BCMA-targeted CAR-T cell therapy for multiple myeloma

Abstract

The present application provides a method of treating a subject suffering from multiple myeloma with siltacabtadine autorucel suspension. The present application further provides a pharmaceutical product containing the siltacabtadine autorucel suspension, instructions for use of the siltacabtadine autorucel suspension, and a method for selling a pharmaceutical product containing the siltacabtadine autorucel suspension.

Description

CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims priority to U.S. Provisional Patent Application No. 63 / 314,968, filed February 28, 2022, the disclosure of which is incorporated by reference in its entirety herein.

[0002] Sequence Listing This application contains a computer readable sequence listing, previously submitted with this application in XML file format, the contents of which are incorporated herein by reference in their entirety. The sequence listing submitted with this application has an XML file title of "14651-054-228_SEQ_LISTING.xml", was created on February 23, 2023, and is 21,916 bytes in size. [Background technology]

[0003] Multiple myeloma is an aggressive plasma cell neoplasm. It is considered to be a B-cell neoplasm that grows uncontrollably in the bone marrow. Symptoms include one or more of hypercalcemia, renal failure, anemia, bone lesions, bacterial infection, hyperviscosity, and amyloidosis. Although available new therapies, including proteasome inhibitors, immunomodulatory drugs, and monoclonal antibodies, have significantly improved patient outcomes, multiple myeloma is still considered to be an incurable disease. Since most patients relapse or become incurable with treatment, there is a continuing need for new therapies for multiple myeloma. Summary of the Invention

[0004] In one aspect, a method of treating a subject suffering from multiple myeloma is provided, the method comprising administering to the subject, by a single intravenous infusion, a composition comprising T cells, the T cells comprising a chimeric antigen receptor (CAR), the chimeric antigen receptor comprising: a) an extracellular antigen binding domain comprising a first anti-BCMA binding moiety and a second BCMA binding moiety; b) a transmembrane domain; and c) an intracellular signaling domain; Thereby, a dose of T cells expressing a CAR (CAR-T cells) is delivered to the subject, wherein the first BCMA binding moiety comprises the amino acid sequence set forth in SEQ ID NO:2 and the second BCMA binding moiety comprises the amino acid sequence set forth in SEQ ID NO:4.

[0005] In some embodiments, the dose is 1.0×10 5 ~5.0×10 6 In some embodiments, the dose comprises 5.0×10 CAR-T cells / kilogram of the subject's mass. 5 ~1.0×10 6 CAR-T cells / kilogram of the subject's mass. In some embodiments, the dose is about 0.75×10 6 In some embodiments, the dose comprises 1.0×10 CAR-T cells per subject. 8 The CAR-T cell is preferably a cellular or a cellular component.

[0006] In some embodiments, the single intravenous infusion is administered with one bag of the CAR-T cells. In some embodiments, the administration of the one bag of the CAR-T cells is completed no later than 3 hours after thawing of the one bag of CAR-T cells.

[0007] In some embodiments, the single intravenous infusion is administered with two bags of the CAR-T cells. In some embodiments, the administration of each of the two bags of the CAR-T cells is completed no later than three hours after thawing each of the two bags of CAR-T cells.

[0008] In some embodiments, the method effectively achieves a negative minimal residual disease (MRD) status in the subject as assessed in bone marrow following the infusion of the CAR-T cells, hi some embodiments, the negative MRD status is achieved at a first follow-up period between about 28 days and about 179 days following the infusion of the CAR-T cells.

[0009] In some embodiments, a lymphodepleting regimen is administered prior to infusion of the CAR-T cells. In some embodiments, the lymphodepleting regimen includes administration of cyclophosphamide or administration of fludarabine. In some embodiments, the lymphodepleting regimen is administered intravenously. In some embodiments, the lymphodepleting regimen is administered 5 to 7 days prior to infusion of the CAR-T cells. In some embodiments, the lymphodepleting regimen includes administration of cyclophosphamide and fludarabine intravenously 5 to 7 days prior to infusion of the CAR-T cells. In some embodiments, the cyclophosphamide is administered at a dose of 300 mg / m 2 In some embodiments, the fludarabine is administered intravenously at a dose of 30 mg / m 2 It is administered intravenously.

[0010] In some embodiments, the method further comprises treating the subject for cytokine release syndrome (CRS) more than 3 days after infusion without significantly reducing in vivo expansion of CAR-T cells. In some embodiments, the CRS treatment comprises administering an IL-6R inhibitor to the subject. In some embodiments, the IL-6R inhibitor is an antibody. In some embodiments, the antibody inhibits IL-6R by binding to the extracellular domain of IL-6R. In some embodiments, the IL-6R inhibitor blocks binding of IL-6 to IL-6R. In some embodiments, the IL-6R inhibitor is tocilizumab.

[0011] In some embodiments, the subject is treated with an infusion prodrug comprising an antipyretic and an antihistamine at most 1 hour prior to the infusion comprising CAR-T cells. In some embodiments, the antipyretic comprises paracetamol or acetaminophen. In some embodiments, the antipyretic is administered orally or intravenously to the subject. In some embodiments, the antipyretic is administered to the subject at a dose of 650 mg to 1000 mg. In some embodiments, the antihistamine comprises diphenhydramine. In some embodiments, the antihistamine is administered to the subject at a dose of 25 mg to 50 mg, or the equivalent thereof.

[0012] In some embodiments, the infusion comprising the CAR-T cells further comprises an excipient selected from dimethylsulfoxide or dextran-40.

[0013] In some embodiments, the subject has received prior therapy including at least three prior lines of therapy, in some embodiments, the at least three prior lines of therapy include treatment with at least one agent, the at least one agent including at least one of a proteasome inhibitor (PI), an immunomodulatory agent (IMiD), or an anti-CD38 antibody.

[0014] In some embodiments, the subject relapses after said at least three prior lines of therapy. In some embodiments, the multiple myeloma is refractory to at least two drugs after said at least three prior lines of therapy. In some embodiments, the at least two drugs that are refractory to curing the subject include a PI and an IMiD. In some embodiments, the subject is refractory to at least three drugs. In some embodiments, the subject is refractory to at least four drugs. In some embodiments, the subject is refractory to at least five drugs.

[0015] In some embodiments, the method effectively obtains an overall response rate of greater than 91%. In some embodiments, the method effectively obtains an overall response rate of greater than 93%. In some embodiments, the method effectively obtains an overall response rate of greater than 95%. In some embodiments, the method effectively obtains an overall response rate of greater than 97%. In some embodiments, the method effectively obtains an overall response rate of greater than 99%. In some embodiments, the overall response rate is assessed at a median follow-up period of at least 12 months following the infusion of the CAR-T cells.

[0016] In some embodiments, the method is effective to obtain a median duration of initial response of less than 1.15 months. In some embodiments, the method is effective to obtain a median duration of initial response of less than 1.10 months. In some embodiments, the method is effective to obtain a median duration of initial response of less than 1.05 months. In some embodiments, the method is effective to obtain a median duration of initial response of less than 1.00 months. In some embodiments, the method is effective to obtain a median duration of initial response of less than 0.95 months.

[0017] In some embodiments, the method is effective to obtain a median duration of best response of less than 2.96 months. In some embodiments, the method is effective to obtain a median duration of best response of less than 2.86 months. In some embodiments, the method is effective to obtain a median duration of best response of less than 2.76 months. In some embodiments, the method is effective to obtain a median duration of best response of less than 2.66 months. In some embodiments, the method is effective to obtain a median duration of best response of less than 2.56 months.

[0018] In some embodiments, the method effectively obtains a negative minimal residual disease (MRD) status in the subject as assessed in bone marrow at a follow-up period of about 28 days or longer after the infusion of the CAR-T cells. In some embodiments, the method effectively maintains a negative MRD status in the subject as assessed in bone marrow at a follow-up period of about 12 months or longer after the infusion of the CAR-T cells.

[0019] In some embodiments, the first BCMA binding moiety and / or the second BCMA binding moiety is an anti-BCMA VHH. In some embodiments, the first BCMA binding moiety is a first anti-BCMA VHH and the second BCMA binding moiety is a second anti-BCMA VHH. In some embodiments, the first BCMA binding moiety comprises a polypeptide encoded by a nucleic acid sequence of SEQ ID NO: 10. In some embodiments, the second BCMA binding moiety comprises a polypeptide encoded by a nucleic acid sequence of SEQ ID NO: 12. In some embodiments, the first BCMA binding moiety and the second BCMA binding moiety are linked to each other via a peptide linker.

[0020] In some embodiments, the peptide linker comprises the amino acid sequence of SEQ ID NO: 3. In some embodiments, the peptide linker comprises a polypeptide encoded by the nucleic acid sequence of SEQ ID NO:11.

[0021] In some embodiments, the CAR polypeptide further comprises a signal peptide located at the N-terminus of the polypeptide. In some embodiments, the signal peptide is derived from CD8α. In some embodiments, the signal peptide comprises the amino acid sequence of SEQ ID NO: 1. In some embodiments, the signal peptide comprises a polypeptide encoded by the nucleic acid sequence of SEQ ID NO: 9.

[0022] In some embodiments, the transmembrane domain comprises the amino acid sequence of SEQ ID NO: 6. In some embodiments, the transmembrane domain comprises a polypeptide encoded by a nucleic acid sequence of SEQ ID NO: 14.

[0023] In some embodiments, the intracellular signaling domain comprises a primary intracellular signaling domain of an immune effector cell. In some embodiments, the intracellular signaling domain is derived from CD3zeta. In some embodiments, the intracellular signaling domain comprises one or more costimulatory signaling domains. In some embodiments, the intracellular signaling domain comprises the amino acid sequence of SEQ ID NO:8. In some embodiments, the intracellular signaling domain comprises a polypeptide encoded by a nucleic acid sequence of SEQ ID NO:16. In some embodiments, the intracellular signaling domain comprises the amino acid sequence of SEQ ID NO:7. In some embodiments, the intracellular signaling domain comprises a polypeptide encoded by a nucleic acid sequence of SEQ ID NO:15.

[0024] In some embodiments, the CAR polypeptide further comprises a hinge domain located between the C-terminus of the extracellular antigen binding domain and the N-terminus of the transmembrane domain. In some embodiments, the hinge domain comprises the amino acid sequence of SEQ ID NO: 5. In some embodiments, the hinge domain comprises a polypeptide encoded by the nucleic acid sequence of SEQ ID NO: 13.

[0025] In some embodiments, the T cells are autologous T cells. In some embodiments, the T cells are allogeneic T cells.

[0026] In some embodiments, the subject is a human.

[0027] In one aspect, a method of treating a subject suffering from multiple myeloma and who has received at least three prior lines of therapy is provided, the method comprising administering to the subject, by a single intravenous infusion, a composition comprising T cells, the T cells comprising a chimeric antigen receptor (CAR), the chimeric antigen receptor comprising the amino acid sequence of SEQ ID NO: 17, about 0.75×10 6 A dose of 100 T cells (CAR-T cells) / kilogram of subject's mass is delivered to a subject, wherein the method effectively obtains a minimal residual disease (MRD) negativity status in the subject, wherein the minimal residual disease negativity status is assessed in bone marrow at a follow-up period of 28 days or more following the infusion of the CAR-T cells.

[0028] In some embodiments, the method includes administering to the patient a 10 to 30-week follow-up period of about 12 months after the infusion of the CAR-T cells. -5 At a sensitivity threshold level of about 44% to about 65% of patients were treated with CAR-T cells for a follow-up period of about 18 months after the infusion of the CAR-T cells. -4 At a sensitivity threshold level of about 57% to about 76% of patients were treated with CAR-T cells for a follow-up period of about 18 months after the infusion of the CAR-T cells. -5 or at a follow-up period of about 18 months after said infusion of said CAR-T cells, -6 At a sensitivity threshold level of about 29% to about 50% of cases effectively achieve the MRD negativity state.

[0029] In some embodiments, the method includes administering the CAR-T cells to a patient in need of treatment for a follow-up period of about 12 months. -5 At a sensitivity threshold level of 100 mg / kg / day, about 55% of patients had a 10% or higher CAR-T cell survival rate at about 18 months of follow-up after the infusion of the CAR-T cells. -4 At a sensitivity threshold level of 100 mg / kg / day, approximately 67% of patients had a 10% or higher CAR-T cell survival rate at a follow-up period of approximately 18 months after the infusion of the CAR-T cells. -5 At a sensitivity threshold level of about 10%, or at a follow-up period of about 18 months after the infusion of the CAR-T cells, -6 At a sensitivity threshold level of 0.1, the MRD negativity is effectively achieved in approximately 39% of cases.

[0030] In some embodiments, the method includes administering to the patient a 10 to 30-week follow-up period of about 12 months after the infusion of the CAR-T cells. -5 At a sensitivity threshold level of about 10 to about 83% in subjects with evaluable samples, or at a follow-up period of about 18 months after the infusion of the CAR-T cells, -5 At a sensitivity threshold level of 0.1, the minimal residual disease (MRD) negativity status is effectively obtained in about 82% to about 97% of subjects with evaluable samples.

[0031] In some embodiments, the method includes administering to the patient a 10 to 30-week follow-up period of about 12 months after the infusion of the CAR-T cells. -5 At a sensitivity threshold level of 100%, in subjects with evaluable samples, a rate of about 93%, or at a follow-up period of about 18 months after the infusion of the CAR-T cells, -5 At this sensitivity threshold level, approximately a 92% rate of subjects with evaluable samples effectively achieves the minimal residual disease (MRD) negative status.

[0032] In some embodiments, the method is effective to obtain at least one response in the subject following said infusion of the CAR-T cells, wherein said at least one response comprises, in order from best to worst, a stringent complete response, a complete response, a best partial response, a partial response, or a minimal response.

[0033] In some embodiments, the method is effective to obtain an initial response before a period of about 27 days to about 321 days after the infusion of the CAR-T cells. In some embodiments, the method is effective to obtain an initial response before a period of about 27 days to about 89 days after the infusion of the CAR-T cells. In some embodiments, the method is effective to obtain an initial response before about 42 days after the infusion of the CAR-T cells. In some embodiments, the method is effective to obtain an initial response before about 29 days after the infusion of the CAR-T cells.

[0034] In some embodiments, the method is effective to obtain a best response of any one of minimal response, partial response, best partial response, complete response, or stringent complete response. In some embodiments, the method is effective to obtain a best response of any one of minimal response, partial response, best partial response, complete response, or stringent complete response at a rate of about 91% to about 99% at about 12 months follow-up period after the infusion of the CAR-T cells, or at a rate of about 93% to about 100% at about 18 months follow-up period after the infusion of the CAR-T cells. In some embodiments, the method is effective to obtain a best response of any one of minimal response, partial response, best partial response, complete response, or stringent complete response at a rate of about 97% at about 12 months follow-up period after the infusion of the CAR-T cells, or at a rate of about 98% at about 18 months follow-up period after the infusion of the CAR-T cells.

[0035] In some embodiments, the method is effective to obtain a best response of any one of partial response, best partial response, complete response, or stringent complete response. In some embodiments, the method is effective to obtain a best response of any one of partial response, best partial response, complete response, or stringent complete response at a rate of about 91% to about 99% at about 12 months follow-up period after the infusion of the CAR-T cells, or at a rate of about 93% to about 100% at about 18 months follow-up period after the infusion of the CAR-T cells. In some embodiments, the method is effective to obtain a best response of any one of partial response, best partial response, complete response, or stringent complete response at a rate of about 97% at about 12 months follow-up period after the infusion of the CAR-T cells, or at a rate of about 97% at about 18 months follow-up period after the infusion of the CAR-T cells.

[0036] In some embodiments, the method is effective to obtain a best response of any one of a best partial response, a complete response, or a stringent complete response. In some embodiments, the method is effective to obtain a best response of any one of a best partial response, a complete response, or a stringent complete response at a rate of about 86% to about 97% at about 12 months follow-up after the infusion of the CAR-T cells, or at a rate of about 88% to about 98% at about 18 months follow-up after the infusion of the CAR-T cells. In some embodiments, the method is effective to obtain a best response of any one of a best partial response, a complete response, or a stringent complete response at a rate of about 93% at about 12 months follow-up after the infusion of the CAR-T cells, or at a rate of about 95% at about 18 months follow-up after the infusion of the CAR-T cells.

[0037] In some embodiments, the method is effective to obtain a best response of complete response or stringent complete response. In some embodiments, the method is effective to obtain the best response of complete response or stringent complete response at a rate of about 57% to about 76% at about 12 months follow-up period after the infusion of the CAR-T cells, or at a rate of about 73% to about 89% at about 18 months follow-up period after the infusion of the CAR-T cells. In some embodiments, the method is effective to obtain the best response of complete response or stringent complete response at a rate of about 67% at about 12 months follow-up period after the infusion of the CAR-T cells, or at a rate of about 83% at about 18 months follow-up period after the infusion of the CAR-T cells.

[0038] In some embodiments, the method is effective to obtain a best response of stringent complete response. In some embodiments, the method is effective to obtain the best response of stringent complete response at a rate of about 57% to about 76% at about 12 months follow-up period after the infusion of the CAR-T cells, or at a rate of about 73% to about 89% at about 18 months follow-up period after the infusion of the CAR-T cells. In some embodiments, the method is effective to obtain the best response of stringent complete response at a rate of about 67% at about 12 months follow-up period after the infusion of the CAR-T cells, or at a rate of about 83% at about 18 months follow-up period after the infusion of the CAR-T cells.

[0039] In some embodiments, the method effectively obtains progression free survival in a subject. In some embodiments, the method effectively obtains progression free survival in a subject from the infusion of the CAR-T cells to about 209 days after the infusion of the CAR-T cells, to about 386 days after the infusion of the CAR-T cells, to about 632 days after the infusion of the CAR-T cells, or to about 684 days after the infusion of the CAR-T cells.

[0040] In some embodiments, the method effectively obtains a progression free survival rate of about 79% to about 93% at about a 6 month follow-up period after the infusion of the CAR-T cells, about 67% to about 84% at about a 12 month follow-up period after the infusion of the CAR-T cells, about 57% to about 75% at about an 18 month follow-up period after the infusion of the CAR-T cells, about 57% to about 75% at about a 21 month follow-up period after the infusion of the CAR-T cells, or about 49% to about 70% at about a 24 month follow-up period after the infusion of the CAR-T cells.

[0041] In some examples, the method effectively obtains the progression free survival at about an 88% rate at about a 6 month follow-up period after the infusion of the CAR-T cells, at about a 76% rate at about a 12 month follow-up period after the infusion of the CAR-T cells, at about a 67% rate at about an 18 month follow-up period after the infusion of the CAR-T cells, at about a 67% rate at about a 21 month follow-up period after the infusion of the CAR-T cells, or at about a 61% rate at about a 24 month follow-up period after the infusion of the CAR-T cells.

[0042] In some embodiments, the method further comprises treating cytokine release syndrome in the subject more than about 1 day after said infusion of said CAR-T cells, hi some embodiments, the method effectively obtains about a 1% to about 99% reversal rate of said cytokine release syndrome within a time period of about 1, 3, 4, 6, 16, or 97 days after said cytokine release syndrome is first observed.

[0043] In some embodiments, the method further comprises treating immune effector cell associated neurotoxicity in the subject more than about 3 days after the infusion of the CAR-T cells. In some embodiments, the method effectively obtains about a 1% to about 17% reversal rate of immune effector cell associated neurotoxicity in a time period of about 1, 4, 5, 8, 12, or 16 days after first observing immune effector cell associated neurotoxicity.

[0044] In some embodiments, the method is effective to obtain the best response before about 27 days to about 534 days after the infusion of the CAR-T cells. In some embodiments, the method is effective to obtain the best response before about 27 days to about 293 days after the infusion of the CAR-T cells. In some embodiments, the method is effective to obtain the best response before about 153 days after the infusion of the CAR-T cells. In some embodiments, the method is effective to obtain the best response before about 78 days after the infusion of the CAR-T cells.

[0045] In some examples, the method effectively maintains the response in the subject for a follow-up period from the time of the initial response to about 180 days after the infusion of the CAR-T cells, about 357 days after the infusion of the CAR-T cells, about 606 days after the infusion of the CAR-T cells, or about 654 days after the infusion of the CAR-T cells. In some examples, the methods effectively maintain a response rate of about 77% to about 91% at about a 6 month follow-up period after the infusion of the CAR-T cells, about 63% to about 81% at about a 12 month follow-up period after the infusion of the CAR-T cells, about 56% to about 75% at about an 18 month follow-up period after the infusion of the CAR-T cells, about 52% to about 72% at about a 21 month follow-up period after the infusion of the CAR-T cells, or about 48% to about 70% at about a 24 month follow-up period after the infusion of the CAR-T cells. In some examples, the methods effectively maintain response at about an 85% rate at about a 6 month follow-up period after the infusion of the CAR-T cells, at about a 74% rate at about a 12 month follow-up period after the infusion of the CAR-T cells, at about a 67% rate at about an 18 month follow-up period after the infusion of the CAR-T cells, at about a 63% rate at about a 21 month follow-up period after the infusion of the CAR-T cells, or at about a 60% rate at about a 24 month follow-up period after the infusion of the CAR-T cells.

[0046] In some embodiments, the method includes administering the CAR-T cells to a patient having a CAR-T cell death within 10 days from the time of administration to about 3 months after administration of the CAR-T cells. -5In some embodiments, the method effectively obtains a minimal residual disease (MRD) negative status in the subject as assessed in bone marrow at a sensitivity threshold level of about 25% to about 44% at about 12 months follow-up period after the infusion of the CAR-T cells, or about 33% to about 54% at about 18 months follow-up period after the infusion of the CAR-T cells. In some embodiments, the method effectively obtains a minimal residual disease (MRD) negative complete response or a minimal residual disease (MRD) negative stringent complete response at about 34% at about 12 months follow-up period after the infusion of the CAR-T cells, or about 43% at about 18 months follow-up period after the infusion of the CAR-T cells.

[0047] The present disclosure further relates to a pharmaceutical product comprising siltacabtadine autorucel suspension for intravenous infusion, wherein the pharmaceutical product is packaged, and wherein the package comprises a label, the label identifying the siltacabtadine autorucel suspension as an approved pharmaceutical product for treating adult patients with relapsed or refractory multiple myeloma after receiving four or more prior lines of therapy, the prior lines of therapy including a proteasome inhibitor, an immunomodulatory agent, and an anti-CD38 monoclonal antibody.

[0048] The present disclosure provides a method of treating relapsed or refractory multiple myeloma in an adult patient in need thereof, comprising administering an approved pharmaceutical product comprising siltacabtadine autorucel suspension in an amount and manner as described in the pharmaceutical product label of the approved pharmaceutical product and / or in a therapeutic treatment regimen described herein.

[0049] The present disclosure further provides a method of marketing an approved pharmaceutical product comprising siltacabtadine autorucel suspension, said method comprising marketing such pharmaceutical product, wherein the pharmaceutical product label of a reference product for such pharmaceutical product comprises instructions for treating patients with relapsed or refractory multiple myeloma after receiving four or more prior lines of therapy, said prior lines of therapy comprising a proteasome inhibitor, an immunomodulatory agent, and an anti-CD38 monoclonal antibody.

