achieved cr+cri received alloSCT as follow-up therapy. alloSCT is not a protocol requirement, but is allowed by researchers as appropriate. Clinical trial-4 was not designed to evaluate the results after subsequent treatment; however, at the discretion of the investigator, most of the responding patients progressed to alloSCT after KTE-X19.
Evaluation of DOR in phase 4 clinical trials without review of subsequent treatments including alloSCT showed a favorable median of 14.2 months. In addition, the median RFS for the subsequent alloSCT examination was 5.2 months, but 9.1 months without examination. Pediatric and young adults with R/R CD19+ ALL who have no history of alloSCT but who receive consolidated alloSCT following anti-CD 19 CAR T-cell therapy have previously been reported to have a tendency to have no increased leukemia survival in a follow-up of > 1 year. In the recently disclosed phase 1 study of anti-CD 19 CAR T-cell therapy in children and young adults with R/R B-ALL in which 75% of MRD-negative response patients continued to perform alloSCT, the median OS at the 4.8 year follow-up was 70.2 months after alloSCT, 5 year event-free survival after alloSCT was 61.9%, and the cumulative incidence of relapse after alloSCT was only 9.5%. Interestingly, a retrospective review in pediatric and young adult patients found that CD34 selected T-cell depleted alloSCT following CAR T-cell therapy could result in improved graft-related mortality and OS compared to unmodified alloSCT.
The data provided herein support the promising potential role of KTE-X19 in prolonging response persistence and survival in pediatric/adolescent patients with R/R B-ALL, particularly if subsequently receiving alloSCT.
While differences in trial design and patient population precluded direct trial-trial comparisons, recent studies with bolaful mab (which also targets CD 19) demonstrated a median OS of only 7.7 months in pediatric R/R B-ALL, similar to the results in adult ALL. 40 also for bolamitraz, consolidation with subsequent alloSCT has shown improved results (12 month RFS rates of 70% versus 30% for patients with and without subsequent alloSCT, respectively). In addition, the remission rate of boscalid was higher in patients with lower baseline tumor burden (< 50% blast; 56% CR) compared to those with higher tumor burden (+.gtoreq.50% blast; 33% CR). In clinical trial-4, the clear correlation between remission rate and baseline bone marrow blast was not evident, as at baseline, CR rates in patients with >5 to ≡25%, >25 to ≡50%, >50 to ≡75% and >75 to 100% blast were 89%, 25%, 100% and 50%, respectively. This is consistent with another pediatric and young adult study using CAR T-cell therapy for CD19, with no differences in response rate based on disease burden. The data from clinical trial-4 demonstrate that KTE-X19 has the potential to provide more beneficial efficacy in patients with high disease burden than reported with boletlizumab. In clinical trial-4, a trend toward lower cr+cri rates appears to be observed in patients with prior bolafuximab.
The AE profile in clinical trial-4 was consistent with previous studies of anti-CD 19 CAR T-cell therapy. For patients receiving KTE-X19, the median time from leukocyte isolation to delivery to the study center was 16.5 days. In contrast, for the first 37 commercially manufactured tisagenlect results for patients with B-ALL, the reported average throughput time from receipt of the leukocyte isolation product to shipment to the clinical center was 23 days. The rapid turnaround time of the patients treated in clinical trial-4 supported the feasibility of setting up rapidly proliferating ALL.
The unmet medical need in R/R pediatric ALL is highest for patients with early relapse or primary refractory disease, with 5 year OS rates of 21% to 28%.2,4,8,43-45 furthermore, the risk of treatment-related morbidity and mortality is 3-to 5-fold higher in patients with MRD-positive disease at the end of the initial and later treatment lines than in patients with undetectable MRD.
3.
To address this ongoing unmet medical need, clinical trial-4 was further modified to expand the eligibility criteria to include patients with MRD positive disease and patients with early first relapse (. Ltoreq.18 months). In addition, a second cohort was opened for pediatric patients with R/R NHL (diffuse large B-cell lymphoma, burkitt's lymphoma, and primary mediastinal B-cell lymphoma). Among the agents approved by the U.S. food and drug administration/European drug administration or study agents in the pediatric R/R B-ALL registration trial, clinical trial 4 was the first CAR T cell therapy trial reporting positive results of follow-up for more than 3 years.
All publications, patents, patent applications, and other documents cited in this application are hereby incorporated by reference in their entirety for all purposes to the same extent as if each individual publication, patent application, or other document was individually indicated to be incorporated by reference for all purposes. While various specific embodiments/aspects have been shown and described, it will be appreciated that various changes can be made therein without departing from the spirit and scope of the disclosure.

Claims (21)

1. A method of treating recurrent/refractory B precursor acute lymphoblastic leukemia in a subject, the method comprising administering to the subject a therapeutically effective amount of immune cells against a tumor antigen, wherein the subject is a pediatric or adolescent subject.
2. The method of claim 1, wherein the therapeutically effective amount of the immune cells is between about 1 x 10 per kg body weight 6 And about 2X 10 6 Between individual immune cells.
3. The method of claim 2, wherein the immune cells are administered in a total volume of between about 40ml to 68 ml.
4. The method of claim 2, wherein the immune cells are administered in a total volume of about 40 ml.
5. The method of claim 1, wherein the therapeutically effective amount of the immune cells is about 1 x 10 per kg body weight 6 And (3) immune cells.
6. The method of claim 1, wherein the immune cells are administered as a first line, a second line, a third line, a fourth line, a fifth line, or a sixth line therapy, or prior to disease progression.
7. The method according to claim 1, wherein the tumor antigen is selected from the group consisting of tumor-associated surface antigen, 5T4, alpha Fetoprotein (AFP), B7-1 (CD 80), B7-2 (CD 86), BCMA, B-human chorionic gonadotrophin, CA-125, carcinoembryonic antigen (CEA), CD123, CD133, CD138, CD19, CD20, CD22, CD23, CD24, CD25, CD30, CD33, CD34, CD4, CD40, CD44, CD56, CD8, CLL-1, c-Met, CMV-specific antigen, CS-1, CSPG4, CTLA-4, DLL3, bisialoganglioside GD2, catheter epithelial mucin, EBV-specific antigen, EGFR variant III (EGFRvIII), ELF2M, endothelial glycoprotein, hepadin B2, epidermal Growth Factor Receptor (EGFR), epithelial cell adhesion molecule (EpCAM), epithelial tumor antigen, epF 2 (HER 2/neu), fibroblast-related protein (fap), FLT3, GD2, GD-3, GD-binding antigen, HEIL-3, HEIL-specific antigen, HIV-1, HIV-2, HIV-specific antigen, HIV-1, HIV-receptor, HIV-specific antigen; CD38, insulin growth factor (IGFl) -l, enterocarboxylesterase, kappa chain, LAGA-la, lambda chain, lasa-specific antigen, lectin-reactive AFP, lineage-specific or tissue-specific antigen such AS CD3, MAGE-A1, major Histocompatibility Complex (MHC) molecule, major Histocompatibility Complex (MHC) molecule presenting tumor-specific peptide epitopes, M-CSF, melanoma-associated antigen, mesothelin, MN-CA IX, MUC-1, mut hsp70-2, mutant p53, mutant ras, neutrophil elastase, NKG2D, nkp, NY-ESO-1, p53, PAP prostases, prostate Specific Antigen (PSA), prostate cancer tumor antigen-1 (PCTA-1), prostate specific antigen proteins, STEAP1, STEAP2, PSMA, RAGE-1, ROR1, RU2 (AS), surface adhesion molecules, survivin and telomerase, TAG-72, the Extra Domain A (EDA) and Extra Domain B (EDB) of fibronectin and the Al domain of tenascin-C (TnC Al), thyroglobulin, tumor matrix antigen, vascular endothelial growth factor receptor-2 (VEGFR 2), virus-specific surface antigens such AS HIV-specific antigens (such AS HIV gpl 20), and any derivatives or variants of these surface antigens.
8. The method of claim 7, wherein the target antigen is CD19.