[0050] The present disclosure further provides a method of authorizing the sale of a pharmaceutical product comprising siltacabtadine autorucel suspension, the method comprising authorizing the sale of such pharmaceutical product, wherein the pharmaceutical product label of a reference product for such pharmaceutical product comprises instructions for treating patients with relapsed or refractory multiple myeloma after receiving four or more prior lines of therapy, the prior lines of therapy comprising a proteasome inhibitor, an immunomodulatory agent, and an anti-CD38 monoclonal antibody. [Brief description of the drawings]

[0051] [Figure 1] Figure 1 shows expression of BCMA antigen on the surface of GC, memory and plasmablast cells in lymph nodes, long-lived plasma cells in bone marrow LN and MALT, and multiple myeloma cells. BAFF-R antigen is not expressed on plasmablasts, long-lived plasma cells, or multiple myeloma cells. TACI is expressed on memory and plasmablast cells, long-lived plasma cells, and multiple myeloma cells. CD138 is expressed only on long-lived plasma cells and multiple myeloma cells.

[0052] [Diagram 2] Figure 1 shows the design of the siltacabtadine autolucel CAR. The siltacabtadine autolucel contains two VHH domains, which differs from the single VL and single VH domains found in various other CARs. The siltacabtadine autolucel contains intracellular CD137 and the human CD3 ζ domain.

[0053] [Diagram 3]FIG. 1 is a schematic diagram of producing a virus encoding a siltacabtadine autolucel CAR, transducing the virus into a patient's T cells, and producing CAR T cells expressing siltacabtadine autolucel.

[0054] [Figure 4] Schematic diagram showing the study design of siltacabtadine autolucel CAR T cells. The patient population includes patients with relapsed or refractory multiple myeloma, incurable in both three previous lines or PI / IMiD, and previously exposed to PI, IMiD and anti-CD38. The primary goal is to establish safety and RP2D, e.g., investigating incidence and severity of adverse events (Stage 1b). Another primary goal is efficacy; ORR-PR or better, as defined by the IMWG (Stage 2). The following are secondary goals: incidence and severity of adverse events (Stage 2) and any further efficacy characterization.

[0055] [Figure 5A] Graph showing response and duration of response (DOR) in the 50 responders with the most follow-up in the all treatment analysis population, as assessed by an Independent Review Committee (IRC), with a median follow-up of 12.4 months, sorted by responder follow-up duration along the vertical axis. [Figure 5B] Graph showing response and duration of response (DOR) in the 44 responders with the shortest follow-up in the all treatment analysis population, sorted by responder follow-up duration along the vertical axis, based on Independent Review Committee (IRC) assessment, with a median follow-up of 12.4 months.

[0056] [Figure 6] Kaplan-Meier plot of duration of response (DOR) of responders in the all-treatment analysis population based on independent review committee (IRC) assessment with a median follow-up of 12.4 months, which showed that responders had approximately 80.2% and 68.2% probability of maintaining response at 9 and 12 months, respectively.

[0057] [Figure 7] Kaplan-Meier plot of overall survival (OS) of all subjects in the all-treatment analysis population with a median follow-up of 12.4 months, which showed that the 9- and 12-month survival rates were approximately 90.7% and 88.5%, respectively.

[0058] [Figure 8] , [Figure 9] , [Figure 10] , [Figure 11] , [Figure 12] , [Figure 13] , [Figure 14] and [Figure 15] Descriptive diagram of the regimen and data obtained from the stage 1b-2 clinical trial described herein, with a median follow-up of 12.4 months, in which patients suffering from relapsed, refractory multiple myeloma were treated with siltacabtadine autorucel.

[0059] [Figure 16] , [Figure 17] , [Figure 18] and [Figure 19] FIG. 1 depicts a description of the assessment and evaluation of cytokine release syndrome (CRS) in the stage 1b-2 clinical trial described herein, in which patients suffering from relapsed, refractory multiple myeloma were treated with siltacabtadine autorucel.

[0060] [Figure 20] Graph showing response and duration of response (DOR) in responders in the all treatment analysis population based on independent review committee (IRC) assessment with a median follow-up of 18 months.

[0061] [Figure 21] Kaplan-Meier plot of duration of response (DOR) of responders in the treated analysis population based on independent review committee (IRC) assessment with a median follow-up of 18 months.

[0062] [Figure 22] Kaplan-Meier plot of progression-free survival (PFS) of responders in the proprietary treatment analysis population based on independent review committee (IRC) assessment with a median follow-up of 18 months.

[0063] [Diagram 23] FIG. 1 is a Kaplan-Meier plot of overall survival (OS) for all subjects in the all treatment analysis population with a median follow-up of 18 months.

[0064] [Figure 24A] , [Figure 24B] and [Figure 24C] Forest plot showing subgroup analysis of overall response rate (ORR) in the all treatment analysis population based on independent review committee (IRC) assessment with a median follow-up of 18 months.

[0065] [Figure 25A] , [Figure 25B] and [Figure 25C] Forest plot showing subgroup analysis of duration of response (DOR) in the all treatment analysis population based on independent review committee (IRC) assessment with a median follow-up of 18 months.

[0066] [Figure 26A] , [Figure 26B] and [Figure 26C] Forest plot showing subgroup analysis of 10-5 overall MRD negativity rate in bone marrow for the all treatment analysis population with a median follow-up of 18 months.

[0067] [Figure 27A] , [Figure 27B] and [Figure 27C] 1 is a forest plot showing subgroup analysis of 10-5 overall MRD negativity rate in bone marrow for subjects with evaluable samples in the all treatment analysis population with a median follow-up of 18 months.

[0068] [Figure 28] FIG. 1 summarizes the selection of comparison groups for indirect treatment comparison (ITC) analyses.

[0069] [Figure 29A] and [Figure 29B] FIG. 1 shows a meta-analysis comparison performed on overall survival for patients treated with siltacabtadine autorucel and those treated by physician's choice of treatment using all index dates.

[0070] [Diagram 30] FIG. 1 shows a meta-analysis comparison performed on progression-free survival for patients treated with siltacabtadine autorucel and those treated by physician's choice of treatment using all index dates.

[0071] [Diagram 31] FIG. 1 shows a meta-analysis comparison performed on overall survival of patients treated with siltacabtadine autorucel and patients treated by physician's choice of treatment using the first index date.

[0072] [Diagram 32] FIG. 1 shows a meta-analysis comparison performed on progression-free survival for patients treated with siltacabtadine autorucel and those treated by physician's choice of treatment using the first index date. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0073] The present disclosure further provides related nucleic acids, recombinant expression vectors, host cells, cell populations, antibodies or antigen-binding portions thereof, and pharmaceutical compositions relating to the immune cells and CAR-expressing T cells of the disclosure. Dosage regimens, dosage forms, and methods of treatment with CAR-T cells are further provided.

[0074] In the following, some aspects of the present disclosure will be described with reference to examples for illustrative purposes only. It should be understood that many specific details, relationships and methods are provided to provide a thorough understanding of the present disclosure. However, as one skilled in the art can easily recognize, the present disclosure can be practiced without one or more of the specific details, or can be practiced using other methods, regimens, reagents, cell lines and animals. The present disclosure is not limited to the order of acts or events shown, since some acts may occur in different orders and / or simultaneously with other acts or events. It should be noted that not all acts, steps or events shown are required to achieve a method according to the present disclosure.

[0075] Unless otherwise defined, all technical terms, symbols and other scientific or technical terms used herein are intended to have the meanings commonly understood by those skilled in the art. In some cases, for clarity and / or ease of reference, the present specification defines terms having commonly understood meanings, and the specification including such definitions should not necessarily be interpreted as indicating a significant difference from the meanings commonly understood in the art. It is further understood that terms (as defined in a general dictionary) should be interpreted as having a meaning consistent with their meaning in the context of the relevant field and / or as otherwise defined in the present specification.

[0076] The term "about" or "approximately" includes within a statistically significant range of values. Such ranges may be within the order of a given value or range, preferably within 50%, more preferably within 20%, even more preferably within 10%, and even more preferably within 5%. The acceptable variation encompassed by the term "about" or "approximately" will depend on the particular system under study and can be readily appreciated by one of ordinary skill in the art.

[0077] The term "antibody" includes monoclonal antibodies (including full-length four-chain antibodies with an immunoglobulin Fc region or full-length heavy chain-only antibodies), antibody compositions with multi-epitopic specificity, multispecific antibodies (e.g., bispecific antibodies, diabbodies and single chain molecules), and antibody fragments (e.g., Fab, F(ab')2 and Fv). The terms "immunoglobulin" (Ig) and "antibody" are used interchangeably herein. Antibodies contemplated herein include single domain antibodies, e.g., heavy chain only antibodies.

[0078] The term "heavy chain-only antibody" or "HCAb" includes functional antibodies, which contain heavy chains but lack the light chains normally found in four-chain antibodies. Camelids (e.g., camels, llamas, alpacas) are known to produce HCAbs.

[0079] The term "single domain antibody" or "sdAb" refers to a single antigen-binding polypeptide having three complementarity determining regions (CDRs). A single sdAb can bind to an antigen but does not pair with a corresponding CDR-containing polypeptide. In some cases, single domain antibodies are engineered from camelid HCAbs and the heavy chain variable domain is referred to herein as a "VHH". Some VHHs may also be referred to as "nanobodies". Camlidid sdAbs are among the smallest antigen-binding antibody fragments known (see, e.g., Hamers-Casterman et al., Nature 363:446-8 (1993); Greenberg et al., Nature 374:168-73 (1995); Hassanzadeh-Ghassabeh et al., Nanomedicine (Lond), 8:1013-26 (2013)). A basic VHH has the structure, from N-terminus to C-terminus, FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4, where FR1 to FR4 refer to framework regions 1 to 4, respectively, and where CDR1 to CDR3 refer to complementarity determining regions 1 to 3.

[0080] The "variable region" or "variable domain" of an antibody refers to the amino-terminal domain of the heavy or light chain of the antibody. The heavy and light chain variable domains may be referred to as "VH" and "VL", respectively. These domains are usually the most variable parts of the antibody (compared to other antibodies of the same class) and contain the antigen binding site. Heavy chain-only antibodies from camelid species have a single heavy chain variable region called a "VHH" domain. Thus, VHH is a special type of variable region.

[0081] The term "variable" refers to the fact that the sequences of some segments of the variable domains vary widely between antibodies. The V domains (i.e., variable domains) mediate antigen binding and define the specificity of a particular antibody for its particular antigen. However, variability is not uniformly distributed over the entire span of the variable domains. Instead, it is concentrated in three segments called hypervariable regions (HVRs) in the light and heavy chain variable domains. The more conserved parts of the variable domains are called framework regions (FRs). Natural heavy and light chain variable domains each contain four FR regions that mainly adopt a β-sheet structure, which are connected via three HVRs that connect to form loops that connect the β-sheet structure and in some cases form part of the β-sheet structure. The HVRs in each chain are tightly bound by the FR regions and favor the formation of the antigen-binding site of the antibody (together with the HVRs from another chain if the antibody is not an sdAb or HCAb) (see Kabat et al., Sequences of Immunological Interest, Fifth Edition, National Institute of Health, Bethesda, Md. (1991)). The constant domains are not directly involved in binding the antibody to an antigen, but exhibit various effector functions, such as the participation of the antibody in antibody-dependent cellular cytotoxicity.

[0082] The terms "fragment of an antibody," "antibody fragment," "functional fragment of an antibody," and "antigen-binding portion" are used interchangeably herein and refer to one or more fragments or portions of an antibody, which fragment or portion retains the ability to specifically bind to an antigen (see generally, Holliger et al., Nat. Biotech., 23(9): 1 126-1129 (2005)). The antigen recognition portion of a CAR encoded by a nucleic acid sequence disclosed herein may contain any BCMA-binding antibody fragment. The antibody fragment desirably includes, for example, one or more CDRs, a variable region (or a portion thereof), a constant region (or a portion thereof), or a combination thereof. Examples of antibody fragments include (i) a Fab fragment, which is a monovalent fragment consisting of the VL, VH, CL, and CHI domains; (ii) a F(ab')2 fragment, which is a bivalent fragment comprising two Fab fragments linked by a disulfide bond at the hinge region; (iii) an Fv fragment consisting of the VL and VH domains of a single arm of an antibody; and (iv) a single-chain Fv (scFv), which is a monovalent molecule consisting of two domains (i.e., VL and VH) of an Fv fragment linked via a synthetic linker, which allows the synthesis of the two domains into a single polypeptide chain (see, for example, Bird et al., Science, 242: 423-426 (1988); Huston et al., Proc. Natl. Acad. Sci. USA, 85: 5879-5883 (1988); and Osbourn et al., Nat. Biotechnol, 16: 778-789 (1989)). (see, e.g., U.S. Pat. No. 6,399,411; U.S. Pat. No. 6,399,425; and (v) diabody fragments that are dimers of polypeptide chains, each polypeptide chain comprising a VH linked to a VL via a peptide linker that is too short to allow pairing between the VH and VL on the same polypeptide chain, thereby driving pairing of complementary domains on different VH-VL polypeptide chains to produce a dimeric molecule having two functional antigen-binding sites. Antibody fragments are known in the art and are described in more detail, for example, in U.S. Patent Application Publication No. 2009 / 0093024 A1.

[0083] As used herein, the terms "specific binding," "specific recognition," or "having specificity for" refer to a measurable and reproducible interaction, such as the binding of a target to an antigen binding protein (e.g., a CAR or a VHH), that determines the presence of the target in the presence of a heteromolecular population that includes a biomolecule.

[0084] The term "specificity" refers to the selective recognition of a particular epitope of an antigen by an antigen binding protein (e.g., a CAR or a VHH). For example, natural antibodies are monospecific.

[0085] A "chimeric antigen receptor" or "CAR" is an artificially constructed hybrid protein or polypeptide that contains an antigen-binding domain of an antibody (or antibody fragment) linked to a T cell signaling domain. CAR features may include the ability to redirect T cell specificity and reactivity to a selected target in a non-MHC-restricted manner (utilizing the antigen-binding properties of monoclonal antibodies). Non-MHC-restricted antigen recognition endows CAR-expressing T cells with the ability to recognize antigens (not dependent on antigen processing), thus circumventing a major mechanism of tumor escape. Advantageously, when expressed in T cells, CARs do not dimerize with endogenous T cell receptor (TCR) α and β chains. T cells expressing CARs are referred to herein as CAR T cells, CAR-T cells, or CAR-modified T cells, and these terms are used interchangeably herein. The cells can be genetically modified to stably express an antibody-binding domain on their surface to confer novel MHC-independent antigen specificity. "BCMA CAR" refers to a CAR that has an extracellular binding domain specific for BCMA. "Dual epitope CAR" refers to a CAR that has an extracellular binding domain specific for two different epitopes of BCMA.

[0086] "Cilta-cel" ("cilta-cel") is a chimeric antigen receptor T cell (CAR-T) therapy that contains two B cell maturation antigen (BCMA)-targeting VHH domains designed to confer avidity for BCMA. Cilta-cel may comprise T lymphocytes transduced with a cilta-cel CAR (a CAR encoded by a lentiviral vector). The CAR targets the human B cell maturation antigen (BCMA CAR). A schematic of the lentiviral vector encoding the cilta-cel CAR is shown in FIG. 2. The amino acid sequence of the cilta-cel CAR is that of SEQ ID NO: 17.

[0087] The terms "express" and "expression" mean to permit or cause the production of the information in a gene or DNA sequence. For example, expression can take the form of producing a protein by activating cellular functions involved in the transcription and translation of the corresponding gene or DNA sequence. A DNA sequence forms an "expression product," such as a protein, in a cell or by cellular expression. The expression product itself, e.g., the resulting protein, can also be referred to as being "expressed" by the cell. Expression products may be characterized as intracellular, extracellular, or transmembrane.

[0088] The term "treat" refers to a therapeutic treatment, where the objective is to alleviate or relieve an undesirable physiological change or disease, or to provide a beneficial or desired clinical outcome during treatment. Beneficial or desired clinical outcomes include alleviation of symptoms, reduction in the extent of disease, stabilization of the disease state (i.e., not worsening), delay or alleviation of disease progression, improvement or alleviation of the disease state, and / or remission (whether partial or complete remission), whether detectable or undetectable. "Treatment" may also mean extending survival compared to the expected survival if the subject were not receiving treatment. Subjects in need of treatment include subjects already suffering from an undesirable physiological change or disease, and subjects who are prone to suffering from a physiological change or disease. Treatment may also refer to a therapeutic agent, referred to herein as a "medicament" or "medication," which, by its action, may be intended to aid in the realization of a beneficial or desired clinical outcome of interest. The therapeutic agent or drug may be administered to the subject via a number of routes, including at least intravenous and oral routes. The term "intravenous" in relation to administration of a therapeutic agent or drug refers to administering said therapeutic agent or drug into one or more veins. The term "oral" in relation to administration of a therapeutic agent or drug refers to administering said therapeutic agent or drug via an oral passageway, such as the mouth.

[0089] As used herein, the term "subject" refers to an animal. The terms "subject" and "patient" are used interchangeably herein when referring to a subject. Thus, "subject" refers to a person being treated for a disease or being prevented from a disease as a patient. The methods described herein may be used to treat animal subjects belonging to any class. Examples of such animals include mammals. Mammals include, but are not limited to, rodent (order Rodentia) mammals, such as mice and hamsters, and lagomorph (order Logomorpha) mammals, such as rabbits. Mammals may be of the order Carnivora, including felines (cats) and canines (dogs). Mammals may be of the order Artiodactyla, including bovidae (cattle) and porcinae (pigs), or of the order Perissodactyla, including equidae (horses). The mammal may be of the order Primate, Ceboid, or Simoid (monkeys) or the order Anthropoid (humans and monkeys). In some embodiments, the mammal is a human.

[0090] The term "effective" as applied to a dose or amount refers to an amount of a compound or pharmaceutical composition sufficient to produce a desired activity after administration to a subject in need thereof. It should be noted that when a combination of active ingredients is administered, the effective amount of the combination may or may not include the amount of each ingredient that is effective when administered individually. The exact amount required will vary from subject to subject, depending on the species, age and general condition of the subject, the severity of the condition being treated, the particular drug or drugs used, the pattern of administration, etc.

[0091] The phrase "pharmacologically acceptable" as used in conjunction with the compositions described herein refers to molecular entities and other components of the composition that are physiologically tolerable and typically do not produce adverse reactions when administered to a mammal (e.g., a human). Preferably, the term "pharmacologically acceptable" means approved by a regulatory agency of the Federal or State government or listed in the United States Pharmacopeia or other generally recognized pharmacopoeias for use in mammals, more specifically, humans.

[0092] The term "line of treatment" used in conjunction with the method of treatment herein refers to one or more cycles of a planned treatment procedure, which may consist of one or more planned cycles of single agent or combination therapy and the treatment sequence administered in a planned manner. For example, a planned treatment regimen of induction therapy followed by autologous stem cell transplantation followed by maintenance is one line of treatment. A new line of treatment is considered to have been started if the planned course of treatment is amended to include other therapeutic agents or drugs (alone or in combination) due to disease progression, recurrence or toxicity. A new line of treatment is also considered to have been started if the observation period after the planned therapy is interrupted due to the need to administer additional treatment to the disease.

[0093] The term "refractory" as used herein in conjunction with treatment with a particular therapeutic agent or drug refers to a disease or subject with a disease that does not respond to said therapeutic agent or drug. The phrase "refractory myeloma" refers to a disease that does not respond to primary or rescue therapy or progresses within 60 days of the last therapy.

[0094] The phrase "refractory disease" refers to a disease in which a minimal response to therapy is not achieved or progresses to disease progression.

[0095] The terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting. As used herein, the indefinite articles "a / an" and "the" should be understood to include plural references unless the context clearly dictates otherwise.

[0096] Throughout the disclosure, each aspect of the disclosure may be presented in the form of a range. It should be understood that the description in range format is merely for convenience and brevity, and should not be interpreted as an inflexible limitation on the scope of the disclosure. Therefore, the description of a range should be considered to have all possible subranges and individual numerical values ​​within the range that are precisely disclosed. For example, the description of a range such as 1 to 6 should be considered to have the precisely disclosed subranges such as 1 to 3, 1 to 4, 1 to 5, 2 to 4, 2 to 6, 3 to 6, and individual numerical values ​​within the range, for example, 1, 2, 2.7, 3, 4, 5, 5.3, and 6. As another example, a range such as 95%-99% identity includes sub-ranges with 95%, 96%, 97%, 98% or 99% identity, including 96%-99%, 96%-98%, 96%-97%, 97%-99%, 97%-98% and 98%-99% identity, etc. This applies regardless of the breadth of the range.

[0097] vector Standard recombinant techniques can be used to obtain polynucleotide sequences encoding the CARs described herein. The desired polynucleotide sequences can be isolated and sequenced from antibody producing cells, such as hybridoma cells. Alternatively, polynucleotides can be synthesized using a nucleotide synthesizer or PCR techniques.

[0098] The present disclosure further provides a vector comprising a nucleic acid sequence encoding a CAR disclosed herein. The vector may be, for example, a plasmid, a cosmid, a viral vector (e.g., retrovirus or adenovirus) or a phage. Suitable vectors and methods for vector production are well known in the art (see, for example, Sambrook et al. and Ausubel et al.).

[0099] In addition to the nucleic acid sequence encoding the CAR disclosed herein, the vector preferably includes expression control sequences that provide for expression of the nucleic acid sequence in a host cell, such as a promoter, enhancer, polyadenylation signal, transcription terminator, internal ribosome entry site (IRES), etc. Exemplary expression control sequences are known in the art and described, for example, in Goeddel, Gene Expression Technology: Methods in Enzymology, Vol. 185, Academic Press, San Diego, Calif. (1990).

[0100] In some embodiments, the vector comprises a promoter. Numerous promoters recognized by many potential host cells are well known. The selected promoter can be operably linked to the cistron DNA encoding the CAR disclosed herein by removing the promoter from the source DNA by restriction enzyme digestion and inserting the isolated promoter sequence into the vector of the present application. Numerous promoters from many different sources are well known in the art, including constitutive, inducible and inhibitable promoters. Representative promoter sources include, for example, viral, mammalian, insect, plant, yeast and bacterial, and suitable promoters from these sources are readily available or can be produced synthetically based on publicly available sequences, for example, from depository centers such as ATCC and other commercial or private sources. Promoters can be unidirectional (i.e., initiate transcription in one direction) or bidirectional (i.e., initiate transcription in the 3' or 5' direction). Non-limiting examples of promoters include, for example, the T7 bacterial expression system, the pBAD(araA) bacterial expression system, the cytomegalovirus (CMV) promoter, the SV40 promoter and the RSV promoter. Inducible promoters include, for example, the Tet system (U.S. Pat. Nos. 5,464,758 and 5,814,618), the ecdysone-inducible system (No et al., Proc. Natl. Acad. Sci., 93: 3346-3351 (1996)), the T-REX™ system (Invitrogen, Carlsbad, Calif.), the LACSWITCH™ system (Stratagene, San Diego, Calif.), and the Cre-ERT tamoxifen-inducible recombinase system (Indra et al., Nuc. Acid. Res., 27: 4324- 4327 (1999); Nuc. Acid. Res., 28: e99 (2000); U.S. Pat. No. 7,112,715; and Kramer & Fussenegger, Methods Mol. Biol, 308: 123-144 (2003)). (2005)).

[0101] In some embodiments, the vector includes an "enhancer." As used herein, the term "enhancer" refers to a DNA sequence that, for example, increases the transcription of a nucleic acid sequence operably linked thereto. Enhancers may be located several kilobases away from the coding region of a nucleic acid sequence and can mediate the binding of regulatory factors, changes in DNA methylation patterns or DNA structure. Many enhancers from a variety of different sources are well known in the art and are available in cloned polynucleotides or within cloned polynucleotides (e.g., from depository centers such as ATCC, and other commercial or private sources). Many polynucleotides that contain a promoter (e.g., the commonly used CMV promoter) also contain enhancer sequences. Enhancers can be located upstream, within, or downstream of a coding sequence. The term "Ig enhancer" refers to an enhancer element derived from an enhancer region that maps within the immunoglobulin (Ig) locus. Such Ig enhancers include, for example, heavy chain (μ) 5' enhancers, light chain (κ) 5' enhancers, κ and μ intronic enhancers, and 3' enhancers (see generally, Paul WE (ed.), Fundamental Immunology, 3rd Edition, Raven Press, New York (1993), pages 353-363, and U.S. Patent No. 5,885,827).

[0102] In some embodiments, a vector contains a "selectable marker gene." As used herein, the term "selectable marker gene" refers to a nucleic acid sequence that allows cells expressing the nucleic acid sequence to be specifically selected or targeted in the presence of a corresponding selection agent. Suitable selectable marker genes are known in the art and are described, for example, in International Patent Application Publication Nos. WO 1992 / 08796 and WO 1994 / 28143; Wigler et al., Proc. Natl. Acad. Sci. USA, 77: 3567 (1980); O'Hare et al., Proc. Natl. Acad. Sci. USA, 78: 1527 (1981); Mulligan & Berg, Proc. Natl. Acad. Sci. USA, 78: 2072 (1981); Colberre-Garapin et al., J. Mol. Biol., 150: 1 (1981); Santerre et al., Gene, 30: 147 (1984); Kent et al., Science, 237: 901-903 (1987); Wigler et al., Cell, IP. 223 (1977), Szybalska & Szybalski, Proc. Natl. Acad. Sci. USA, 48: 2026 (1962), Lowy et al., Cell, 22: 817 (1980), and U.S. Patent Nos. 5,122,464 and 5,770,359.

[0103] In some embodiments, a vector is an "episomal expression vector" or "episome," which is replicable in a host cell and persists as an extrachromosomal segment of DNA in the host cell under appropriate selective pressure (see, e.g., Conese et al., Gene Therapy, 11: 1735-1742 (2004)). Representative commercially available episomal expression vectors include, but are not limited to, episomal plasmids that utilize the Epstein-Barr virus nuclear antigen 1 (EBNA1) and the Epstein-Barr virus (EBV) origin of replication (oriP). The vectors pREP4, pCEP4, pREP7, and pcDNA3.1 from Invitrogen (Carlsbad, Calif.) and pB-CMV from Stratagene (La Jolla, Calif.) represent non-limiting examples of episomal vectors that use T antigen and the SV40 origin of replication instead of EBNA1 and oriP.

[0104] In some embodiments, the vector is an "integrative expression vector," which may integrate randomly into the DNA of a host cell, or may contain recombination sites to allow recombination between the expression vector and specific sites in the chromosomal DNA of the host cell. Such integrative expression vectors can utilize endogenous expression control sequences of the host cell chromosome to achieve expression of the desired protein. Examples of vectors that integrate in a site-specific manner include, for example, components of the flp-in system (e.g., pcDNA™5 / FRT) from Invitrogen (Carlsbad, CA), or the cre-lox system, which may be found, for example, in the pExchange-6 core vector from Stratagene (La Jolla, CA). Examples of vectors that randomly integrate into a host cell chromosome include, for example, pcDNA3.1 (introduced in the absence of T antigen) from Invitrogen (Carlsbad, CA), and pCI or pFNI OA (ACT) FLEXI™ from Promega (Madison, WI).

[0105] In some embodiments, the vector is a viral vector. Exemplary viral expression vectors include, but are not limited to, adenovirus-based vectors (e.g., the adenovirus-based Per.C6 system available from Crucell, Inc. (Leiden, The Netherlands)), lentivirus-based vectors (e.g., lentivirus-based pLP1 from Life Technologies (Carlsbad, Calif.)), and retroviral vectors (e.g., pFB-ERV plus pCFB-EGSH from Stratagene (La Jolla, Calif.)). In a preferred embodiment, the viral vector is a lentiviral vector.

[0106] A vector containing a nucleic acid of the invention encoding a CAR is introduced into a host cell, which is capable of expressing the encoded CAR, and includes any suitable prokaryotic or eukaryotic cell. Preferred host cells are those that can be easily and reliably grown, have reasonably fast growth rates, have well-characterized expression systems, and can be easily and efficiently transformed or transfected.

[0107] As used herein, the term "host cell" refers to any type of cell that may contain an expression vector. The host cell may be a eukaryotic cell, such as a plant, animal, fungus, or algae, or a prokaryotic cell, such as a bacterium or a protozoan. The host cell may be a cultured cell or a primary cell, i.e., directly isolated from a living organism, such as a human. The host cell may be an adherent cell or a suspension cell, i.e., a cell that grows in suspension. Suitable host cells are known in the art and include, for example, DH5α E. coli cells, Chinese hamster ovary cells, monkey VERO cells, COS cells, HEK 293 cells, and the like. In a preferred embodiment, the host cell is a HEK 293 cell. In some embodiments, the HEK 293 cell is derived from the ATCC SD-3515 line. In some embodiments, the HEK 293 cell is derived from the IU-VPF MCB line. In some embodiments, the HEK 293 cell is derived from the IU-VPF MWCB line. In some embodiments, the host cell may be a peripheral blood lymphocyte (PBL), a peripheral blood mononuclear cell (PBMC), or a natural killer cell (NK). Preferably, the host cell is a natural killer (NK) cell. More preferably, the host cell is a T cell.

[0108] To amplify or replicate a recombinant expression vector, the host cell may be a prokaryotic cell, e.g., a DH5α cell. To produce a virus from a viral expression vector, the host cell may be a eukaryotic cell, e.g., a HEK 293 cell. To produce a recombinant CAR, the host cell may be a mammalian cell. The host cell is preferably a human cell. The host cell may be any cell type, from any type of tissue, and at any developmental stage. Methods for selecting suitable mammalian host cells, and for transforming, culturing, amplifying, screening, and purifying the cells are known in the art.

[0109] In some examples, the disclosure provides an isolated host cell that expresses a nucleic acid sequence encoding a CAR described herein.

[0110] In some embodiments, the host cell is a T cell. The T cells of the present disclosure may be any T cell, for example, cultured T cells, such as primary T cells, or T cells from a cultured T cell line, or T cells obtained from a mammal. If obtained from a mammal, the T cells may be obtained from many sources, including, but not limited to, blood, bone marrow, lymph nodes, thymus, or other tissues or fluids. Additionally, the T cells may be enriched or purified. The T cells are preferably human T cells (e.g., isolated from a human). The T cells may be at any stage of development, including, but not limited to, CD4+ / CD8+ double positive T cells, CD4+ helper T cells, e.g., Th and Th2 cells, CD8+ T cells (e.g., cytotoxic T cells), tumor infiltrating cells, memory T cells, naive T cells, and the like. In one embodiment, the T cells are CD8+ T cells or CD4+ T cells. T cell lines are available, for example, from the American Type Culture Collection (ATCC, Manassas, VA) and the German Collection of Microbial Cell Cultures (DSMZ), and include, for example, Jurkat cells (ATCC TIB-152), Sup-Tl cells (ATCC CRL-1942), RPMI 8402 cells (DSMZ ACC-290), Karpas 45 cells (DSMZ ACC-545), and derivatives thereof.

[0111] In some embodiments, the host cell is a natural killer (NK) cell. NK cells are cytotoxic lymphocytes that function in the innate immune system. NK cells are defined as large granular lymphocytes and constitute a third class of cells differentiated from the common lymphoid progenitor, which also produces B and T lymphocytes (see, e.g., Immunobiology, 5th ed., Janeway et al., eds., Garland Publishing, New York, NY (2001)). NK cells differentiate and mature in the bone marrow, lymph nodes, spleen, tonsils, and thymus. After maturation, NK cells enter the cycle as large lymphocytes with unique cytotoxic granules. NK cells can recognize and kill some abnormal cells, e.g., some tumor cells and virus-infected cells, and are believed to be important in the innate immune defense against intracellular pathogens. As discussed above with respect to T cells, the NK cells may be any NK cells, e.g., cultured NK cells, such as primary NK cells, or NK cells from a cultured NK cell line, or NK cells obtained from a mammal. If obtained from a mammal, the NK cells may be obtained from a number of sources, including, but not limited to, blood, bone marrow, lymph nodes, thymus, or other tissues or fluids. Additionally, the NK cells may be enriched or purified. The NK cells are preferably human NK cells (e.g., isolated from a human). NK cell lines are available, for example, from the American Type Culture Collection (ATCC, Manassas, VA), and include, for example, NK-92 cells (ATCC CRL-2407), NK92MI cells (ATCC CRL-2408), and derivatives thereof.

[0112] In some examples, the nucleic acid sequence encoding a CAR can be introduced into a cell by "transfection," "transformation," or "transduction." As used herein, "transfection," "transformation," or "transduction" refers to the introduction of one or more exogenous polynucleotides into a host cell by using physical or chemical methods. Many transfection techniques are known in the art and include, for example, calcium phosphate DNA coprecipitation (e.g., Murray EJ (ed.), Methods in Molecular Biology, Vol. 7, Gene Transfer and Expression Protocols, Humana Press (1991)), DEAE-dextran, electroporation, cationic liposome-mediated transfection, microparticle bombardment facilitated by tungsten particles (Johnston, Nature, 346: 776-777 (1990)), and strontium phosphate DNA coprecipitation (Brash et al., Mol. Cell Biol., 7: 2031-2034 (1987)). After the infected granules have grown in suitable packaging cells, the phage or viral vector may be introduced into the host cell, where many packaging cells are commercially available.

[0113] Chimeric Antigen Receptor International Patent Publication No. WO 2018 / 028647 is incorporated herein by reference in its entirety. U.S. Patent Publication No. 2018 / 0230225 is incorporated herein by reference in its entirety.

[0114] The present disclosure provides a method of treating a subject with a cell expressing a chimeric antigen receptor (CAR). The CAR comprises an extracellular antigen binding domain that contains one or more single domain antibodies. In various embodiments, a BCMA-targeting CAR (also referred to herein as a "BCMA CAR") is provided, which comprises a polypeptide, the polypeptide comprising (a) an extracellular antigen binding domain comprising an anti-BCMA binding moiety, (b) a transmembrane domain, and (c) an intracellular signaling domain. In some embodiments, the anti-BCMA binding moiety is camelid, chimeric, human, or humanized. In some embodiments, the intracellular signaling domain comprises a primary intracellular signaling domain of an immune effector cell (e.g., a T cell). In some embodiments, the primary intracellular signaling domain is derived from CD4. In some embodiments, the primary intracellular signaling domain is derived from CD3zeta. In some embodiments, the intracellular signaling domain comprises a costimulatory signaling domain. In some embodiments, the costimulatory signaling domain is derived from a costimulatory molecule selected from the group consisting of CD27, CD28, CD137, OX40, CD30, CD40, CD3, LFA-1, ICOS, CD2, CD7, LIGHT, NKG2C, B7-H3, a ligand of CD83, and combinations thereof. In some embodiments, the transmembrane domain is derived from CD137.

[0115] In some embodiments, the BCMA CAR further comprises a hinge domain (e.g., a CD8α hinge domain) located between the C-terminus of the extracellular antigen binding domain and the N-terminus of the transmembrane domain. In some embodiments, the BCMA CAR further comprises a signal peptide (e.g., a CD8α signal peptide) located at the N-terminus of the polypeptide. In some embodiments, the polypeptide comprises, from N-terminus to C-terminus, a CD8α signal peptide, an extracellular antigen binding domain, a CD8α hinge domain, a CD28 transmembrane domain, a first costimulatory signaling domain derived from CD28, a second costimulatory signaling domain derived from CD137, and a primary intracellular signaling domain derived from CD4. In some embodiments, the polypeptide comprises, from N-terminus to C-terminus, a CD8α signal peptide, an extracellular antigen binding domain, a CD8α hinge domain, a CD8α transmembrane domain, a second costimulatory signaling domain derived from CD137, and a primary intracellular signaling domain derived from CD3ζ. In some embodiments, the BCMA CAR is monospecific. In some embodiments, the BCMA CAR is monovalent.

[0116] The present application further provides a CAR having two or more (including but not limited to any one of 2, 3, 4, 5, 6 or more) binding moieties that specifically bind to an antigen, such as BCMA. In some embodiments, one or more binding moieties are antigen-binding fragments. In some embodiments, one or more binding moieties comprise a single domain antibody. In some embodiments, one or more binding moieties comprise a VHH.

[0117] In some embodiments, the CAR is a multivalent (e.g., bivalent, trivalent or higher) CAR comprising a polypeptide, the polypeptide comprising (a) an extracellular antigen-binding domain that comprises a plurality (e.g., at least about any one of 2, 3, 4, 5, 6 or more) of binding moieties that specifically bind to an antigen (e.g., a tumor antigen); (b) a transmembrane domain; and (c) an intracellular signaling domain.

[0118] In some embodiments, the binding moieties, e.g., VHHs (including multiple VHHs or being a first VHH and / or a second VHH) are camelid, chimeric, human or humanized. In some embodiments, the binding moieties or VHHs are linked to each other via peptide bonds or peptide linkers. In some embodiments, each peptide linker is no more than about 50 (e.g., no more than about any one of 35, 25, 20, 15, 10 or 5) amino acids in length.

[0119] In some embodiments the first BCMA binding moiety and / or the second BCMA binding moiety is an anti-BCMA VHH. In some embodiments the first BCMA binding moiety is a first anti-BCMA VHH and the second BCMA binding moiety is a second anti-BCMA VHH.

[0120] In some embodiments, the first BCMA binding moiety and the second BCMA binding moiety are linked to each other via a peptide linker. In some embodiments, the peptide linker comprises the amino acid sequence of SEQ ID NO: 3. In some embodiments, the peptide linker comprises a polypeptide encoded by the nucleic acid sequence of SEQ ID NO: 11.

[0121] In some embodiments, the CAR further comprises a hinge domain (e.g., a CD8α hinge domain) located between the C-terminus of the extracellular antigen binding domain and the N-terminus of the transmembrane domain. In some embodiments, the CAR further comprises a signal peptide (e.g., a CD8α signal peptide) located at the N-terminus of the polypeptide.

[0122] Without wishing to be bound by theory, multivalent CARs or those CARs comprising an extracellular antigen binding domain containing a first BCMA binding moiety and a second BCMA binding moiety may be particularly suitable for targeting multimeric antigens through synergistic binding of different antigen binding sites or for enhancing binding affinity or avidity for antigens. Improved avidity may allow for a significant reduction in the dose of CAR-T cells required to achieve a therapeutic effect, for example a dose range of 4.0×10 4 to 1.0×10 6 CAR-T cells / kilogram subject mass, or 3.0 x 10 6 to 1.0×10 8 total CAR-T expressing cells. Monovalent CARs, such as bb2121, may need to be administered at 5 to 10 times these amounts to achieve comparable efficacy. In various embodiments, the reduced dose ranges can provide a significant reduction in cytokine release syndrome (CRS) and other potentially dangerous side effects of CAR-T therapy.

[0123] The various binding moieties (e.g., extracellular antigen binding domains including a first BCMA binding moiety and a second BCMA binding moiety) in the CAR described herein can be linked to each other via a peptide linker. The peptide linkers linking the different binding moieties (e.g., VHHs) can be the same or different. The different domains of the CAR can also be linked to each other via a peptide linker. In some examples, the binding moieties (e.g., VHHs) are directly linked without any peptide linker.

[0124] The peptide linker in the CAR described herein may have any suitable length. In some embodiments, the length of the peptide linker is at least about any one of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 25, 30, 35, 40, 50, 75, 100 or more amino acids. In some embodiments, the length of the peptide linker is no more than about any one of 100, 75, 50, 40, 35, 30, 25, 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5 or fewer amino acids: In some embodiments, the length of the peptide linker is any one of about 1 amino acid to about 10 amino acids, about 1 amino acid to about 20 amino acids, about 1 amino acid to about 30 amino acids, about 5 amino acids to about 15 amino acids, about 10 amino acids to about 25 amino acids, about 5 amino acids to about 30 amino acids, about 10 amino acids to about 30 amino acids, about 30 amino acids to about 50 amino acids, about 50 amino acids to about 100 amino acids, or about 1 amino acid to about 100 amino acids.

[0125] The CAR of the present application comprises a transmembrane domain that can be directly or indirectly linked to an extracellular antigen-binding domain.

[0126] The CAR may comprise a T cell activation moiety. The T cell activation moiety may be any suitable moiety derived or derived from any suitable molecule. In one embodiment, for example, the T cell activation moiety comprises a transmembrane domain. The transmembrane domain may be any transmembrane domain derived or derived from any molecule known in the art. For example, the transmembrane domain may be derived or derived from a CD8a molecule or a CD28 molecule. Without wishing to be bound by theory, CD8 is a transmembrane glycoprotein that functions as a co-receptor for the T cell receptor (TCR) and is primarily expressed on the surface of cytotoxic T cells. The most common form of CD8 exists as a dimer composed of CD8α and CD8β chains. CD28 is expressed on T cells and provides a costimulatory signal necessary for T cell activation. CD28 is a receptor for CD80 (B7.1) and CD86 (B7.2). In a preferred embodiment, CD8α and CD28 are human.

[0127] In addition to the transmembrane domain, the T cell activation moiety may further comprise an intracellular (i.e., cytoplasmic) T cell signaling domain. The intracellular T cell signaling domain may be obtained or derived from a CD28 molecule, a CD3ζ molecule or modified forms thereof, a human Fc receptor gamma (FcRγ) chain, a CD27 molecule, an OX40 molecule, a 4-1BB molecule, or other intracellular signaling molecules known in the art. Without wishing to be bound by theory, (1) CD28 is a T cell marker important for T cell costimulation, (2) CD3ζ associates with the TCR to produce a signal and contains an immunoreceptor tyrosine-based activation motif (ITAM), and (3) 4-1BB, also called CD137, promotes differentiation of T lymphocytes and improves long-term survival of T lymphocytes by delivering a strong costimulatory signal to T cells. In a preferred embodiment, CD28, CD3ζ, 4-1BB, OX40, and CD27 are human.

[0128] The T cell stimulatory domain of a CAR encoded by a nucleic acid sequence disclosed herein may comprise any one of the above transmembrane domains in any combination with any one or more of the above intercellular T cell signaling domains. For example, a nucleic acid sequence disclosed herein can encode a CAR comprising a CD28 transmembrane domain and the intracellular T cell signaling domains of CD28 and CD3ζ. Alternatively, for example, a nucleic acid sequence disclosed herein can encode a CAR comprising a CD8α transmembrane domain and the intracellular T cell signaling domains of CD28, CD3ζ, Fc receptor gamma (FcRγ) chain, and / or 4-1BB.

[0129] In some embodiments, the CAR polypeptide further comprises a signal peptide located at the N-terminus of the polypeptide. In some embodiments, the signal peptide is derived from CD8α. In some embodiments, the signal peptide comprises the amino acid sequence of SEQ ID NO: 1. In some embodiments, the signal peptide comprises a polypeptide encoded by the nucleic acid sequence of SEQ ID NO: 9.

[0130] In some embodiments, the transmembrane domain comprises the amino acid sequence of SEQ ID NO: 6. In some embodiments, the transmembrane domain comprises a polypeptide encoded by a nucleic acid sequence of SEQ ID NO: 14.

[0131] In some embodiments, the intracellular signaling domain comprises a primary intracellular signaling domain of an immune effector cell. In some embodiments, the intracellular signaling domain is derived from CD3zeta. In some embodiments, the intracellular signaling domain comprises at least one costimulatory signaling domain. In some embodiments, the intracellular signaling domain comprises the amino acid sequence of SEQ ID NO:8. In some embodiments, the intracellular signaling domain comprises a polypeptide encoded by a nucleic acid sequence of SEQ ID NO:16. In some embodiments, the intracellular signaling domain comprises the amino acid sequence of SEQ ID NO:7. In some embodiments, the intracellular signaling domain comprises a polypeptide encoded by a nucleic acid sequence of SEQ ID NO:15.

[0132] In some embodiments, the CAR polypeptide further comprises a hinge domain located between the C-terminus of the extracellular antigen binding domain and the N-terminus of the transmembrane domain. In some embodiments, the hinge domain comprises the amino acid sequence of SEQ ID NO: 5. In some embodiments, the hinge domain comprises a polypeptide encoded by the nucleic acid sequence of SEQ ID NO: 13.

[0133] In some embodiments, the CAR comprises one or more or all of SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, and SEQ ID NO:8. In one embodiment, the CAR comprises SEQ ID NO:17. In some embodiments, the CAR comprises a polypeptide encoded by one or more or all of the nucleic acid sequences of SEQ ID NO:9, SEQ ID NO:10, SEQ ID NO:11, SEQ ID NO:12, SEQ ID NO:13, SEQ ID NO:14, SEQ ID NO:15, and SEQ ID NO:16.

[0134] Immune effector cell compositions "Immune effector cell" refers to an immune cell capable of exerting an immune effector function. In some embodiments, the immune effector cell expresses at least FcγRIII and exerts ADCC effector function. Examples of immune effector cells that mediate ADCC include peripheral blood mononuclear cells (PBMCs), natural killer (NK) cells, monocytes, cytotoxic T cells, neutrophils, and eosinophils. In some embodiments, the immune effector cell is a T cell. In some embodiments, the T cell is an autologous T cell. In some embodiments, the T cell is an allogeneic T cell. In some embodiments, the T cell is CD4+ / CD8-, CD4- / CD8+, CD4+ / CD8+, CD4- / CD8-, or a combination thereof. In some embodiments, the T cell expresses a CAR and produces IL-2, TNF, and / or TNF after binding to a target cell (e.g., a CD20+ or ​​CD19+ tumor cell). In some examples, CD8+ T cells express the CAR and lyse the antigen-specific target cells after binding to the target cells.

[0135] Biological methods for introducing vectors into immune effector cells include using DNA and RNA vectors. Viral vectors have become the most widely used method for inserting genes into mammalian, e.g., human, cells. Chemical methods for introducing vectors into immune effector cells include colloidal dispersion systems, such as macromolecular complexes, nanocapsules, microspheres, beads, and lipid-based systems, including oil-in-water emulsions, micelles, mixed micelles, and liposomes. An exemplary colloidal system as an in vitro delivery vehicle is a liposome (e.g., an artificial membrane vesicle).