9. The method of claim 8, further comprising preconditioning the subject with one or more preconditioning agents, wherein the one or more preconditioning agents are selected from at least one of an alkylating agent and a platinum-based agent, wherein the alkylating agent is selected from the group consisting of: melphalan, chlorambucil, cyclophosphamide, dichloromethyldiethylamine, nitrogen mustard (HN 2), uratemustine, uracil nitrogen mustard, melphalan, chlorambucil, ifosfamide, bendamustine, carmustine, lomustine, streptozotocin, alkyl sulfonates, busulfan, thiotepa or the like, and any combination thereof, and wherein the platinum-based preconditioning agent is selected from the group consisting of: platinum, cisplatin, carboplatin, nedaplatin, oxaliplatin, satraplatin, triplatinum tetranitrate, procarbazine, altretamine, triazene, dacarbazine, mitozolomide, temozolomide, dacarbazine, temozolomide, and any combination thereof.
10. The method of claim 9, wherein the preconditioning agent comprises cyclophosphamide and fludarabine.
11. The method of claim 9, wherein the cyclophosphamide is present at between 200mg/m 2 Day and 2000mg/m 2 Between/day, and wherein the fludarabine is administered at a dose of between 20mg/m 2 Day and 900mg/m 2 Dosage between/day.
12. The method of claim 9, wherein the administration of the one or more preconditioning agents begins at least seven days, at least six days, at least five days, at least four days, at least three days, at least two days, or at least one day prior to the administration of the immune cells.
13. The method of claim 1, wherein the subject has a high tumor burden.
14. The method of claim 1, further comprising at least one of the following steps: tozucchini is administered only in the case of cytokine release syndrome for management of neurological events, and corticosteroids are administered for management of grade 2 neurological events.
15. The method of claim 1, wherein the subject is at high risk for disease progression, wherein the subject is at high risk if the subject shows disease progression within 24 months after initial diagnosis.
16. The method of claim 1, wherein the immune cells are selected from Tumor Infiltrating Lymphocytes (TILs), NK cells, autologous T cells, allogeneic T cells, and engineered autologous T cells (eacts), and any combination thereof.
17. The method of claim 1, wherein the immune cell is a CAR T cell.
18. A method of treating cancer in a subject in need thereof, wherein the cancer is non-hodgkin lymphoma (NHL) or relapsed/refractory B-precursor acute lymphoblastic leukemia or relapsed/refractory B-cell non-hodgkin lymphoma (R/R B-ALL), the method comprising administering to the subject a therapeutically effective amount of immune cells that are anti-tumor antigens, and wherein the immune cells are autologous T cells that express the anti-CD 19 Chimeric Antigen Receptor (CAR).
19. The method of claim 18, wherein the cancer is NHL and the NHL is Mantle Cell Lymphoma (MCL) or Indolent NHL (iNHL).
20. The method of claim 19, wherein the iNHL is an edge zone lymphoma (MZL) or Follicular Lymphoma (FL).
21. The method of claim 19, wherein the cancer is NHL, and the NHL is diffuse large B-cell lymphoma (DLBCL) non-specific, primary mediastinum large B-cell lymphoma, high grade B-cell lymphoma, or DLBCL caused by follicular lymphoma.
CN202280034095.1A 2021-05-14 2022-05-12 Chimeric antigen receptor T cell therapies Pending CN117295507A (en)

Applications Claiming Priority (5)

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US63/188,916 2021-05-14
US63/248,941 2021-09-27
US202263328364P 2022-04-07 2022-04-07
US63/328,364 2022-04-07
PCT/US2022/029047 WO2022241151A2 (en) 2021-05-14 2022-05-12 Chimeric antigen receptor t cell therapy

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