[0136] The dosage form according to the present specification is 3.0 x 10 7 to 1.0×10 8In some embodiments, the dosage form comprises 3.0×10 CAR-T cells, the CAR-T cells comprising a CAR containing a polypeptide, the polypeptide comprising (a) an extracellular antigen-binding domain comprising a first BCMA binding moiety that specifically binds to a first epitope of BCMA and a second BCMA binding moiety that specifically binds to a second epitope of BCMA, (b) a transmembrane domain, and (c) an intracellular signaling domain, wherein the first epitope and the second epitope are distinct. In some embodiments, the dosage form comprises 3.0×10 7 From 4.0×10 7 In some embodiments, the dosage form comprises 3.5×10 CAR-T cells. 7 From 4.5×10 7 In some embodiments, the dosage form comprises 4.0×10 CAR-T cells. 7 From 5.0×10 7 In some embodiments, the dosage form comprises 4.5×10 CAR-T cells. 7 From 5.5 × 10 7 In some embodiments, the dosage form comprises 5.0×10 CAR-T cells. 7 From 6.0×10 7 In some embodiments, the dosage form contains 5.5×10 CAR-T cells. 7 From 6.5×10 7 In some embodiments, the dosage form contains 6.0×10 CAR-T cells. 7 From 7.0×10 7 In some embodiments, the dosage form comprises 6.5×10 CAR-T cells. 7 From 7.5 × 10 7 In some embodiments, the dosage form contains 7.0×10 CAR-T cells. 7 From 8.0×10 7 In some embodiments, the dosage form comprises 7.5×10 CAR-T cells. 7 From 8.5×10 7 In some embodiments, the dosage form contains 8.0×10 CAR-T cells. 7 From 9.0×10 7 In some embodiments, the dosage form contains 8.5×10 CAR-T cells.7 From 9.5×10 7 In some embodiments, the dosage form contains 9.0×10 CAR-T cells. 7 to 1.0×10 8 Contains CAR-T cells.

[0137] In some embodiments, the dosage form provided is 3.0×10 7 to 1.0×10 8 In some embodiments, the dosage form comprises 3.0×10 engineered immune effector cells (e.g., T cells), the engineered immune effector cells comprising a CAR containing a polypeptide, the polypeptide comprising (a) an extracellular antigen binding domain comprising a first anti-BCMA VHH that specifically binds to a first epitope of BCMA and a second anti-BCMA VHH that specifically binds to a second epitope of BCMA, (b) a transmembrane domain, and (c) an intracellular signaling domain, where the first epitope and the second epitope are different. In some embodiments, the dosage form comprises 3.0×10 7 From 4.0×10 7 In some embodiments, the dosage form comprises 3.5×10 CAR-T cells. 7 From 4.5×10 7 In some embodiments, the dosage form comprises 4.0×10 CAR-T cells. 7 From 5.0×10 7 In some embodiments, the dosage form comprises 4.5×10 CAR-T cells. 7 From 5.5 × 10 7 In some embodiments, the dosage form comprises 5.0×10 CAR-T cells. 7 From 6.0×10 7 In some embodiments, the dosage form contains 5.5×10 CAR-T cells. 7 From 6.5×10 7 In some embodiments, the dosage form contains 6.0×10 CAR-T cells. 7 From 7.0×10 7 In some embodiments, the dosage form comprises 6.5×10 CAR-T cells. 7 From 7.5 × 107 In some embodiments, the dosage form contains 7.0×10 CAR-T cells. 7 From 8.0×10 7 In some embodiments, the dosage form comprises 7.5×10 CAR-T cells. 7 From 8.5×10 7 In some embodiments, the dosage form contains 8.0×10 CAR-T cells. 7 From 9.0×10 7 In some embodiments, the dosage form contains 8.5×10 CAR-T cells. 7 From 9.5×10 7 In some embodiments, the dosage form contains 9.0×10 CAR-T cells. 7 to 1.0×10 8 Contains CAR-T cells.

[0138] In some embodiments, the cell population of the CAR-T formulation described herein includes T cells or T cell populations, e.g., at different stages of differentiation. Stages of T cell differentiation include naive T cells, stem central memory T cells, central memory T cells, effector memory T cells, and final effector T cells, from the least differentiated to the most differentiated. After antigen exposure, naive T cells proliferate and differentiate into memory T cells, e.g., stem central memory T cells and central memory T cells, and then into effector memory T cells. After receiving the appropriate T cell receptor, costimulatory signals, and inflammatory signals, memory T cells further differentiate into final effector T cells. See, e.g., Restifo. Blood. 124.4(2014):476-77, and Joshi et al. J. Immunol. 180.3(2008):1309-15.

[0139] Naive T cells may have the following expression pattern of cell surface markers: CCR7+, CD62L+, CD45RO-, CD95-. Stem central memory T cells (Tscm) may have the following expression pattern of cell surface markers: CCR7+, CD62L+, CD45RO-, CD95+. Central memory T cells (Tcm) may have the following expression pattern of cell surface markers: CCR7+, CD62L+, CD45RO+, CD95+. Effector memory T cells (Tem) may have the following expression pattern of cell surface markers: CCR7-, CD62L-, CD45RO+, CD95+. Final effector T cells (Teff) may have the following expression pattern of cell surface markers: CCR7-, CD62L-, CD45RO-, CD95+. See, e.g., Gattinoni et al. Nat. Med. 17(2011):1290-7, and Flynn et al. Clin. Translat. Immunol. 3(2014):e20.

[0140] Pharmaceutical Compositions and Formulations The present application further provides a pharmaceutical composition, which comprises any one of the engineered immune effector cells comprising any one of the anti-BCMA antibodies disclosed herein or any one of the CARs (e.g., BCMA CARs) described herein, and a pharma- ceutical acceptable vector. The pharmaceutical composition may be prepared in the form of a lyophilized formulation or aqueous solution by mixing any of the immune effector cells having the desired purity described herein with an optional pharma- ceutical acceptable vector, excipient or stabilizer (Remington's Pharmaceutical Sciences 16th Edition, edited by Osol, A. (1980)). In some embodiments, the pharmaceutical composition of the CAR-T cells further comprises an excipient selected from dimethyl sulfoxide or dextran-40.

[0141] The compositions described herein may be administered as part of a pharmaceutical composition comprising one or more vectors. The choice of vector is determined in part by the particular nucleic acid sequence, vector or host cell expressing the CAR disclosed herein, and the particular method for administering the nucleic acid sequence, vector or host cell expressing the CAR disclosed herein. As such, there are multiple suitable formulations of the pharmaceutical compositions of the present disclosure.

[0142] For example, the pharmaceutical composition may contain a preservative. Suitable preservatives may include, for example, methylparaben, propylparaben, sodium benzoate and benzalkonium chloride. Optionally, a mixture of two or more preservatives may be used. The preservative or mixture thereof is typically present in an amount of about 0.0001% to about 2% by weight of the total composition.

[0143] A buffering agent may also be used in the composition. Suitable buffering agents include, for example, citric acid, sodium citrate, phosphoric acid, potassium phosphate, and various other acids and salts. Optionally, a mixture of two or more buffering agents may be used. The buffering agent or mixture thereof is typically present in an amount of about 0.001% to about 4% by weight of the total composition.

[0144] Compositions comprising a nucleic acid sequence encoding a CAR disclosed herein or a host cell expressing a CAR disclosed herein may be prepared as an inclusion complex, such as a cyclodextrin inclusion complex, or liposomes. Liposomes may be used to target host cells (e.g., T cells or NK cells) or the nucleic acid sequences disclosed herein to a specific tissue. Liposomes may also be used to increase the half-life of the nucleic acid sequences disclosed herein. Many methods may be used to prepare liposomes, such as those described in, for example, Szoka et al., Ann. Rev. Biophys. Bioeng., 9: 467 (1980), and U.S. Pat. Nos. 4,235,871, 4,501,728, 4,837,028, and 5,019,369. The compositions may use timed, delayed, and sustained release delivery systems, such that delivery of the compositions disclosed herein occurs prior to sensitization of the site to be treated and has sufficient time to cause sensitization of the site to be treated. Many types of release delivery systems are available and known to those of skill in the art. Such systems increase convenience for the subject and the physician by avoiding repeated administration of the composition, and may be particularly suitable for some exemplary compositions of the present disclosure.

[0145] In some embodiments, the CAR-T cells are about 1.0×10 5 From 2.0×10 5 Cells / kg, 1.5 x 10 5 From 2.5 × 10 5 Cells / kg, 2.0 x 10 5 From 3.0×10 5 Cells / kg, 2.5 x 10 5 From 3.5 × 10 5 Cells / kg, 3.0 x 10 5 From 4.0×10 5 Cells / kg, 3.5 x 10 5 From 4.5×10 5 Cells / kg, 4.0 x 10 5 From 5.0×10 5 Cells / kg, 4.5 x 10 5From 5.5 × 10 5 Cells / kg, 5.0 x 10 5 From 6.0×10 5 Cells / kg, 5.5 x 10 5 From 6.5×10 5 Cells / kg, 6.0 x 10 5 From 7.0×10 5 Cells / kg, 6.5 x 10 5 From 7.5 × 10 5 Cells / kg, 7.0 x 10 5 From 8.0×10 5 Cells / kg, 7.5 x 10 5 From 8.5×10 5 Cells / kg, 8.0 x 10 5 From 9.0×10 5 Cells / kg, 8.5 x 10 5 From 9.5×10 5 Cells / kg, 9.0 x 10 5 to 1.0×10 6 In a preferred embodiment, the dose is about 0.75×10 6 In some embodiments, the CAR-T cells are formulated at 1.0×10 cells / kg. 8 It is formulated in doses of less than one cell / subject.

[0146] Treatment method The present application further relates to methods and compositions for use in cellular immunotherapy. In some embodiments, the cellular immunotherapy is for treating cancer in a subject, including but not limited to hematological malignancies and solid tumors. In some embodiments, the subject is a human. In some embodiments, the method is suitable for treating adult and pediatric populations, including all age subgroups, and may be used as any line of treatment, including first or subsequent lines.

[0147] Any of the anti-BCMA VHHs, CARs and engineered immune effector cells (e.g., CAR-T cells) described herein can be used in methods to treat cancer. In some embodiments, the immune effector cells are autologous. In some embodiments, the immune effector cells are allogeneic.

[0148] In some embodiments, the CAR-T cells are about 1.0×10 5 From 2.0×10 5 Cells / kg, 1.5 x 10 5 From 2.5 × 10 5 Cells / kg, 2.0 x 10 5 From 3.0×10 5 Cells / kg, 2.5 x 10 5 From 3.5 × 10 5 Cells / kg, 3.0 x 10 5 From 4.0×10 5 Cells / kg, 3.5 x 10 5 From 4.5×10 5 Cells / kg, 4.0 x 10 5 From 5.0×10 5 Cells / kg, 4.5 x 10 5 From 5.5 × 10 5 Cells / kg, 5.0 x 10 5 From 6.0×10 5 Cells / kg, 5.5 x 10 5 From 6.5×10 5 Cells / kg, 6.0 x 10 5 From 7.0×10 5 Cells / kg, 6.5 x 10 5 From 7.5 × 10 5 Cells / kg, 7.0 x 10 5 From 8.0×10 5 Cells / kg, 7.5 x 10 5 From 8.5×10 5 Cells / kg, 8.0 x 10 5 From 9.0×10 5 Cells / kg, 8.5 x 10 5 From 9.5×10 5 Cells / kg, 9.0 x 10 5to 1.0×10 6 Cells / kg, 1.0 x 10 6 From 2.0×10 6 Cells / kg, 1.5 x 10 6 From 2.5 × 10 6 Cells / kg, 2.0 x 10 6 From 3.0×10 6 Cells / kg, 2.5 x 10 6 From 3.5 × 10 6 Cells / kg, 3.0 x 10 6 From 4.0×10 6 Cells / kg, 3.5 x 10 6 From 4.5×10 6 Cells / kg, 4.0 x 10 6 From 5.0×10 6 Cells / kg, 4.5 x 10 6 From 5.5 × 10 6 cells / kg or 5.0 x 10 6 From 6.0×10 6 In a preferred embodiment, the dose is about 0.75×10 cells / kg. 6 In some embodiments, the CAR-T cells are about 1.0×10 cells / kg per subject. 8 The cells are administered in doses of 100 cells.

[0149] In some embodiments, the CAR-T cells are administered at a dose of 1.0×10 per subject. 8 In some embodiments, the CAR-T cells are administered at a dose of about 3.0 to 4.0 x 10 7 In some embodiments, the CAR-T cells are administered at a dose of about 3.5 to 4.5 x 10 cells. 7 In some embodiments, the CAR-T cells are administered at a dose of about 4.0 to 5.0 x 10 cells. 7 In some embodiments, the CAR-T cells are administered at a dose of about 4.5 to 5.5 x 10 cells. 7 In some embodiments, the CAR-T cells are administered at a dose of about 5.0 to 6.0 x 10 cells. 7In some embodiments, the CAR-T cells are administered at a dose of about 5.5 to 6.5 x 10 cells. 7 In some embodiments, the CAR-T cells are administered at a dose of about 6.0 to 7.0 x 10 cells. 7 In some embodiments, the CAR-T cells are administered at a dose of about 6.5 to 7.5 x 10 cells. 7 In some embodiments, the CAR-T cells are administered at a dose of about 7.0 to 8.0 x 10 cells. 7 In some embodiments, the CAR-T cells are administered at a dose of about 7.5 to 8.5 x 10 cells. 7 In some embodiments, the CAR-T cells are administered at a dose of about 8.0 to 9.0 x 10 cells. 7 In some embodiments, the CAR-T cells are administered at a dose of about 8.5 to 9.5 x 10 cells. 7 In some embodiments, the CAR-T cells are administered at a dose of about 9.0×10 7 to 1.0×10 8 The cells are administered in doses of 100 cells.

[0150] In some embodiments, the CAR-T cells are about 0.693×10 6 In some embodiments, the CAR-T cells are administered at a dose of about 0.52×10 live CAR-positive T cells / kg. 6 In some embodiments, the CAR-T cells are administered at a dose of about 0.94×10 live CAR-positive T cells / kg. 6 In some embodiments, the CAR-T cells are administered at a dose of about 0.709×10 live CAR-positive T cells / kg. 6 In some embodiments, the CAR-T cells are administered at a dose of about 0.51×10 live CAR-positive T cells / kg. 6 In some embodiments, the CAR-T cells are administered at a dose of about 0.95×10 live CAR-positive T cells / kg. 6 The CAR-T cells are administered at a dose of 100 mg / kg of live CAR-positive T cells. In some embodiments, the CAR-T cells are administered in an outpatient setting.

[0151] In some examples, the composition comprising CAR-T cells administered to the subject further comprises an excipient selected from dimethylsulfoxide or dextran-40.

[0152] In some embodiments, the CAR-T cells are administered (e.g., at any of the above-mentioned doses) in one or more intravenous infusions. In some embodiments, the administration of the CAR-T cells is by a single intravenous infusion. In some embodiments, a single intravenous infusion is administered using one bag of CAR-T cells. In some embodiments, the administration of the one bag of CAR-T cells is completed from the time the one bag of CAR-T cells is thawed to three hours after the one bag of CAR-T cells is thawed. In some embodiments, a single intravenous administration is administered using two bags of the CAR-T cells. In some embodiments, the administration of each bag of the two bags of CAR-T cells is completed from the time the first bag of the two bags of CAR-T cells is thawed to three hours after the first bag of CAR-T cells is thawed.

[0153] In some embodiments, the time from initial apheresis to administration of the CAR-T cells is less than 41, 47, 54, 61, 68, 75, 82, 89, 96, 103, 110, 117, 124, 131, 138, 145, 152, 159, 166, or 167 days. In some embodiments, the time from initial apheresis to administration of the CAR-T cells is more than 41, 47, 54, 61, 68, 75, 82, 89, 96, 103, 110, 117, 124, 131, 138, 145, 152, 159, 166, or 167 days.

[0154] In some embodiments, the lymphodepleting regimen is administered prior to administration of the CAR-T cells. In some embodiments, the lymphodepleting regimen includes administration of cyclophosphamide and / or administration of fludarabine. In some embodiments, the lymphodepleting regimen is administered intravenously. In some embodiments, the lymphodepleting regimen is administered 5 to 7 days prior to administration of the CAR-T cells. In some embodiments, the lymphodepleting regimen is administered 2 to 4 days prior to administration of the CAR-T cells. In some embodiments, the lymphodepleting regimen includes intravenous administration of cyclophosphamide and fludarabine 5 to 7 days prior to administration of the CAR-T cells. In some embodiments, the lymphodepleting regimen includes intravenous administration of cyclophosphamide and fludarabine 2 to 4 days prior to administration of the CAR-T cells. In some embodiments, the lymphodepleting regimen includes intravenous administration of cyclophosphamide and fludarabine 2 to 4 days prior to administration of the CAR-T cells. In some embodiments, the lymphodepleting regimen includes intravenous administration of 300 mg / m 2 In some embodiments, the lymphodepleting regimen includes administering cyclophosphamide intravenously at a dose of 30 mg / m 2 In some embodiments, the lymphodepleting regimen is administered daily for three days. If the delay in administration of CAR-T cells exceeds 14 days, the lymphodepleting regimen may be repeated.

[0155] In some embodiments, the method of treating with CAR-T cells further comprises treating the subject for cytokine release syndrome (CRS) within 3 days of administering the CAR-T cells without significantly reducing in vivo expansion of the CAR-T cells. In some embodiments, treating CRS comprises administering an IL-6R inhibitor to the subject. In some embodiments, the IL-6R inhibitor is an antibody. In some embodiments, the IL-6 inhibitor inhibits IL-6R by binding to the extracellular domain of IL-6R. In some embodiments, the IL-6R inhibitor blocks the binding of IL-6 to IL-6R. In some embodiments, the IL-6R inhibitor is tocilizumab. CRS may be identified based on clinical symptoms [see approved labeling herein]. In some embodiments, other causes of fever, hypoxia, and hypotension are assessed and treated. Laboratory tests may monitor disseminated intravascular coagulation, hematological parameters, and pulmonary, cardiac, renal, and hepatic function. CRS may be managed based on the suggestions in Table 1 of the approved labeling disclosed herein. These methods may include administering levetiracetam to patients experiencing CRS for antiepileptic prophylaxis. In some embodiments, these methods include monitoring patients experiencing grade 2 or higher CRS (e.g., hypotension unresponsive to fluids, or oxygen deprivation requiring supplemental oxygenation) with continuous cardiac telemetry and pulse oximetry. In some embodiments, intensive care unit level monitoring and supportive care may be used for severe or life-threatening CRS. For CRS that is refractory to frontline interventions (e.g., tocilizumab or tocilizumab and corticosteroids), these methods include alternative treatment regimens (i.e., higher corticosteroid doses, alternative anti-cytokine agents such as anti-IL1 and / or anti-TNFα, anti-T cell therapy). Refractory CRS is characterized by fever, end-organ toxicity (e.g., oxygen deprivation, hypotension) not improved within 12 hours of frontline intervention, or development of HLH / MAS.

[0156] In some embodiments, the method of treating with CAR-T cells further comprises treating the subject with an infused prodrug comprising an antipyretic and an antihistamine at most 1 hour prior to administration of the CAR-T cells. In some embodiments, the antipyretic comprises paracetamol or acetaminophen. In some embodiments, the antipyretic is administered orally or intravenously to the subject. In some embodiments, the antipyretic is administered to the subject at a dose of 650 mg to 1000 mg. In some embodiments, the antihistamine comprises diphenhydramine. In some embodiments, the antihistamine is administered to the subject at a dose of 25 mg to 50 mg, or an equivalent dose. The composition may be administered to the mammal using standard administration techniques, including host cells expressing a CAR-encoding nucleic acid sequence disclosed herein or a vector comprising a CAR-encoding nucleic acid sequence disclosed herein, including oral, intravenous, intraperitoneal, subcutaneous, pulmonary, transdermal, intramuscular, intranasal, buccal, sublingual, or suppository administration. The composition is preferably suitable for parenteral administration. As used herein, the term "parenteral" includes intravenous, intramuscular, subcutaneous, rectal, intravaginal, and intraperitoneal administration. More preferably, the composition is administered to the mammal by intravenous, intraperitoneal, or subcutaneous injection using peripheral systemic delivery. Most preferably, the composition is administered by intravenous infusion.

[0157] A composition comprising a host cell expressing a CAR-encoding nucleic acid sequence disclosed herein or a vector comprising a CAR-encoding nucleic acid sequence disclosed herein can be administered with one or more additional therapeutic agents, which can be co-administered to a mammal. "Co-administration" refers to administration of one or more additional therapeutic agents and a composition comprising a host cell disclosed herein or a vector disclosed herein sufficiently close in time that a CAR disclosed herein can potentiate the effect of the one or more other therapeutic agents, or vice versa. In this embodiment, a composition comprising a host cell disclosed herein or a vector disclosed herein can be administered first, and then one or more additional therapeutic agents can be administered, or vice versa.

[0158] The CAR-expressing cells described herein and at least one additional therapeutic agent may be administered simultaneously (in the same or separate compositions) or sequentially. In the case of sequential administration, the CAR-expressing cells described herein may be administered first and the additional agent may be administered second, or the order of administration may be reversed.

[0159] In some embodiments, the lymphodepleting regimen is administered prior to administration of the CAR-T cells. In some embodiments, the lymphodepleting regimen is administered about 2 to about 7 days prior to administration of the CAR-T cells. In some embodiments, the lymphodepleting regimen is administered intravenously. In some embodiments, the lymphodepleting regimen includes administration of cyclophosphamide or administration of fludarabine. In some embodiments, the cyclophosphamide is administered at a dose of 300 mg / m 2 In some embodiments, the fludarabine is administered intravenously at a dose of 30 mg / m 2 It is administered intravenously.

[0160] In some embodiments, cyclophosphamide at 300 mg / m 2 and fludarabine 30 mg / m 2 and intravenously administering the CAR-T cells about 2 to about 7 days prior to the administration of the CAR-T cells.

[0161] In some embodiments, the subject further undergoes a bridging therapy, wherein the bridging therapy includes short-term treatment with at least one bridging drug between apheresis and the lymphodepleting regimen, and wherein the at least one bridging drug has previously resulted in disease stabilization, minimal response, partial response, best partial response, complete response, or stringent complete response in the subject. In some embodiments, the subject's tumor burden increases despite the bridging therapy. In some embodiments, the subject's tumor burden increases by about 25% or more despite the bridging therapy. Suitable bridging therapies include, for example, dexamethasone, bortezomib, cyclophosphamide, and pomalidomide. In some embodiments, the bridging therapy includes dexamethasone. In some embodiments, the bridging therapy includes bortezomib. In some embodiments, the bridging therapy includes cyclophosphamide. In some embodiments, the bridging therapy includes pomalidomide.

[0162] In some embodiments, at most about 1 hour prior to the administration of the CAR-T cells, the subject is treated with an administered prodrug comprising an antipyretic and an antihistamine. In some embodiments, the antipyretic comprises paracetamol or acetaminophen. In some embodiments, the antipyretic is administered orally or intravenously to the subject. In some embodiments, the antipyretic is administered to the subject at a dose of 650 mg to 1000 mg. In some embodiments, the antihistamine comprises diphenhydramine. In some embodiments, the antihistamine is administered to the subject at a dose of 25 mg to 50 mg, or the equivalent thereof. In some embodiments, the antipyretic comprises paracetamol or acetaminophen, wherein the antipyretic comprises a dose of 650 mg to 1000 mg, and wherein the antihistamine comprises diphenhydramine, wherein the antihistamine comprises a dose of 25 mg to 50 mg, or an equivalent thereof, and wherein the antihistamine comprises a dose of 25 mg to 50 mg, or an equivalent thereof, and wherein the antipyretic comprises a dose of 25 mg to 50 mg, or an equivalent thereof, and wherein the antipyretic comprises a dose of 25 mg to 50 mg, or an equivalent thereof, and wherein the antihistamine ...

[0163] In some embodiments, the method includes administering cyclophosphamide 300 mg / m daily for three days prior to administration of the CAR-T cells. 2 Intravenous (IV) and fludarabine 30 mg / m 2 administer a lymphodepleting chemotherapy regimen including IV and infusion of a prodrug containing an antipyretic (e.g., acetaminophen 650 to 1000 mg orally or intravenously) and an antihistamine (e.g., diphenhydramine 25 to 50 mg or equivalent orally or intravenously), where: CAR-T cells were administered 2 to 4 days after lymphodepleting chemotherapy was completed. The CAR-T cells are administered 30-60 minutes after the infusion prodrug is administered.

[0164] In some embodiments, CAR-T cell administration is not administered or CAR-T cell administration is delayed if the patient has any one of the following: clinically significant active infection or inflammatory disorder, or cyclophosphamide and fludarabine conditioned non-hematological toxicity > Grade 3, excluding grade 3 nausea, vomiting, diarrhea, or constipation. Administration of CAR-T cells should be delayed until these events have resolved < Grade 1. In some embodiments, prophylactic systemic corticosteroids are not administered.

[0165] In some embodiments, the method further comprises diagnosing the subject with cytokine release syndrome (CRS). In a preferred embodiment, the diagnosis is based on the American Society for Transplantation and Cellular Therapy (ASTCT) consensus classification, formerly the American Society for Blood and Marrow Transplantation (ASBMT). Table 13 provides a non-limiting summary of the ASTCT consensus classification of CRS diagnoses. In some embodiments, CRS is assessed by assessing the levels of one or more or all of IL-6, IL-10, IFN-□, C-reactive protein (CRP) and ferritin.

[0166] In some embodiments, the method further comprises treating the subject for cytokine release syndrome (CRS). In some embodiments, the CRS is treated with an antipyretic. In some embodiments, the CRS is treated with an anticytokine therapy. In some embodiments, the treatment of CRS occurs more than about 3 days after infusion. In some embodiments, the treatment of CRS occurs without significantly reducing in vivo expansion of the CAR-T cells. In some embodiments, the method further comprises treating the subject for cytokine release syndrome more than about 3 days after administration of the CAR-T cells without significantly reducing in vivo expansion of the CAR-T cells. In some embodiments, the treatment of CRS comprises administering an IL-6R inhibitor to the subject. In some embodiments, the IL-6R inhibitor is an antibody. In some embodiments, the antibody inhibits IL-6R by binding to the extracellular domain of IL-6R. In some embodiments, the IL-6R inhibitor blocks the binding of IL-6 to IL-6R. In some embodiments, the IL-6R inhibitor is tocilizumab. In some embodiments, the anti-cytokine therapy includes administration of tocilizumab. In some embodiments, the anti-cytokine therapy includes administration of a steroid. In some embodiments, the treatment of CRS includes treatment with a monoclonal antibody other than tocilizumab. In some embodiments, the antibody other than tocilizumab targets a cytokine. In some embodiments, the cytokine targeted by the antibody other than tocilizumab is IL-1. In some embodiments, the antibody targeting IL-1 is anakinra. In some embodiments, the cytokine targeted by the antibody other than tocilizumab is TNFα. In some embodiments, the treatment of CRS includes administering a corticosteroid to the subject. In some embodiments, the treatment of CRS includes the use of a vasoconstrictor. In some embodiments, the treatment of CRS includes intubation or mechanical ventilation. In some embodiments, the treatment of CRS includes administering cyclophosphamide to the subject. In some embodiments, treating CRS comprises administering etanercept to the subject.In some embodiments, treating CRS comprises administering levetiracetam to the subject. In some embodiments, treating CRS comprises supportive care.

[0167] In some embodiments, the method further comprises diagnosing immune cell effector associated neurotoxicity (ICANS) in the subject. In some embodiments, the diagnosis is made based on the National Cancer Institute Common Terminology Standard for Adverse Events (NCI CTCAE) standard. In some embodiments, the diagnosis is made based on the NCI CTCAE standard version 5.0. In some embodiments, the diagnosis is made based on the American Society for Transplant and Cellular Therapy (ASTCT) consensus classification system. In some embodiments, the subject has neurotoxicity consistent with ICAN. Table 14 provides a non-limiting summary of the ASTCT consensus classification system for ICANS diagnosis. In some embodiments, the treatment of ICANS comprises administering an IL-6R inhibitor to the subject. In some embodiments, the IL-6R inhibitor is an antibody. In some embodiments, the antibody inhibits IL-6R by binding to the extracellular domain of IL-6R. In some embodiments, the IL-6R inhibitor blocks the binding of IL-6 to IL-6R. In some embodiments, the IL-6R inhibitor is tocilizumab. In some embodiments, the treatment of ICANS comprises administering to the subject an IL-1 inhibitor. In some embodiments, the IL-1 inhibitor is an antibody. In a preferred embodiment, the IL-1 inhibitory antibody is anakinra. In some embodiments, the treatment of ICANS comprises administering to the subject a corticosteroid. In some embodiments, the treatment of ICANS comprises administering to the subject levetiracetam. In some embodiments, the treatment of ICANS comprises administering to the subject dexamethasone. In some embodiments, the treatment of ICANS comprises administering to the subject methylprednisolone sodium succinate. In some embodiments, the treatment of ICANS comprises administering to the subject pethidine. In some embodiments, the treatment of ICANS comprises administering to the subject one or more or all of tocilizumab, anakinra, a corticosteroid, levetiracetam, dexamethasone, methylprednisolone sodium succinate, or pethidine.

[0168] If a neurotoxic complication is suspected during CRS, or vice versa, the method may include administering: Corticosteroids with more invasive interventions for CRS and neurotoxicity levels in Tables 1 and 2 based on the approved labeling disclosed herein Tocilizumab according to CRS levels in Table 1 of the approved labeling disclosed herein Antiepileptic drugs with neurotoxicity in Table 2 of the approved labeling disclosed herein

[0169] In some embodiments, the method further comprises diagnosing cytopenia in the subject. In some embodiments, the cytopenia comprises one or more or all of lymphopenia, neutropenia, and thrombocytopenia. Without being bound by theory, grade 3 or grade 4 lymphopenia, but not grade 2 or less, is considered to be a condition in which the lymphocyte count is greater than or equal to 0.5×10 9Grade 3 or 4 neutropenia, but not grade 2 or less, is characterized by a neutrophil count of less than 1000 cells / microliter of the subject's blood sample, and Grade 3 or 4 thrombocytopenia, but not grade 2 or less, is characterized by a platelet count of less than 50,000 cells / microliter of the subject's blood sample. In some examples, more than 75% of subjects suffering from Grade 3 or Grade 4 lymphopenia after administration of the CAR-T cells have recovered to Grade 2 or less lymphopenia at day 60 after administration of the CAR-T cells. In some examples, more than 80% of subjects suffering from Grade 3 or Grade 4 lymphopenia after administration of the CAR-T cells have recovered to Grade 2 or less lymphopenia at day 60 after administration of the CAR-T cells. In some examples, greater than 85% of subjects with grade 3 or grade 4 lymphopenia after administration of the CAR-T cells have recovered to grade 2 or less lymphopenia at day 60 after administration of the CAR-T cells. In some examples, greater than 90% of subjects with grade 3 or grade 4 lymphopenia after administration of the CAR-T cells have recovered to grade 2 or less lymphopenia at day 60 after administration of the CAR-T cells. In some examples, greater than 70% of subjects with grade 3 or grade 4 neutropenia after administration of the CAR-T cells have recovered to grade 2 or less neutropenia at day 60 after administration of the CAR-T cells. In some examples, greater than 75% of subjects with grade 3 or grade 4 neutropenia after administration of the CAR-T cells have recovered to grade 2 or less neutropenia at day 60 after administration of the CAR-T cells. In some examples, greater than 80% of subjects with grade 3 or grade 4 neutropenia after administration of the CAR-T cells have recovered to grade 2 or less neutropenia at day 60 after administration of the CAR-T cells. In some examples, greater than 85% of subjects with grade 3 or grade 4 neutropenia after administration of the CAR-T cells have recovered to grade 2 or less neutropenia at day 60 after administration of the CAR-T cells.In some embodiments, more than 30% of subjects with grade 3 or grade 4 thrombocytopenia after administration of the CAR-T cells have recovered to grade 2 or less thrombocytopenia at day 60 after administration of the CAR-T cells. In some embodiments, more than 34% of subjects with grade 3 or grade 4 thrombocytopenia after administration of the CAR-T cells have recovered to grade 2 or less thrombocytopenia at day 60 after administration of the CAR-T cells. In some embodiments, more than 38% of subjects with grade 3 or grade 4 thrombocytopenia after administration of the CAR-T cells have recovered to grade 2 or less thrombocytopenia at day 60 after administration of the CAR-T cells. In some embodiments, more than 42% of subjects with grade 3 or grade 4 thrombocytopenia after administration of the CAR-T cells have recovered to grade 2 or less thrombocytopenia at day 60 after administration of the CAR-T cells.

[0170] When a composition comprising a host cell expressing a CAR-encoding nucleic acid sequence disclosed herein or a vector comprising a CAR-encoding nucleic acid sequence disclosed herein is administered to a mammal (e.g., a human), the biological activity of the CAR can be measured by any suitable method known in the art. According to the methods disclosed herein, the CAR binds to BCMA in multiple myeloma cells, and these multiple myeloma cells are destroyed. The binding of the CAR to BCMA on the surface of multiple myeloma cells can be measured by any suitable method known in the art, including, for example, ELISA and flow cytometry. The ability of the CAR to destroy multiple myeloma cells can be measured by any suitable method known in the art, such as the cytotoxicity assays described in, for example, Kochenderfer et al., J. Immunotherapy, 32(7): 689-702 (2009), and Herman et al. J. Immunological Methods, 285(1): 25-40 (2004). The biological activity of the CAR can also be measured by assaying the expression of several cytokines, such as CD 107a, IFNγ, IL-2, and TNF.

[0171] The methods described herein may be used to treat a variety of cancers, including solid and liquid cancers. In some embodiments, these methods are for treating multiple myeloma. The methods described herein may be used in adjunctive or novel adjunctive treatments, as a combination therapy with first, second, third, or other types of cancer therapy known in the art (e.g., chemotherapy, surgery, radiation, gene therapy, immunotherapy, bone marrow transplantation, stem cell transplantation, targeted therapy, cryotherapy, ultrasound therapy, photodynamic therapy, radiofrequency ablation, etc.).

[0172] In some embodiments, the cancer is multiple myeloma. In some embodiments, the cancer is stage I, stage II or stage III, and / or stage A or stage B multiple myeloma according to the Durie-Salmon classification system. In some embodiments, the cancer is stage I, stage II or stage III multiple myeloma according to the International Classification System published by the International Myeloma Working Group (IMWG). In some embodiments, the multiple myeloma is aggressive.

[0173] In some embodiments, the subject has received prior treatment including at least three prior lines of treatment. In some embodiments, the median number of prior lines of treatment is six. In some embodiments, the prior lines of treatment include surgery, radiation therapy, or autologous or allogeneic transplantation, or any combination of these treatments. In some embodiments, the at least three prior lines of treatment include drug treatment, and the drug is a proteasome inhibitor (PI). Non-limiting examples of PIs include bortezomib, carbenzimib, and izomib. In some embodiments, the at least three prior lines of treatment include drug treatment, and the drug is an immunomodulatory drug (IMiD). Non-limiting examples of IMiDs include lenalidomide, pomalidomide, and thalidomide. In some embodiments, the at least three prior lines of treatment include drug treatment, and the drug is a corticosteroid. Non-limiting examples of corticosteroids include dexamethasone and prednisone. In some embodiments, at least three prior lines of treatment include treatment with a drug, the drug being an alkylating agent. In some embodiments, at least three prior lines of treatment include treatment with a drug, the drug being an anthracycline. In some embodiments, at least three prior lines of treatment include treatment with a drug, the drug being an anti-CD38 antibody. Non-limiting examples of anti-CD38 antibodies include daratumumab, isatuximab, and the research antibody TAK-079. In some embodiments, at least three prior lines of treatment include treatment with a drug, the drug being elotuzumab. In some embodiments, at least three prior lines of treatment include treatment with a drug, the drug being panobinostat. In some embodiments, at least three prior lines of treatment include treatment with at least one drug, the at least one drug being at least one of a PI, an IMiD, and an anti-CD38 antibody. In some embodiments, the at least three prior lines of treatment include treatment with at least one drug, and the at least one drug includes at least one of a PI, an IMiD, and an alkylating agent.In some embodiments, the subject relapses after the at least three prior lines of therapy. In some embodiments, the multiple myeloma is refractory to one or more or all of bortezomib, carbenzimib, izomib, lenalidomide, pomalidomide, thalidomide, dexamethasone, prednisone, alkylating agents, daratumumab, isatuximab, TAK-079, elotuzumab, and panobinostat. In some embodiments, the multiple myeloma is refractory to at least two drugs after the at least three prior lines of therapy. In some embodiments, the at least two drugs that are refractory to multiple myeloma include a PI and an IMiD. In some embodiments, the multiple myeloma is refractory to at least three drugs after the at least three prior lines of therapy. In some embodiments, the multiple myeloma is refractory to at least four drugs after the at least three prior lines of therapy. In some embodiments, the at least four prior lines of therapy include treatment with at least one agent, the at least one agent including at least one of a PI, an IMiD, an anti-CD38 antibody, and an alkylating agent. In some embodiments, the multiple myeloma is incurable with at least five agents after the at least three prior lines of therapy.

[0174] In some embodiments, the subject has about 10% to about 30% bone marrow plasma cells prior to said administration of said CAR-T cells.

[0175] In some embodiments, a bone marrow aspirate or biopsy may be performed for clinical evaluation, or a bone marrow aspirate may be performed for biomarker evaluation. In some embodiments, clinical classification (morphology, cytogenetics, immunohistochemistry or immunofluorescence or flow cytometry) may be performed. In some embodiments, immunophenotyping may be performed on a portion of the bone marrow aspirate to monitor checkpoint ligand expression in BCMA, CD138 positive multiple myeloma cells, and checkpoint expression in T cells. In some embodiments, next generation sequencing (NGS) of bone marrow aspirate DNA may be used to monitor minimal residual disease (MRD) in subjects. NGS of bone marrow aspirate DNA is known to those of skill in the art. In some embodiments, NGS is performed via clonoSeq. In some embodiments, a baseline bone marrow aspirate may be used to define a myeloma clone, and a post-treatment sample may be used to assess MRD negativity. In some embodiments, MRD negativity status may be based on evaluable samples. In some embodiments, an evaluable sample is one or more or all of correction, quality control, and evaluable cell suitability under a particular sensitivity level. In some embodiments, the sensitivity level is 10 -6 In some embodiments, the sensitivity level is 10 -6 and the sensitivity level is 10 -5 In some embodiments, the sensitivity level is 10 -4 In some embodiments, the sensitivity level is 10 -3 It is.

[0176] In some embodiments, the International Myeloma Working Group (IMWG) response criteria are used to assess the subject's response to the treatment method, and these criteria are summarized in Table 6. In some embodiments, the response may be classified as stringent complete response (sCR). In some embodiments, the response may be classified as complete response (CR), which is worse than stringent complete response (sCR). In some embodiments, the response may be classified as very good partial response (VGPR), which is worse than complete response (CR). In some embodiments, the response may be classified as partial response (PR), which is worse than very good partial response (VGPR). In some embodiments, the response may be classified as minimal response (MR), which is worse than partial response (PR). In some embodiments, the response may be classified as stable disease (SD), which is worse than minimal response (MR). In some embodiments, the response may be classified as progressive disease (PD), which is worse than stable disease.

[0177] In some embodiments, tests to assess response criteria based on the International Myeloma Working Group (IMWG) are measurement of myeloma protein (M-protein) in serum and urine, serum calcium corrected for albumin, bone marrow examination, bone survey, and documentation of extramedullary plasmacytoma.

[0178] Non-limiting examples of tests for the measurement of blood and urinary M-protein are known to those skilled in the art and include serum quantitative Ig, serum protein electrophoresis (SPEP), serum immunofixation electrophoresis, serum FLC assay, 24-hour urinary M-protein electrophoresis quantitative (UPEP), urine immunofixation electrophoresis and serum β2-microglobulin.

[0179] It is known to those skilled in the art to calculate serum calcium corrected for albumin in a blood sample to detect hypercalcemia. Without wishing to be bound by theory, calcium is bound to albumin, and only unbound (free) calcium has biological activity, therefore serum calcium levels must be adjusted for abnormal albumin levels ("corrected serum calcium").

[0180] In some embodiments, a bone survey of any or all of the skull, entire spine, pelvis, thorax, humerus, femur, and any other bones may be performed and assessed by radiography ("X-ray") or low-dose computed tomography (CT) diagnostic quality scans without the use of IV contrast, both of which are known to those of skill in the art. In some embodiments, X-ray or CT scans may be performed locally after T cell administration and before disease progression is confirmed to document response or progression with or without clinical indication based on symptoms. In some embodiments, magnetic resonance imaging (MRI) may be used to assess bone disease, but does not replace a bone survey. MRI is known to those of skill in the art. In some embodiments, a radionuclide bone scan may be used at screening in addition to a complete bone survey to document disease status in two ways. Radionuclide bone scans are known to those of skill in the art. In some embodiments, a radionuclide bone scan and a complete bone survey may be performed simultaneously. In some embodiments, a radionuclide bone scan may not replace a complete bone survey. In some embodiments, when a subject shows disease progression manifested by painful symptoms in bone changes, disease progression can be documented by bone surveys or other radiographs depending on the symptoms experienced by the subject.

[0181] In some embodiments, extramedullary plasmacytoma can be documented by clinical examination or MRI. In some embodiments, extramedullary plasmacytoma can be documented by CT scan if there are no contraindications to the use of IV contrast. In some embodiments, extramedullary plasmacytoma can be documented by fusion of a positron emission tomography (PET) scan with a CT scan if the CT portion is of sufficient diagnostic quality. In some embodiments, subjects may undergo local assessment, measurement, or evaluation of measurable sites of extramedullary disease every 4 weeks until a confirmed CR or confirmed disease progression occurs. In some embodiments, evaluation of extramedullary plasmacytoma may be performed every 12 weeks.

[0182] In some embodiments, the sum of the products of perpendicular diameters of previous extramedullary plasmacytomas may be decreased by more than 90% or at least 50% to qualify for VGPR or PR or MR, respectively. In some embodiments, the sum of the products of perpendicular diameters of previous extramedullary plasmacytomas must increase by at least 50%, or the longest diameter of a previous lesion that is >1 cm on the short axis must increase by at least 50%, or a new plasmacytoma must develop to qualify for disease progression. In some embodiments, if not all extramedullary plasmacytomas present are reported, the sum of the products of perpendicular diameters of reported plasmacytomas is increased by at least 50% to qualify for disease progression. In some embodiments, if the study treatment interferes with the immunofixation assay, CR may be defined as the disappearance of the original M protein associated with multiple myeloma upon immunofixation.

[0183] In some embodiments, the response in a subject to a therapeutic method is assessed based on a change in disease burden or tumor burden. Disease burden or tumor burden refers to the type of measurable disease in a subject. In some embodiments, the change in tumor burden can be assessed based on a change in paraprotein levels after treatment. In some embodiments, the paraprotein is M-protein in serum. In some embodiments, the paraprotein is M-protein in serum. In some embodiments, the change in tumor burden can be assessed based on the difference between impaired and intact free light chains (dFLC). In some embodiments, the change in tumor burden is assessed based on the maximum paraprotein reduction relative to baseline, i.e., before administration of the CAR-T cells. In some embodiments, the change in tumor burden is assessed at a median follow-up period of 28 days or more after administration of the CAR-T cells. In some embodiments, the change in tumor burden is assessed at a median follow-up period of 1 month or more after administration of the CAR-T cells. In some embodiments, the change in tumor burden is assessed at a median follow-up period of 3 months or more after administration of the CAR-T cells. In some embodiments, the change in tumor burden is assessed at a median follow-up period of 6 months or more after administration of the CAR-T cells. In some embodiments, the change in tumor burden is assessed at a median follow-up period of 9 months or more after administration of the CAR-T cells. In some embodiments, the change in tumor burden is assessed at a median follow-up period of 12 months or more after administration of the CAR-T cells.

[0184] In some embodiments, the subject is re-treated by administration of a second dose of CAR-T cells via a second intravenous infusion. In some embodiments, the re-treatment dose is 1.0×10 5 From 5.0×10 6 In some embodiments, the retreatment dose comprises about 0.75×10 CAR-T cells / kilogram of subject mass. 5In some embodiments, the subject is re-treated if the subject exhibits disease progression after a best response of minimal response or better response after the first infusion of CAR-T cells. In some embodiments, the time from the first infusion of CAR-T cells to detection of disease progression comprises at least six months.

[0185] In one aspect, a method of treating a subject suffering from multiple myeloma is provided, the method comprising administering to the subject a composition comprising a therapeutically effective number of T cells that comprise a chimeric antigen receptor (CAR) to deliver a dose of T cells expressing a CAR (CAR-T cells) to the subject by a single intravenous infusion.

[0186] In some embodiments, the subject has undergone prior therapy including at least three prior lines of therapy. In some embodiments, the at least three prior lines of therapy include treatment with at least one agent, the at least one agent including at least one of a PI, an IMiD, and an anti-CD38 antibody. In some embodiments, the subject relapses after the at least three prior lines of therapy.

[0187] In some embodiments, the multiple myeloma is refractory to at least two drugs after said at least three prior lines of therapy. In some embodiments, the at least two drugs that are refractory to curing the subject include a PI and an IMiD. In some embodiments, the multiple myeloma is refractory to at least three drugs after said at least three prior lines of therapy. In some embodiments, the multiple myeloma is refractory to at least four drugs after said at least three prior lines of therapy. In some embodiments, the multiple myeloma is refractory to at least five drugs after said at least three prior lines of therapy.

[0188] In some embodiments, the subject age is greater than 65 years. In some embodiments, the subject is black or African American. In some embodiments, the subject has received three prior lines of therapy. In some embodiments, the subject has received at least four prior lines of therapy. In some embodiments, the multiple myeloma or subject is three drug refractory, i.e., the multiple myeloma or subject is three drug refractory. In some embodiments, the multiple myeloma or subject is five drug refractory, i.e., the multiple myeloma or subject is five drug refractory. In some embodiments, the subject has standard risk cytogenetics. In some embodiments, the subject has high risk cytogenetics. In some embodiments, the subject or multiple myeloma is characterized as stage III according to the International Classification System. In some embodiments, the subject has about 10% to about 30% bone marrow plasma cells prior to said administration of said CAR-T cells. In some embodiments, the subject has about 31% to about 59% bone marrow plasma cells prior to said administration of said CAR-T cells. In some embodiments, the subject has about 60% to about 100% bone marrow plasma cells prior to said administration of said CAR-T cells. In some embodiments, BCMA expression in the subject's tumor is lower than the median in a multiple myeloma patient population or any randomly selected population. In some embodiments, BCMA expression in the subject's tumor is equal to or greater than the median in a multiple myeloma patient population or any randomly selected population. In some embodiments, plasmacytoma is present in the subject. In some embodiments, the plasmacytoma is bone-based. In some embodiments, the plasmacytoma is extramedullary. In some embodiments, the plasmacytoma is bone-based and extramedullary.

[0189] In some embodiments, the method of treatment effectively obtains a reduced tumor burden in the subject. In some embodiments, the method of treatment effectively obtains a reduced tumor burden in the subject by about 1% to about 100%, about 60% to about 100%, about 65% to about 100%, about 70% to about 100%, about 75% to about 100%, about 80% to about 100%, about 85% to about 100%, about 90% to about 100%, about 92% to about 100%, about 95% to about 100%, about 96% to about 100%, about 97% to about 100%, about 98% to about 100%, or about 99% to about 100%. In some embodiments, the method of treatment effectively obtains a reduced tumor burden in the subject by about 100%. In some embodiments, the treatment method is effective to obtain about 1% to about 100% reduced tumor burden in the subject at a rate of about 1% to about 100%. In some embodiments, the treatment method is effective to obtain about 60% to about 100% reduced tumor burden in the subject at a rate of about 1% to about 100%. In some embodiments, the treatment method is effective to obtain about 65% to about 100% reduced tumor burden in the subject at a rate of about 1% to about 92%. In some embodiments, the treatment method is effective to obtain about 70% to about 100% reduced tumor burden in the subject at a rate of about 1% to about 88%. In some embodiments, the treatment method is effective to obtain about 90% to about 100% reduced tumor burden in the subject at a rate of about 1% to about 88%. In some embodiments, the treatment method is effective to obtain about a 95% to about a 100% reduction in tumor burden in a subject at a rate of about 1% to about 88%. In some embodiments, the treatment method is effective to obtain about a 99% to about a 100% reduction in tumor burden in a subject at a rate of about 1% to about 88%. In some embodiments, the treatment method is effective to obtain about a 100% reduction in tumor burden in a subject at a rate of about 1% to about 83%.

[0190] In some embodiments, the method of treatment effectively achieves a negative minimal residual disease (MRD) status or effectively maintains the minimal residual disease (MRD) status in the subject. -6In some embodiments, the method of treatment effectively achieves a negative minimal residual disease (MRD) status in the subject at a sensitivity level of 10 -5 In some embodiments, the method of treatment effectively achieves a negative minimal residual disease (MRD) status in the subject at a sensitivity level of 10 -4 In some embodiments, the method of treatment effectively achieves a negative minimal residual disease (MRD) status in the subject at a sensitivity level of 10 -3 In some embodiments, the method of treatment effectively obtains a negative minimal residual disease (MRD) status in the subject at a sensitivity level of 0.1 to 1.0%. In some embodiments, the method of treatment effectively obtains a negative MRD status in the subject as assessed in bone marrow. In some embodiments, the method of treatment effectively maintains a negative MRD status in the subject as assessed in an evaluable bone marrow sample. In some embodiments, the method of treatment effectively obtains a negative MRD status in the subject as assessed in bone marrow at a follow-up period of about 28 days or more after the administration of the CAR-T cells, about 2 months or more after the administration of the CAR-T cells, about 3 months or more after the administration of the CAR-T cells, about 6 months or more after the administration of the CAR-T cells, about 9 months or more after the administration of the CAR-T cells, or about 12 months or more after the administration of the CAR-T cells. In some embodiments, the minimal residual disease (MRD) negativity status is obtained at a first follow-up period between about 28 days and about 179 days after the infusion of the CAR-T cells.

[0191] In some embodiments, the treatment method effectively maintains the initially achieved minimal residual disease (MRD) negativity in the subject. -5 In some embodiments, the treatment method effectively maintains MRD negativity at a sensitivity level of 10 -6 In some embodiments, the method of treatment effectively achieves a negative minimal residual disease (MRD) status in the subject at a sensitivity level of 10 -4 In some embodiments, the treatment method effectively maintains MRD negativity at a sensitivity level of 10-3 In some embodiments, the treatment method effectively maintains MRD negativity at a sensitivity level of 0.1 to 1.0. In some embodiments, the treatment method effectively maintains MRD negativity when assessed in a bone marrow sample. In some embodiments, the treatment method effectively maintains MRD negativity when assessed in an evaluable bone marrow sample. In some embodiments, the treatment method effectively maintains MRD negativity when assessed in bone marrow DNA. In some embodiments, the method effectively maintains the minimal residual disease (MRD) negativity in the subject as assessed in bone marrow at a second follow-up period of about 29 days to about 359 days after the administration of the CAR-T cells, about 29 days to about 9 months after the administration of the CAR-T cells, about 29 days to about 6 months after the administration of the CAR-T cells, about 29 days to about 3 months after the administration of the CAR-T cells, or about 29 days to about 2 months after the administration of the CAR-T cells. In some examples, the method effectively maintains in the subject a negative status of the minimal residual disease (MRD) as assessed in bone marrow at a second follow-up period of about 180 days to about 359 days after the infusion of the CAR-T cells. In some examples, the method effectively maintains in the subject a negative status of the minimal residual disease (MRD) as assessed in bone marrow at a second follow-up period of about 360 days to about 539 days after the infusion of the CAR-T cells.

[0192] In some embodiments, the efficacy of the treatment method is assessed by evaluating the proportion of subjects with MRD negative status. -6 The efficacy of the treatment method is assessed by assessing the proportion of subjects with MRD negative status at a sensitivity level of 10 -5 The efficacy of the treatment method is assessed by assessing the proportion of subjects with MRD negative status at a sensitivity level of 10 -4 The efficacy of the treatment method is assessed by assessing the proportion of subjects with MRD negative status at a sensitivity level of 10 -3The efficacy of the treatment method is assessed by assessing the proportion of subjects with MRD negative status at a median follow-up period of about 359 days after CAR-T cell administration, about 9 months after CAR-T cell administration, about 6 months after CAR-T cell administration, about 3 months after CAR-T cell administration, about 2 months after CAR-T cell administration, or about 29 days after CAR-T cell administration. In some examples, the efficacy of the treatment method is assessed by assessing the proportion of subjects with MRD negative status at a median follow-up period of about 12 months after the infusion of the CAR-T cells. -5 At a sensitivity threshold level of about 44% or less, at a follow-up period of about 12 months after said administration of said CAR-T cells, -5 At a sensitivity threshold level of about 100%, about 55% of patients had a 10% or greater survival rate at about 12 months follow-up after the infusion of the CAR-T cells. -5 At a sensitivity threshold level of about 65% or less, at a follow-up period of about 18 months after the infusion of the CAR-T cells, -4 At a sensitivity threshold level of about 57% or less, at a follow-up period of about 18 months after the infusion of the CAR-T cells, -4 At a sensitivity threshold level of 100 mg / kg / day, approximately 67% of patients had a 10% or higher CAR-T cell survival rate at a follow-up period of approximately 18 months after the infusion of the CAR-T cells. -4 At a sensitivity threshold level of about 76% or less, at a follow-up period of about 18 months after the infusion of the CAR-T cells, -5 At a sensitivity threshold level of about 47% or less, at a follow-up period of about 18 months after the infusion of the CAR-T cells, -5 At a sensitivity threshold level of 100 mg / kg / day, approximately 58% of patients had a 10% or higher CAR-T cell survival rate at a follow-up period of approximately 18 months after the infusion of the CAR-T cells. -5 At a sensitivity threshold level of about 68% or less, at a follow-up period of about 18 months after the infusion of the CAR-T cells, -6 At a sensitivity threshold level of about 10%, a rate of about 29% or less was observed during a follow-up period of about 18 months after the infusion of the CAR-T cells. -6At a sensitivity threshold level of about 10%, or at a follow-up period of about 18 months after the infusion of the CAR-T cells, -6 In some embodiments, the method effectively achieves a minimal residual disease (MRD) negative status in a rate of about 50% or less at a sensitivity threshold level of 100% or less at a follow-up period of about 12 months after the infusion of the CAR-T cells. -5 At a sensitivity threshold level of about 44% to about 65% of patients were treated with CAR-T cells for a follow-up period of about 18 months after the infusion of the CAR-T cells. -4 At a sensitivity threshold level of about 57% to about 76% of patients were treated with CAR-T cells for a follow-up period of about 18 months after the infusion of the CAR-T cells. -5 or at a follow-up period of about 18 months after said infusion of said CAR-T cells, -6 At a sensitivity threshold level of about 100%, the method effectively achieves a minimal residual disease (MRD) negative status in about 29% to about 50% of patients. In some embodiments, the method effectively achieves a minimal residual disease (MRD) negative status in about 10% to about 50% of patients at a follow-up period of about 12 months after the administration of the CAR-T cells. -5 At a sensitivity threshold level of 100 mg / kg / day, about 55% of patients had a 10% or higher CAR-T cell survival rate at about 18 months of follow-up after the infusion of the CAR-T cells. -4 At a sensitivity threshold level of 100 mg / kg / day, approximately 67% of patients had a 10% or higher CAR-T cell survival rate at a follow-up period of approximately 18 months after the infusion of the CAR-T cells. -5 At a sensitivity threshold level of about 10%, or at a follow-up period of about 18 months after the infusion of the CAR-T cells, -6 At a sensitivity threshold level of 0.1, the minimal residual disease (MRD) negativity status is effectively achieved in approximately 39% of cases.

[0193] In some embodiments, the efficacy of the treatment method is assessed by evaluating the proportion of subjects with evaluable bone marrow and negative MRD status. -6 The efficacy of the treatment method is assessed by assessing the proportion of subjects with evaluable bone marrow and MRD negative status at a sensitivity level of 10 -5The efficacy of the treatment method is assessed by assessing the proportion of subjects with evaluable bone marrow and MRD negative status at a sensitivity level of 10 -4 The efficacy of the treatment method is assessed by assessing the proportion of subjects with evaluable bone marrow and MRD negative status at a sensitivity level of 10 -3 The efficacy of the treatment method is assessed by assessing the proportion of subjects with evaluable bone marrow and MRD negative status at a median follow-up period of about 359 days after CAR-T cell administration, about 9 months after CAR-T cell administration, about 6 months after CAR-T cell administration, about 3 months after CAR-T cell administration, about 2 months after CAR-T cell administration, or about 29 days after CAR-T cell administration. In some embodiments, the method assesses the efficacy of the treatment method by assessing the proportion of subjects with evaluable bone marrow and MRD negative status at a median follow-up period of about 12 months after the infusion of the CAR-T cells. -5 At a sensitivity threshold level of 100%, about 83% or less of subjects with evaluable samples have a 10% or greater CAR-T cell response rate at a follow-up period of about 12 months after the infusion of the CAR-T cells. -5 At a sensitivity threshold level of 100%, about 93% of subjects with evaluable samples had a 10% or greater survival rate at about 12 months follow-up after the infusion of the CAR-T cells. -5 At a sensitivity threshold level of 100%, about 98% of subjects with evaluable samples had a 10% or greater survival rate at about 18 months follow-up after the infusion of the CAR-T cells. -5 At a sensitivity threshold level of 100%, about 82% or less of subjects with evaluable samples have a 100% or greater CAR-T cell response rate at a follow-up period of about 18 months after the infusion of the CAR-T cells. -5 At a sensitivity threshold level of 100%, in subjects with evaluable samples, a rate of about 92% or a follow-up period of about 18 months after the infusion of the CAR-T cells is about 10%. -5At a sensitivity threshold level of 0.1%, the method effectively obtains a minimal residual disease (MRD) negative status in about 97% or less of subjects with evaluable samples. In some embodiments, the method effectively obtains a minimal residual disease (MRD) negative status in about 10% or less of subjects with evaluable samples at a follow-up period of about 12 months after the infusion of the CAR-T cells. -5 At a sensitivity threshold level of about 10 to about 83% in subjects with evaluable samples, or at a follow-up period of about 18 months after the infusion of the CAR-T cells, -5 At a sensitivity threshold level of about 100%, the method effectively obtains a minimal residual disease (MRD) negative status in about 82% to about 97% of subjects with evaluable samples. In some embodiments, the method effectively obtains a minimal residual disease (MRD) negative status in about 100% to about 97% of subjects with evaluable samples at a follow-up period of about 12 months after the infusion of the CAR-T cells. -5 At a sensitivity threshold level of 100%, in subjects with evaluable samples, a rate of about 93%, or at a follow-up period of about 18 months after the infusion of the CAR-T cells, -5 At this sensitivity threshold level, approximately a 92% rate of subjects with evaluable samples effectively achieves the minimal residual disease (MRD) negative status.

[0194] In some embodiments, the method is effective to obtain at least one response in the subject following said infusion of the CAR-T cells, wherein said at least one response comprises, in order from best to worst, a stringent complete response, a complete response, a best partial response, a partial response, or a minimal response.

[0195] In some embodiments, the method is effective to obtain an initial response about 27 days or more, about 29 days or more, about 42 days or more, about 89 days or more, or about 321 days or more after the infusion of the CAR-T cells. In some embodiments, the method is effective to obtain an initial response before about 27 days to about 321 days after the infusion of the CAR-T cells. In some embodiments, the method is effective to obtain an initial response before about 27 days to about 89 days after the infusion of the CAR-T cells. In some embodiments, the method is effective to obtain an initial response before about 42 days after the infusion of the CAR-T cells. In some embodiments, the method is effective to obtain an initial response before about 29 days after the infusion of the CAR-T cells.

[0196] In some embodiments, the effectiveness of the treatment method is assessed by assessing the proportion of subjects having a stringent complete response. In some embodiments, the effectiveness of the treatment method is assessed by assessing the proportion of subjects having a complete response or better. In some embodiments, the effectiveness of the treatment method is assessed by assessing the proportion of subjects having a best partial response or better. In some embodiments, the effectiveness of the treatment method is assessed by assessing the proportion of subjects having a partial response or better. In some embodiments, the effectiveness of the treatment method is assessed by assessing the proportion of subjects having a minimal response or better.

[0197] In some embodiments, the method effectively obtains a best response of any one of minimal response, partial response, best partial response, complete response, or stringent complete response, i.e., a best response of minimal response or better. In some embodiments, the rate at which the method effectively obtains a best response of minimal response or better is referred to as the clinical benefit rate. In some examples, the method effectively obtains the best response of a minimal response or better at about 91% or less at about 12 months follow-up period after the infusion of the CAR-T cells, at about 97% or less at about 12 months follow-up period after the infusion of the CAR-T cells, at about 99% or less at about 12 months follow-up period after the infusion of the CAR-T cells, at about 93% or less at about 18 months follow-up period after the infusion of the CAR-T cells, at about 98% or less at about 18 months follow-up period after the infusion of the CAR-T cells, or at about 100% or less at about 18 months follow-up period after the infusion of the CAR-T cells. In some embodiments, the method is effective to obtain the best response of any one of minimal response, partial response, best partial response, complete response, or stringent complete response at about a 91% to about 99% rate at about a 12 month follow-up period after the infusion of the CAR-T cells, or at about a 93% to about 100% rate at about an 18 month follow-up period after the infusion of the CAR-T cells. In some embodiments, the method is effective to obtain the best response of any one of minimal response, partial response, best partial response, complete response, or stringent complete response at about a 97% rate at about a 12 month follow-up period after the infusion of the CAR-T cells, or at about a 98% rate at about an 18 month follow-up period after the infusion of the CAR-T cells.

[0198] In some embodiments, the method effectively achieves a best response of any one of partial response, best partial response, complete response, or stringent complete response, i.e., a best response of partial response or better. In some embodiments, the rate at which the method effectively achieves a best response of partial response or better is referred to as the overall survival rate or overall response rate. In some examples, the methods are effective to obtain a best response of a partial response or better at about 91% or less at about 12 months follow-up period after the infusion of the CAR-T cells, at about 97% or less at about 12 months follow-up period after the infusion of the CAR-T cells, at about 99% or less at about 12 months follow-up period after the infusion of the CAR-T cells, at about 93% or less at about 18 months follow-up period after the infusion of the CAR-T cells, at about 97% or less at about 18 months follow-up period after the infusion of the CAR-T cells, or at about 100% or less at about 18 months follow-up period after the infusion of the CAR-T cells. In some embodiments, the method is effective to obtain the best response of any one of partial response, best partial response, complete response, or stringent complete response at about a 91% to about 99% rate at about a 12 month follow-up period after the infusion of the CAR-T cells, or at about a 93% to about 100% rate at about an 18 month follow-up period after the infusion of the CAR-T cells. In some embodiments, the method is effective to obtain the best response of any one of partial response, best partial response, complete response, or stringent complete response at about a 97% rate at about a 12 month follow-up period after the infusion of the CAR-T cells, or at about a 97% rate at about an 18 month follow-up period after the infusion of the CAR-T cells.

[0199] In some embodiments, the method effectively obtains a best response of any one of a best partial response, a complete response, or a stringent complete response, i.e., a best partial response or better. In some embodiments, the method effectively obtains the best response of a best partial response or better in about 86% or less at about 12 months follow-up period after the infusion of the CAR-T cells, about 93% or less at about 12 months follow-up period after the infusion of the CAR-T cells, about 97% or less at about 12 months follow-up period after the infusion of the CAR-T cells, about 88% or less at about 18 months follow-up period after the infusion of the CAR-T cells, about 95% or less at about 18 months follow-up period after the infusion of the CAR-T cells, or about 98% or less at about 18 months follow-up period after the infusion of the CAR-T cells. In some embodiments, the method is effective to obtain the best response, either a best partial response, a complete response, or a stringent complete response, at about an 86% to about 97% rate at about a 12 month follow-up period after the infusion of the CAR-T cells, or at about an 88% to about 98% rate at about an 18 month follow-up period after the infusion of the CAR-T cells. In some embodiments, the method is effective to obtain the best response, either a best partial response, a complete response, or a stringent complete response, at about an 93% rate at about a 12 month follow-up period after the infusion of the CAR-T cells, or at about an 95% rate at about an 18 month follow-up period after the infusion of the CAR-T cells.

[0200] In some embodiments, the method effectively obtains a best response of a complete response or a stringent complete response, i.e., a best response of a complete response or better. In some embodiments, the method effectively obtains the best response of a complete response or better in about 57% or less at about 12 months follow-up period after the infusion of the CAR-T cells, about 67% or less at about 12 months follow-up period after the infusion of the CAR-T cells, about 76% or less at about 12 months follow-up period after the infusion of the CAR-T cells, about 73% or less at about 18 months follow-up period after the infusion of the CAR-T cells, about 83% or less at about 18 months follow-up period after the infusion of the CAR-T cells, or about 89% or less at about 18 months follow-up period after the infusion of the CAR-T cells. In some embodiments, the method is effective to obtain the best response of complete response or stringent complete response at about a 57% to about a 76% rate at about a 12 month follow-up period after the infusion of the CAR-T cells, or at about a 73% to about a 89% rate at about a 18 month follow-up period after the infusion of the CAR-T cells. In some embodiments, the method is effective to obtain the best response of complete response or stringent complete response at about a 67% rate at about a 12 month follow-up period after the infusion of the CAR-T cells, or at about a 83% rate at about a 18 month follow-up period after the infusion of the CAR-T cells.

[0201] In some embodiments, the method is effective to obtain a best response of stringent complete response. In some embodiments, the method is effective to obtain the best response of stringent complete response in about 57% or less at about 12 months follow-up period after the infusion of the CAR-T cells, in about 67% or less at about 12 months follow-up period after the infusion of the CAR-T cells, in about 76% or less at about 12 months follow-up period after the infusion of the CAR-T cells, in about 73% or less at about 18 months follow-up period after the infusion of the CAR-T cells, in about 83% or less at about 18 months follow-up period after the infusion of the CAR-T cells, or in about 89% or less at about 18 months follow-up period after the infusion of the CAR-T cells. In some embodiments, the method is effective to obtain the best response of stringent complete response at about a 57% to about a 76% rate at about a 12 month follow-up period after the infusion of the CAR-T cells, or at about a 73% to about a 89% rate at about a 18 month follow-up period after the infusion of the CAR-T cells. In some embodiments, the method is effective to obtain the best response of stringent complete response at about a 67% rate at about a 12 month follow-up period after the infusion of the CAR-T cells, or at about a 83% rate at about a 18 month follow-up period after the infusion of the CAR-T cells.

[0202] In some embodiments, the method is effective to obtain the best response at about 27 days or more, 78 days or more, 153 days or more, 293 days or more, or about 534 days or more after the infusion of the CAR-T cells. In some embodiments, the method is effective to obtain the best response at about 27 days to about 534 days after the infusion of the CAR-T cells. In some embodiments, the method is effective to obtain the best response at about 27 days to about 293 days after the infusion of the CAR-T cells. In some embodiments, the method is effective to obtain the best response at about 153 days after the infusion of the CAR-T cells. In some embodiments, the method is effective to obtain the best response at about 78 days after the infusion of the CAR-T cells.

[0203] In some examples, the method effectively maintains a response in the subject for a follow-up period from the time of the initial response to about 180 days after the infusion of the CAR-T cells, from the time of the initial response to about 357 days after the infusion of the CAR-T cells, from the time of the initial response to about 606 days after the infusion of the CAR-T cells, or from the time of the initial response to about 654 days after the infusion of the CAR-T cells. In some embodiments, the method comprises the step of: at about 77% or less at about a 6 month follow-up period after the infusion of the CAR-T cells; at about 85% or less at about a 6 month follow-up period after the infusion of the CAR-T cells; at about 91% or less at about a 6 month follow-up period after the infusion of the CAR-T cells; at about 63% or less at about a 12 month follow-up period after the infusion of the CAR-T cells; at about 74% or less at about a 12 month follow-up period after the infusion of the CAR-T cells; at about 81% or less at about a 12 month follow-up period after the infusion of the CAR-T cells; at about 56% or less at about a 18 month follow-up period after the infusion of the CAR-T cells; at about 67% or less at about 18 months follow-up period after the infusion of the CAR-T cells, at about 75% or less at about 18 months follow-up period after the infusion of the CAR-T cells, at about 52% or less at about 21 months follow-up period after the infusion of the CAR-T cells, at about 63% or less at about 21 months follow-up period after the infusion of the CAR-T cells, at about 72% or less at about 21 months follow-up period after the infusion of the CAR-T cells, at about 48% or less at about 24 months follow-up period after the infusion of the CAR-T cells, at about 60% or less at about 24 months follow-up period after the infusion of the CAR-T cells, or at about 70% or less at about 24 months follow-up period after the infusion of the CAR-T cells.In some examples, the methods effectively maintain a response rate of about 77% to about 91% at about a 6 month follow-up period after the infusion of the CAR-T cells, about 63% to about 81% at about a 12 month follow-up period after the infusion of the CAR-T cells, about 56% to about 75% at about an 18 month follow-up period after the infusion of the CAR-T cells, about 52% to about 72% at about a 21 month follow-up period after the infusion of the CAR-T cells, or about 48% to about 70% at about a 24 month follow-up period after the infusion of the CAR-T cells. In some examples, the methods effectively maintain response at about an 85% rate at about a 6 month follow-up period after the infusion of the CAR-T cells, at about a 74% rate at about a 12 month follow-up period after the infusion of the CAR-T cells, at about a 67% rate at about an 18 month follow-up period after the infusion of the CAR-T cells, at about a 63% rate at about a 21 month follow-up period after the infusion of the CAR-T cells, or at about a 60% rate at about a 24 month follow-up period after the infusion of the CAR-T cells.

[0204] In some embodiments, the method comprises administering 10 to about 3 months after administration of the CAR-T cells. -5at about 12 months follow-up period after the infusion of the CAR-T cells, about 34% or less, at about 12 months follow-up period after the infusion of the CAR-T cells, about 44% or less, at about 12 months follow-up period after the infusion of the CAR-T cells, about 33% or less, at about 18 months follow-up period after the infusion of the CAR-T cells, about 43% or less, or about 54% or less, at about 18 months follow-up period after the infusion of the CAR-T cells. In some embodiments, the method is effective to obtain a minimal residual disease (MRD) negative complete response or a minimal residual disease (MRD) negative stringent complete response at a rate of about 25% to about 44% at about a 12 month follow-up period after the infusion of the CAR-T cells, or at a rate of about 33% to about 54% at about an 18 month follow-up period after the infusion of the CAR-T cells. In some embodiments, the method is effective to obtain a minimal residual disease (MRD) negative complete response or a minimal residual disease (MRD) negative stringent complete response at a rate of about 34% at about a 12 month follow-up period after the infusion of the CAR-T cells, or at a rate of about 43% at about an 18 month follow-up period after the infusion of the CAR-T cells.

[0205] In some embodiments, the method effectively obtains progression free survival in a subject. In some embodiments, the method effectively obtains progression free survival in a subject between about 209 days after the infusion of the CAR-T cells, between about 386 days after the infusion of the CAR-T cells, between about 632 days after the infusion of the CAR-T cells, or between about 684 days after the infusion of the CAR-T cells. In some embodiments, the method comprises the step of: at about 79% or less at about a 6 month follow-up period after the infusion of the CAR-T cells; at about 88% or less at about a 6 month follow-up period after the infusion of the CAR-T cells; at about 93% or less at about a 6 month follow-up period after the infusion of the CAR-T cells; at about 67% or less at about a 12 month follow-up period after the infusion of the CAR-T cells; at about 76% or less at about a 12 month follow-up period after the infusion of the CAR-T cells; at about 84% or less at about a 12 month follow-up period after the infusion of the CAR-T cells; at about 57% or less at about a 18 month follow-up period after the infusion of the CAR-T cells; at about 67% or less at about 18 months follow-up period after the infusion of the CAR-T cells, at about 75% or less at about 18 months follow-up period after the infusion of the CAR-T cells, at about 57% or less at about 21 months follow-up period after the infusion of the CAR-T cells, at about 67% or less at about 21 months follow-up period after the infusion of the CAR-T cells, at about 75% or less at about 21 months follow-up period after the infusion of the CAR-T cells, at about 49% or less at about 24 months follow-up period after the infusion of the CAR-T cells, at about 61% or less at about 24 months follow-up period after the infusion of the CAR-T cells, or at about 70% or less at about 24 months follow-up period after the infusion of the CAR-T cells.In some embodiments, the method effectively obtains a progression free survival rate of about 79% to about 93% at about a 6 month follow-up period after the infusion of the CAR-T cells, about 67% to about 84% at about a 12 month follow-up period after the infusion of the CAR-T cells, about 57% to about 75% at about an 18 month follow-up period after the infusion of the CAR-T cells, about 57% to about 75% at about a 21 month follow-up period after the infusion of the CAR-T cells, or about 49% to about 70% at about a 24 month follow-up period after the infusion of the CAR-T cells. In some examples, the method effectively obtains the progression free survival at about an 88% rate at about a 6 month follow-up period after the infusion of the CAR-T cells, at about a 76% rate at about a 12 month follow-up period after the infusion of the CAR-T cells, at about a 67% rate at about an 18 month follow-up period after the infusion of the CAR-T cells, at about a 67% rate at about a 21 month follow-up period after the infusion of the CAR-T cells, or at about a 61% rate at about a 24 month follow-up period after the infusion of the CAR-T cells.

[0206] In some embodiments, the method further comprises treating cytokine release syndrome in the subject more than about 1 day after said infusion of said CAR-T cells, hi some embodiments, the method effectively obtains about a 1% to about 99% reversal rate of said cytokine release syndrome within a time period of about 1, 3, 4, 6, 16, or 97 days after said cytokine release syndrome is first observed.

[0207] In some embodiments, the method further comprises treating immune effector cell associated neurotoxicity in the subject more than about 3 days after the infusion of the CAR-T cells. In some embodiments, the method effectively obtains about a 1% to about 17% reversal rate of immune effector cell associated neurotoxicity in a time period of about 1, 4, 5, 8, 12, or 16 days after first observing immune effector cell associated neurotoxicity.

[0208] In some embodiments, the treatment method is effective to obtain a reduced tumor burden in the subject. In some embodiments, the treatment method is effective to obtain a reduced tumor burden in greater than 90% of the subjects. In some embodiments, the treatment method is effective to obtain a reduced tumor burden in greater than 91% of the subjects. In some embodiments, the treatment method is effective to obtain a reduced tumor burden in greater than 92% of the subjects. In some embodiments, the treatment method is effective to obtain a reduced tumor burden in greater than 93% of the subjects. In some embodiments, the treatment method is effective to obtain a reduced tumor burden in greater than 94% of the subjects. In some embodiments, the treatment method is effective to obtain a reduced tumor burden in greater than 95% of the subjects. In some embodiments, the treatment method is effective to obtain a reduced tumor burden in greater than 96% of the subjects. In some embodiments, the treatment method is effective to obtain a reduced tumor burden in greater than 97% of the subjects. In some embodiments, the treatment method is effective to obtain a reduced tumor burden in greater than 98% of the subjects. In some embodiments, the treatment method is effective to obtain a reduced tumor burden in greater than 99% of subjects, hi some embodiments, the treatment method is effective to obtain a reduced tumor burden in 100% of subjects.

[0209] In some embodiments, the method of treatment effectively obtains a minimal residual disease (MRD) negative status or effectively maintains the minimal residual disease (MRD) status in the subject. In some embodiments, the method of treatment effectively obtains a minimal residual disease (MRD) negative status in the subject. In some embodiments, the method of treatment effectively obtains a minimal residual disease (MRD) negative status in the subject. In some embodiments, the method of treatment effectively obtains a minimal residual disease (MRD) negative status in the subject. -6 In some embodiments, the method of treatment effectively achieves a negative minimal residual disease (MRD) status in the subject at a sensitivity level of 10 -5 In some embodiments, the method of treatment effectively achieves a negative minimal residual disease (MRD) status in the subject at a sensitivity level of 10 -4In some embodiments, the method of treatment effectively achieves a negative minimal residual disease (MRD) status in the subject at a sensitivity level of 10 -3 In some embodiments, the method of treatment effectively obtains a negative minimal residual disease (MRD) status in the subject when assessed in bone marrow. In some embodiments, the method of treatment effectively maintains a negative MRD status when assessed in an assessable bone marrow sample. In some embodiments, the method of treatment effectively obtains a negative MRD status when assessed in bone marrow DNA. In some embodiments, the method of treatment effectively obtains a negative MRD status when assessed at a follow-up period of 28 days or more after administration of the CAR-T cells. In some embodiments, the method of treatment effectively obtains a negative MRD status when assessed at a follow-up period of 1 month or more after administration of the CAR-T cells. In some embodiments, the method of treatment effectively obtains a negative MRD status when assessed at a follow-up period of 3 months or more after administration of the CAR-T cells. In some embodiments, the method of treatment effectively obtains a negative MRD status when assessed at a follow-up period of 6 months or more after administration of the CAR-T cells. In some embodiments, the treatment method effectively achieves MRD negativity when assessed at a follow-up period of 9 months or more after administration of the CAR-T cells. In some embodiments, the treatment method effectively achieves MRD negativity when assessed at a follow-up period of 12 months or more after administration of the CAR-T cells.

[0210] In some embodiments, the treatment method effectively maintains minimal residual disease (MRD) negativity initially achieved in the subject. -5 In some embodiments, the treatment method effectively maintains MRD negativity at a sensitivity level of 10 -6 In some embodiments, the method of treatment effectively achieves a negative minimal residual disease (MRD) status in the subject at a sensitivity level of 10 -4 In some embodiments, the treatment method effectively maintains MRD negativity at a sensitivity level of 10 -3In some embodiments, the method of treatment effectively maintains MRD negativity when assessed with a bone marrow sample. In some embodiments, the method of treatment effectively maintains MRD negativity when assessed with an evaluable bone marrow sample. In some embodiments, the method of treatment effectively maintains MRD negativity when assessed with bone marrow DNA. In some embodiments, the method of treatment effectively maintains MRD negativity when assessed with a one month or greater follow-up period after administration of the CAR-T cells. In some embodiments, the method of treatment effectively maintains MRD negativity when assessed with a three month or greater follow-up period after administration of the CAR-T cells. In some embodiments, the method of treatment effectively maintains MRD negativity when assessed with a six month or greater follow-up period after administration of the CAR-T cells. In some embodiments, the method of treatment effectively maintains MRD negativity when assessed with a nine month or greater follow-up period after administration of the CAR-T cells. In some embodiments, the treatment method effectively maintains MRD negativity when assessed over a follow-up period of 12 months or more following administration of the CAR-T cells.

[0211] In some embodiments, the efficacy of the treatment method is assessed by evaluating the proportion of subjects with MRD negative status. -6 The efficacy of the treatment method is assessed by assessing the proportion of subjects with MRD negative status at a sensitivity level of 10 -5 The efficacy of the treatment method is assessed by assessing the proportion of subjects with MRD negative status at a sensitivity level of 10 -4 The efficacy of the treatment method is assessed by assessing the proportion of subjects with MRD negative status at a sensitivity level of 10 -3The efficacy of the treatment method is assessed by assessing the proportion of subjects with MRD negative status at a median follow-up period of 28 days or more after administration of the CAR-T cells. In some embodiments, the efficacy of the treatment method is assessed by assessing the proportion of subjects with MRD negative status at a median follow-up period of 1 month or more after administration of the CAR-T cells. In some embodiments, the efficacy of the treatment method is assessed by assessing the proportion of subjects with MRD negative status at a median follow-up period of 3 months or more after administration of the CAR-T cells. In some embodiments, the efficacy of the treatment method is assessed by assessing the proportion of subjects with MRD negative status at a median follow-up period of 6 months or more after administration of the CAR-T cells. In some embodiments, the efficacy of the treatment method is assessed by assessing the proportion of subjects with MRD negative status at a median follow-up period of 9 months or more after administration of the CAR-T cells. In some examples, the efficacy of the treatment method is assessed by evaluating the proportion of subjects with MRD-negative status at a median follow-up period of 12 months or more after administration of the CAR-T cells.

[0212] In some embodiments, the efficacy of the treatment method is assessed by evaluating the proportion of subjects with evaluable bone marrow and negative MRD status. -6 The efficacy of the treatment method is assessed by assessing the proportion of subjects with evaluable bone marrow and MRD negative status at a sensitivity level of 10 -5 The efficacy of the treatment method is assessed by assessing the proportion of subjects with evaluable bone marrow and MRD negative status at a sensitivity level of 10 -4 The efficacy of the treatment method is assessed by assessing the proportion of subjects with evaluable bone marrow and MRD negative status at a sensitivity level of 10 -3The efficacy of the treatment method is assessed by assessing the proportion of subjects with evaluable bone marrow and MRD negative status at a median follow-up of 28 days or more after administration of the CAR-T cells. In some embodiments, the efficacy of the treatment method is assessed by assessing the proportion of subjects with evaluable bone marrow and MRD negative status at a median follow-up of 1 month or more after administration of the CAR-T cells. In some embodiments, the efficacy of the treatment method is assessed by assessing the proportion of subjects with evaluable bone marrow and MRD negative status at a median follow-up of 3 months or more after administration of the CAR-T cells. In some embodiments, the efficacy of the treatment method is assessed by assessing the proportion of subjects with evaluable bone marrow and MRD negative status at a median follow-up of 6 months or more after administration of the CAR-T cells. In some embodiments, the efficacy of the treatment method is assessed by evaluating the proportion of subjects with evaluable bone marrow and MRD negative status at a median follow-up of at least 9 months after administration of the CAR T cells. In some embodiments, the efficacy of the treatment method is assessed by evaluating the proportion of subjects with evaluable bone marrow and MRD negative status at a median follow-up of at least 12 months after administration of the CAR T cells.

[0213] In some embodiments, the effectiveness of the treatment method is assessed by assessing the proportion of subjects having a robust complete response. In some embodiments, the effectiveness of the treatment method is assessed by assessing the proportion of subjects having a complete response or better. In some embodiments, the effectiveness of the treatment method is assessed by assessing the proportion of subjects having a best partial response or better. In some embodiments, the effectiveness of the treatment method is assessed by assessing the proportion of subjects having a partial response or better. In some embodiments, the overall response rate is used to assess the effectiveness of the treatment method. In some embodiments, the overall response rate is the proportion of subjects having a partial response or better.

[0214] In some embodiments, the method comprises: -5 At a sensitivity threshold level of 100 μg / mL, the method effectively obtains a minimal residual disease (MRD) negativity rate of greater than 39%. -5 At a sensitivity threshold level of 100 μg / mL, the method effectively obtains a minimal residual disease (MRD) negativity rate of greater than 44%. -5 At a sensitivity threshold level of 100 μg / mL, the method effectively obtains a minimal residual disease (MRD) negativity rate of greater than 49%. -5 At a sensitivity threshold level of 100 μg / mL, the method effectively achieves a minimal residual disease (MRD) negativity rate of greater than 54%. -5 At a sensitivity threshold level of 10, the method effectively obtains a minimal residual disease (MRD) negativity rate of greater than 59%. -5 At a sensitivity threshold level of 100 μg / mL, the method effectively achieves a minimal residual disease (MRD) negativity rate of greater than 64%. -5 At a sensitivity threshold level of 100 μg / mL, the method effectively achieves a minimal residual disease (MRD) negativity rate of greater than 69%. -5 At a sensitivity threshold level of 100 μg / mL, the method effectively achieves a minimal residual disease (MRD) negativity rate of greater than 74%. -5 At a sensitivity threshold level of 100 μg / mL, the method effectively obtains a minimal residual disease (MRD) negativity rate of greater than 70%. In some embodiments, the method effectively obtains a minimal residual disease (MRD) negativity rate of greater than 70% in assessable bone marrow. -5 At a sensitivity threshold level of 100 μg / mL, the method effectively obtains a minimal residual disease (MRD) negativity rate of greater than 75%. In some embodiments, the method effectively obtains a minimal residual disease (MRD) negativity rate of greater than 75% in assessable bone marrow. -5 At a sensitivity threshold level of 100 μg / mL, the method effectively obtains a minimal residual disease (MRD) negativity rate of greater than 80%. In some embodiments, the method effectively obtains a minimal residual disease (MRD) negativity rate of greater than 80% in assessable bone marrow. -5 At a sensitivity threshold level of 100 μg / mL, the method effectively achieves a minimal residual disease (MRD) negativity rate of greater than 85%. In some embodiments, the method achieves a minimal residual disease (MRD) negativity rate of greater than 85% in assessable bone marrow. -5At a sensitivity threshold level of 100 μg / mL, the method effectively achieves a minimal residual disease (MRD) negativity rate of greater than 90%. In some embodiments, the method achieves a minimal residual disease (MRD) negativity rate of greater than 90% in assessable bone marrow. -5 At a sensitivity threshold level of 100 μg / mL, the method effectively achieves a minimal residual disease (MRD) negativity rate of greater than 95%. In some embodiments, the method achieves a minimal residual disease (MRD) negativity rate of greater than 95% in assessable bone marrow. -5 At this sensitivity threshold level, a 100% minimal residual disease (MRD) negativity rate is effectively obtained.

[0215] In some embodiments, the method of treatment effectively obtains an overall response rate of greater than 90%. In some embodiments, the method of treatment effectively obtains an overall response rate of greater than 91%. In some embodiments, the method effectively obtains an overall response rate of greater than 93%. In some embodiments, the method effectively obtains an overall response rate of greater than 95%. In some embodiments, the method effectively obtains an overall response rate of greater than 97%. In some embodiments, the method effectively obtains an overall response rate of greater than 99%. In some embodiments, the method effectively obtains an overall response rate of 100%. In some embodiments, the overall response rate is assessed at a median follow-up period of at least 6 months after said infusion of said CAR-T cells. In some embodiments, the overall response rate is assessed at a median follow-up period of at least 12 months after said infusion of said CAR-T cells.

[0216] In some embodiments, greater than 70% of subjects respond to the treatment method 9 months after administration of the CAR-T cells. In some embodiments, greater than 72% of subjects respond to the treatment method 9 months after administration of the CAR-T cells. In some embodiments, greater than 74% of subjects respond to the treatment method 9 months after administration of the CAR-T cells. In some embodiments, greater than 76% of subjects respond to the treatment method 9 months after administration of the CAR-T cells. In some embodiments, greater than 78% of subjects respond to the treatment method 9 months after administration of the CAR-T cells. In some embodiments, greater than 80% of subjects respond to the treatment method 9 months after administration of the CAR-T cells. In some embodiments, greater than 82% of subjects respond to the treatment method 9 months after administration of the CAR-T cells. In some embodiments, greater than 84% of subjects respond to the treatment method 9 months after administration of the CAR-T cells. In some examples, greater than 86% of subjects respond to the treatment regimen 9 months after administration of the CAR-T cells.

[0217] In some embodiments, greater than 54% of responding subjects respond to the treatment method 12 months after administration of the CAR-T cells. In some embodiments, greater than 58% of responding subjects respond to the treatment method 12 months after administration of the CAR-T cells. In some embodiments, greater than 62% of responding subjects respond to the treatment method 12 months after administration of the CAR-T cells. In some embodiments, greater than 66% of responding subjects respond to the treatment method 12 months after administration of the CAR-T cells. In some embodiments, greater than 70% of responding subjects respond to the treatment method 12 months after administration of the CAR-T cells. In some embodiments, greater than 74% of responding subjects respond to the treatment method 12 months after administration of the CAR-T cells. In some embodiments, greater than 78% of responding subjects respond to the treatment method 12 months after administration of the CAR-T cells.

[0218] In some embodiments, the treatment methods are effective in providing a duration of response greater than 9 months, 10 months, 11 months, 12 months, 13 months, 14 months, or 15 months. In some embodiments, the treatment methods are effective in providing a duration of response greater than 12.4 months. In some embodiments, the treatment methods are effective in providing a duration of response greater than 15.9 months.

[0219] In some embodiments, the treatment methods are effective to obtain a median duration of response of greater than 9 months, 10 months, 11 months, 12 months, 13 months, 14 months, or 15 months. In some embodiments, the treatment methods are effective to obtain a median duration of response of greater than 12.4 months. In some embodiments, the treatment methods are effective to obtain a median duration of response of greater than 15.9 months.

[0220] In some embodiments, the treatment method is effective in obtaining a complete response or better in greater than 60% of subjects. In some embodiments, the treatment method is effective in obtaining a complete response or better in greater than 61% of subjects. In some embodiments, the treatment method is effective in obtaining a complete response or better in greater than 62% of subjects. In some embodiments, the treatment method is effective in obtaining a complete response or better in greater than 63% of subjects. In some embodiments, the treatment method is effective in obtaining a complete response or better in greater than 64% of subjects. In some embodiments, the treatment method is effective in obtaining a complete response or better in greater than 65% of subjects. In some embodiments, the treatment method is effective in obtaining a complete response or better in greater than 66% of subjects. In some embodiments, the treatment method is effective in obtaining a complete response or better in greater than 67% of subjects. In some examples, a complete response or better is assessed less than one month after administration of the CAR-T cells. In some examples, a complete response or better is assessed less than three months after administration of the CAR-T cells. In some examples, a complete response or better is assessed less than six months after administration of the CAR-T cells. In some examples, a complete response or better is assessed less than nine months after administration of the CAR-T cells. In some examples, a complete response or better is assessed less than 12 months after administration of the CAR-T cells. In some examples, a complete response or better is assessed less than 15 months after administration of the CAR-T cells. In some examples, a complete response or better is assessed more than 15 months after administration of the CAR-T cells.

[0221] In some embodiments, the treatment method is effective in obtaining a best partial response or better in greater than 85% of subjects. In some embodiments, the treatment method is effective in obtaining a best partial response or better in greater than 86% of subjects. In some embodiments, the treatment method is effective in obtaining a best partial response or better in greater than 87% of subjects. In some embodiments, the treatment method is effective in obtaining a best partial response or better in greater than 88% of subjects. In some embodiments, the treatment method is effective in obtaining a best partial response or better in greater than 89% of subjects. In some embodiments, the treatment method is effective in obtaining a best partial response or better in greater than 90% of subjects. In some embodiments, the treatment method is effective in obtaining a best partial response or better in greater than 91% of subjects. In some embodiments, the treatment method is effective to obtain a best partial response or better in greater than 92% of subjects. In some embodiments, the best partial response or better is assessed less than 1 month after administration of the CAR-T cells. In some embodiments, the best partial response or better is assessed less than 3 months after administration of the CAR-T cells. In some embodiments, the best partial response or better is assessed less than 6 months after administration of the CAR-T cells. In some embodiments, the best partial response or better is assessed less than 9 months after administration of the CAR-T cells. In some embodiments, the best partial response or better is assessed less than 12 months after administration of the CAR-T cells. In some embodiments, the best partial response or better is assessed less than 15 months after administration of the CAR-T cells. In some embodiments, the best partial response or better is assessed more than 15 months after administration of the CAR-T cells.

[0222] In some embodiments, the method of treatment is effective to obtain a median duration of initial response of less than 1.15 months. In some embodiments, the method of treatment is effective to obtain a median duration of initial response of less than 1.10 months. In some embodiments, the method of treatment is effective to obtain a median duration of initial response of less than 1.05 months. In some embodiments, the method of treatment is effective to obtain a median duration of initial response of less than 1.00 months. In some embodiments, the method of treatment is effective to obtain a median duration of initial response of less than 0.95 months.

[0223] In some embodiments, the method of treatment is effective to obtain a median duration of best response of less than 2.96 months. In some embodiments, the method of treatment is effective to obtain a median duration of best response of less than 2.86 months. In some embodiments, the method of treatment is effective to obtain a median duration of best response of less than 2.76 months. In some embodiments, the method of treatment is effective to obtain a median duration of best response of less than 2.66 months. In some embodiments, the method of treatment is effective to obtain a median duration of best response of less than 2.56 months.

[0224] In some embodiments, the method effectively obtains an overall survival rate of greater than 80% at 9 months after administration of the CAR-T cells. In some embodiments, the method effectively obtains an overall survival rate of greater than 82% at 9 months after administration of the CAR-T cells. In some embodiments, the method effectively obtains an overall survival rate of greater than 85% at 9 months after administration of the CAR-T cells. In some embodiments, the method effectively obtains an overall survival rate of greater than 87% at 9 months after administration of the CAR-T cells. In some embodiments, the method effectively obtains an overall survival rate of greater than 90% at 9 months after administration of the CAR-T cells. In some embodiments, the method effectively obtains an overall survival rate of greater than 92% at 9 months after administration of the CAR-T cells. In some embodiments, the method effectively obtains an overall survival rate of greater than 95% at 9 months after administration of the CAR-T cells.

[0225] In some embodiments, the method effectively obtains an overall survival rate of greater than 80% at 12 months after administration of the CAR-T cells. In some embodiments, the method effectively obtains an overall survival rate of greater than 83% at 12 months after administration of the CAR-T cells. In some embodiments, the method effectively obtains an overall survival rate of greater than 86% at 12 months after administration of the CAR-T cells. In some embodiments, the method effectively obtains an overall survival rate of greater than 89% at 12 months after administration of the CAR-T cells. In some embodiments, the method effectively obtains an overall survival rate of greater than 92% at 12 months after administration of the CAR-T cells. In some embodiments, the method effectively obtains an overall survival rate of greater than 93% at 12 months after administration of the CAR-T cells.

[0226] In some embodiments, the method effectively obtains a progression-free survival rate of greater than 70% at 9 months after administration of the CAR-T cells. In some embodiments, the method effectively obtains a progression-free survival rate of greater than 72% at 9 months after administration of the CAR-T cells. In some embodiments, the method effectively obtains a progression-free survival rate of greater than 75% at 9 months after administration of the CAR-T cells. In some embodiments, the method effectively obtains a progression-free survival rate of greater than 77% at 9 months after administration of the CAR-T cells. In some embodiments, the method effectively obtains a progression-free survival rate of greater than 80% at 9 months after administration of the CAR-T cells. In some embodiments, the method effectively obtains a progression-free survival rate of greater than 82% at 9 months after administration of the CAR-T cells. In some embodiments, the method effectively obtains a progression-free survival rate of greater than 85% at 9 months after administration of the CAR-T cells. In some embodiments, the method is effective to obtain a progression-free survival rate of 87% or more at 9 months after administration of the CAR-T cells.

[0227] In some embodiments, the method effectively obtains a progression-free survival rate of greater than 66% at 12 months after administration of the CAR-T cells. In some embodiments, the method effectively obtains a progression-free survival rate of greater than 69% at 12 months after administration of the CAR-T cells. In some embodiments, the method effectively obtains a progression-free survival rate of greater than 72% at 12 months after administration of the CAR-T cells. In some embodiments, the method effectively obtains a progression-free survival rate of greater than 76% at 12 months after administration of the CAR-T cells. In some embodiments, the method effectively obtains a progression-free survival rate of greater than 80% at 12 months after administration of the CAR-T cells. In some embodiments, the method effectively obtains a progression-free survival rate of greater than 84% at 12 months after administration of the CAR-T cells.

[0228] In some embodiments, the method of treatment effectively results in greater than 86% of subjects recovered from cytokine release syndrome. In some embodiments, the method of treatment effectively results in greater than 88% of subjects recovered from cytokine release syndrome. In some embodiments, the method of treatment effectively results in greater than 90% of subjects recovered from cytokine release syndrome. In some embodiments, the method of treatment effectively results in greater than 92% of subjects recovered from cytokine release syndrome. In some embodiments, the method of treatment effectively results in greater than 94% of subjects recovered from cytokine release syndrome. In some embodiments, the method of treatment effectively results in greater than 96% of subjects recovered from cytokine release syndrome. In some embodiments, the method of treatment effectively results in greater than 98% of subjects recovered from cytokine release syndrome. In some embodiments, the method of treatment effectively results in greater than 99% of subjects recovered from cytokine release syndrome. In some embodiments, the method of treatment effectively results in 100% of subjects recovered from cytokine release syndrome.

[0229] In some embodiments, the treatment method effectively results in greater than 90% of subjects recovered from immune effector cell associated neurotoxicity (if any). In some embodiments, the treatment method effectively results in greater than 92% of subjects recovered from immune effector cell associated neurotoxicity (if any). In some embodiments, the treatment method effectively results in greater than 94% of subjects recovered from immune effector cell associated neurotoxicity (if any). In some embodiments, the treatment method effectively results in greater than 96% of subjects recovered from immune effector cell associated neurotoxicity (if any). In some embodiments, the treatment method effectively results in greater than 98% of subjects recovered from immune effector cell associated neurotoxicity (if any). In some embodiments, the treatment method effectively results in 100% of subjects recovered from immune effector cell associated neurotoxicity (if any).

[0230] In some embodiments, the method further comprises diagnosing cytopenia in the subject. In some embodiments, the cytopenia comprises one or more or all of lymphopenia, neutropenia, and thrombocytopenia. Without being bound by theory, grade 3 or grade 4 lymphopenia, but not grade 2 or less, is considered to be a condition in which the lymphocyte count is greater than or equal to 0.5×10 9Grade 3 or 4 neutropenia, but not grade 2 or less, is characterized by a neutrophil count of less than 1000 cells / microliter of the subject's blood sample, and Grade 3 or 4 thrombocytopenia, but not grade 2 or less, is characterized by a platelet count of less than 50,000 cells / microliter of the subject's blood sample. In some examples, more than 75% of subjects suffering from Grade 3 or Grade 4 lymphopenia after administration of the CAR-T cells have recovered to Grade 2 or less lymphopenia at day 60 after administration of the CAR-T cells. In some examples, more than 80% of subjects suffering from Grade 3 or Grade 4 lymphopenia after administration of the CAR-T cells have recovered to Grade 2 or less lymphopenia at day 60 after administration of the CAR-T cells. In some examples, greater than 85% of subjects with grade 3 or grade 4 lymphopenia after administration of the CAR-T cells have recovered to grade 2 or less lymphopenia at day 60 after administration of the CAR-T cells. In some examples, greater than 90% of subjects with grade 3 or grade 4 lymphopenia after administration of the CAR-T cells have recovered to grade 2 or less lymphopenia at day 60 after administration of the CAR-T cells. In some examples, greater than 70% of subjects with grade 3 or grade 4 neutropenia after administration of the CAR-T cells have recovered to grade 2 or less neutropenia at day 60 after administration of the CAR-T cells. In some examples, greater than 75% of subjects with grade 3 or grade 4 neutropenia after administration of the CAR-T cells have recovered to grade 2 or less neutropenia at day 60 after administration of the CAR-T cells. In some examples, greater than 80% of subjects with grade 3 or grade 4 neutropenia after administration of the CAR-T cells have recovered to grade 2 or less neutropenia at day 60 after administration of the CAR-T cells. In some examples, greater than 85% of subjects with grade 3 or grade 4 neutropenia after administration of the CAR-T cells have recovered to grade 2 or less neutropenia at day 60 after administration of the CAR-T cells.In some embodiments, more than 30% of subjects with grade 3 or grade 4 thrombocytopenia after administration of the CAR-T cells have recovered to grade 2 or less thrombocytopenia at day 60 after administration of the CAR-T cells. In some embodiments, more than 34% of subjects with grade 3 or grade 4 thrombocytopenia after administration of the CAR-T cells have recovered to grade 2 or less thrombocytopenia at day 60 after administration of the CAR-T cells. In some embodiments, more than 38% of subjects with grade 3 or grade 4 thrombocytopenia after administration of the CAR-T cells have recovered to grade 2 or less thrombocytopenia at day 60 after administration of the CAR-T cells. In some embodiments, more than 42% of subjects with grade 3 or grade 4 thrombocytopenia after administration of the CAR-T cells have recovered to grade 2 or less thrombocytopenia at day 60 after administration of the CAR-T cells.

[0231] In some embodiments, the subject is re-treated by administration of a second dose of CAR-T cells via a second intravenous infusion. In some embodiments, the re-treatment dose is 1.0×10 5 From 5.0×10 6 In some embodiments, the retreatment dose comprises about 0.75×10 CAR-T cells / kilogram of subject mass. 5 In some embodiments, the subject is re-treated if the subject shows disease progression after a best response of minimal response or better response after the first infusion of CAR-T cells. In some embodiments, the time from the first infusion of CAR-T cells to detection of disease progression comprises at least six months.

[0232] Sets (kits) and products Any of the compositions described herein may be included in a set. In some examples, the engineered immortalized CAR-T cells are provided in a set, which may further include reagents suitable for expanding the cells, such as media.

[0233] In non-limiting examples, the components are a chimeric receptor expression construct, one or more reagents for producing the chimeric receptor expression construct, cells for transfecting the expression construct, and / or one or more instruments for obtaining immortalized T cells for transfecting the expression construct (such instruments may be a syringe, pipette, forceps, and / or any such medically approved device).

[0234] In some embodiments, the set comprises reagents or equipment for cell electroporation.

[0235] In some embodiments, the set comprises artificial antigen presenting cells.

[0236] The set may include one or more compositions of the present disclosure or reagents for producing compositions of the present disclosure, appropriately aliquoted. The components of the set may be packaged in aqueous media or lyophilized form. The container device of the set may include at least one vial, test tube, flask, bottle, syringe, or other container device into which the components can be placed, and preferably appropriately aliquoted. If there is more than one component in the set, the set will typically further contain a second, third, or other additional container, in which the additional components may be separately placed. However, various combinations of components may be included in the vial. The set of the present disclosure will typically further include a device for containing the chimeric receptor construct and any other reagent containers in a closed confine for commercial sale. Such containers may include, for example, injection-molded or blow-molded plastic containers into which the desired vials are held.

[0237] Approved Pharmaceutical Products The disclosure further provides a method of treating an adult patient with relapsed or refractory multiple myeloma, comprising administering to said patient an approved pharmaceutical product comprising siltacabtadine autorucel suspension in an amount and manner as described in the pharmaceutical product label of the approved pharmaceutical product and / or in the dosing and / or treatment regimen described herein.

[0238] The present disclosure further relates to a pharmaceutical product comprising siltacabtadine autorucel suspension, wherein said pharmaceutical product is packaged, and wherein said package comprises a label, said label identifying siltacabtadine autorucel suspension as an approved pharmaceutical product for treating adult patients with relapsed or refractory multiple myeloma after receiving four or more prior lines of therapy, wherein said prior lines of therapy include a proteasome inhibitor, an immunomodulatory agent, and an anti-CD38 monoclonal antibody.

[0239] The disclosure further provides a method of marketing an approved pharmaceutical product comprising siltacabtadine autorucel suspension, the method comprising marketing the approved pharmaceutical product, wherein the pharmaceutical product label of a reference product for the approved pharmaceutical product comprises instructions for treating patients with relapsed or refractory multiple myeloma after receiving four or more prior lines of therapy, the prior lines of therapy comprising a proteasome inhibitor, an immunomodulatory agent, and an anti-CD38 monoclonal antibody.

[0240] In some embodiments, the method of marketing includes a method of marketing a biosimilar of an approved pharmaceutical product comprising siltacabtadine autorucel suspension, the method including marketing the biosimilar, where the pharmaceutical product label of the biosimilar reference product comprises instructions for treating patients with relapsed or refractory multiple myeloma after receiving four or more prior lines of therapy, the prior lines of therapy including a proteasome inhibitor, an immunomodulatory agent, and an anti-CD38 monoclonal antibody.

[0241] The present disclosure further provides a method of authorizing the sale of a pharmaceutical product comprising siltacabtadine autorucel suspension, the method comprising authorizing the sale of such pharmaceutical product, wherein the pharmaceutical product label of a reference product for such pharmaceutical product comprises instructions for treating patients with relapsed or refractory multiple myeloma after receiving four or more prior lines of therapy, the prior lines of therapy comprising a proteasome inhibitor, an immunomodulatory agent, and an anti-CD38 monoclonal antibody.

[0242] In some embodiments, the method of authorizing marketing includes a method of authorizing marketing of a biosimilar of an approved drug comprising siltacabtadine autorucel suspension, the method comprising authorizing marketing of the biosimilar, wherein the pharmaceutical product label of the biosimilar reference product comprises instructions for treating patients with relapsed or refractory multiple myeloma after receiving four or more prior lines of therapy, the prior lines of therapy comprising a proteasome inhibitor, an immunomodulatory agent, and an anti-CD38 monoclonal antibody.

[0243] The terms "marketing" or "sale" as used herein refer to the transfer of a pharmaceutical product, e.g., a pharmaceutical composition or dosage form, from a seller to a buyer.

[0244] The term "license to sell" as used herein refers to a seller submitting to a buyer an offer to sell a pharmaceutical product, e.g., a pharmaceutical composition or dosage form. These processes include licensing a pharmaceutical product for sale.

[0245] The term "pharmaceutical product" refers to a product containing an active drug ingredient that has been approved for sale by a governmental authority, such as the Food and Drug Administration or a similar authority in other countries.

[0246] "Label" or "pharmaceutical product labeling" refers to information provided to a patient that provides information regarding a pharmaceutical product. Such information may include, but is not limited to, one or more of: pharmaceutical product description, clinical pharmacology, indications (use of the pharmaceutical product), contraindications (who should not take the pharmaceutical product), warnings, precautions, adverse events (side effects), drug abuse and dependence, dosage and administration, use during pregnancy, use in nursing mothers, use in children and the elderly, mode of delivery of the pharmaceutical product, patient safety information, or any combination thereof. In some embodiments, the label or pharmaceutical product labeling provides instructions for a patient in need of the BCMA CAR T cells. In further embodiments, the label or pharmaceutical product labeling identifies the siltacabtadine autolucel suspension and provides instructions for a patient in need of the BCMA CAR T cells.

[0247] The term "reference product" refers to an FDA approved biological product (approved pharmaceutical product) to which the proposed biosimilar product is compared. In addition, the approval of the reference product is based on a full complement of safety and efficacy data. The proposed biosimilar product is compared and evaluated with the reference product to ensure that the products are highly similar and have no clinically meaningful differences.

[0248] "Biosimilar" refers to a biological product that is highly similar to a conventional FDA approved reference product (approved pharmaceutical product) and has no clinically meaningful differences. By extensively analyzing (i.e., characterizing) the structure and function of the reference product and the submitted biosimilar, and comparing the product properties, e.g., purity, chemical properties, and biological activity, the high similarity between the biosimilar and the reference product can be demonstrated. Minor differences in clinical inactive ingredients between the reference product and the submitted biosimilar product (e.g., minor differences compared to stabilizers or buffers used in the reference product) are acceptable. FDA will carefully evaluate any differences between the submitted biosimilar product and the reference product to ensure that the biosimilar meets FDA's high approval standards. During the manufacturing of a biological product, minor differences (i.e., acceptable intra-product variations) are expected, whether the product is a biosimilar or a reference product. The manufacturer must further demonstrate that the submitted biosimilar product has no clinically relevant differences from the reference product in terms of safety, purity, and potency (safety and effectiveness), typically demonstrated through human pharmacokinetic (exposure) and pharmacodynamic (response) studies, clinical immunogenicity assessments, and (if necessary) additional clinical studies.

[0249] The manufacturer of the submitted biosimilar product will generate data to demonstrate biosimilarity of the submitted product to the FDA-approved reference product. Applications for biosimilar products must include data demonstrating biosimilarity to the reference product, where: - analytical studies have shown that the biological product is highly similar to the reference product, without any subtle differences in the active ingredients in clinical practice; - Animal studies, including toxicity assessments; and - One or more clinical studies sufficient to demonstrate that the submitted biosimilar product achieves the safety, purity, and potency in the indication or indications approved for the reference product, and will usually include data from immunogenicity, pharmacokinetics (PK), and possibly pharmacodynamics (PD) assessments, and potentially comparative clinical studies.

[0250] Rather than generating the same complete nonclinical and clinical data as the reference product, manufacturers who demonstrate that the submitted biosimilar and the FDA-approved reference product are highly similar and have no clinically meaningful differences may rely in part on the safety and effectiveness determinations that FDA previously approved for the reference product. Therefore, biosimilar manufacturers may not need to conduct as many clinical trials.

[0251] The present specification further discloses a method of supplying, storing and processing an approved pharmaceutical product comprising siltacabtadine autorucel suspension. The approved pharmaceutical product comprising siltacabtadine autorucel suspension is provided in an infusion bag containing frozen siltacabtadine autorucel suspension in 5% DMSO, a 70 mL suspension in an infusion bag and metal case (NDC 57894-111-01) or a 30 mL suspension in an infusion bag and metal case (NDC 57894-111-02). Each infusion bag is individually packaged in an aluminum freezer case. The infusion bag is stored and shipped in a cryogenic container at or below -120°C, for example in liquid nitrogen vapor, and the approved pharmaceutical product is stored in the original packaging including the case protecting the infusion bag. In some embodiments, the time from leukapheresis to product availability may be 27 to 66 days. In some embodiments, the time from leukapheresis to product availability may be 32 days.

[0252] The following illustrative examples are intended to be merely examples of the present disclosure and should not be construed as limiting the disclosure in any way. Illustrative Examples Example 1. A method of treating a subject, comprising administering to the subject a single infusion of a dose of a composition comprising T cells containing a chimeric antigen receptor (CAR); wherein the CAR comprises the amino acid sequence of SEQ ID NO: 17; wherein the dose is 0.5×10 6 to 1.0×10 6 T cells / kg subject weight, and wherein the method comprises completing administration of the dose of T cells to the subject within about 2.5 hours at a temperature of about 20° C. to 25° C. (Example 2) The method of Example 1, wherein the subject has relapsed or refractory multiple myeloma that has received multiple prior lines of therapy, and optionally, the subject has received three or more prior lines of therapy. (Example 3) The method of Example 1 or 2, wherein the subject has received four or more prior lines of treatment. (Example 4) The method described in Example 2 or 3, wherein the prior line of treatment includes a proteasome inhibitor, an immunomodulator, and an anti-CD38 monoclonal antibody. (Example 5) The method according to any one of Examples 1 to 4, wherein the T cells are autologous T cells. (Example 6) The method of any one of Examples 1-5, wherein the method further comprises administering to the subject a lymphodepleting chemotherapy regimen prior to administering to the subject the T cells. (Example 7) The method described in Example 6, wherein the lymphodepleting chemotherapy regimen comprises administering to the subject cyclophosphamide and fludarabine. (Example 8) The method described in Example 7, wherein the lymphodepleting chemotherapy regimen comprises intravenously administering cyclophosphamide and fludarabine to the subject. Example 9: The lymphodepleting chemotherapy regimen is administered to the subject at about 300 mg / m 2 and cyclophosphamide at a dose of 30 mg / m 2 The method of Example 8, comprising administering a dose of fludarabine intravenously daily. (Example 10) The method of any one of Examples 6-9, wherein the lymphodepleting chemotherapy regimen is about 3 days. (Example 11) The method of any one of Examples 6-10, wherein the method comprises administering the lymphodepleting chemotherapy regimen to the subject for at least about 2-4 days prior to administering the T cells to the subject. (Example 12) The method of any one of Examples 1-11, wherein the method further comprises administering a premedication to the subject at most 60 minutes prior to administering the T cells to the subject, and wherein the premedication comprises an antipyretic and an antihistamine. (Example 13) The method of Example 12, wherein the method includes administering the premedication to the subject about 30 to 60 minutes prior to administering the T cells to the subject. (Example 14) The method described in Example 12 or 13, wherein the antipyretic includes paracetamol or acetaminophen. (Example 15) The method described in Example 14, wherein the antipyretic comprises acetaminophen in a dose of about 650 mg to 1000 mg. (Example 16) The method of any one of Examples 12 to 15, wherein the antihistamine comprises diphenhydramine. (Example 17) The method described in Example 16, wherein the dose of diphenhydramine is about 25 mg to 50 mg or equivalent. (Example 18) The method according to any one of Examples 12 to 17, wherein the premedication is administered orally or intravenously. (Example 19) The method of any one of Examples 12-18, wherein the premedication does not include a systemic corticosteroid. (Example 20) The method of any one of Examples 1-19, wherein the method includes thawing the dose of T cells prior to administration, and wherein the thawing is completed within about 15 minutes. (Example 21) The method of Example 20, wherein the dose of T cells is thawed at a temperature of about 37°C ± 2°C. (Example 22) The method of any one of Examples 1-21, wherein the method further comprises treating cytokine release syndrome (CRS) in the subject after administering the dose of T cells. (Example 23) The method of Example 22, wherein CRS includes fever, fever, hypotension, elevated aspartate aminotransferase, chills, elevated alanine aminotransferase, sinus tachycardia, hyperbilirubinemia, hypoxia, respiratory failure, acute kidney injury, disseminated intravascular coagulation and hemorrhage (e.g., retroperitoneal, intracerebral or gastrointestinal hemorrhage), hemophagocytic lymphohistiocytosis (HLH), macrophage activation syndrome (MAS), angina pectoris, supraventricular and ventricular tachycardia, malaise, myalgia, increased C-reactive protein, ferritin, blood alkaline phosphatase, gamma-glutamyltransferase, organ toxicity, or any combination thereof. (Example 24) The method of Example 22 or 23, wherein treating CRS in the subject comprises administering to the subject an anti-cytokine agent or a corticosteroid. (Example 25) The method described in Example 24, wherein the anti-cytokine agent comprises a monoclonal antibody that targets a cytokine. (Example 26) The method described in Example 25, wherein the monoclonal antibody targeting the cytokine is an IL-6R inhibitor. (Example 27) The method described in Example 26, wherein the IL-6R inhibitor is tocilizumab. (Example 28) The method described in Example 27, wherein the method comprises intravenously administering tocilizumab at a dose of about 8 mg / kg within about 1 hour. (Example 29) The method described in Example 28, wherein the dose of tocilizumab does not exceed about 800 mg. (Example 30) The method described in Example 28 or 29, wherein the dose of tocilizumab does not exceed three doses within a 24-hour period. (Example 31) The method of any one of Examples 28-30, wherein the dose of tocilizumab does not exceed a total of four doses. (Example 32) The method described in Example 24, wherein the anti-cytokine agent further comprises an anti-cytokine agent other than tocilizumab. (Example 33) The method described in Example 24, wherein the anti-cytokine agent further comprises a monoclonal antibody that targets a cytokine other than tocilizumab. (Example 34) The method according to any one of Examples 24 to 33, wherein the corticosteroid comprises dexamethasone or methylprednisolone. (Example 35) The method described in Example 34, wherein the corticosteroid is dexamethasone. (Example 36) The method of Example 35, wherein the method comprises administering dexamethasone intravenously to the subject at a dose of about 10 mg every 12 to 24 hours. (Example 37) The method of Example 36, wherein the method comprises administering dexamethasone intravenously to the subject at a dose of about 10 mg every 12 hours. (Example 38) The method of Example 35, wherein the method comprises administering dexamethasone intravenously to the subject at a dose of about 20 mg every about 6 to 12 hours. (Example 39) The method of Example 38, wherein the method comprises administering dexamethasone intravenously to the subject at a dose of about 20 mg about every 6 hours. (Example 40) The method described in Example 34, wherein the corticosteroid is methylprednisolone. (Example 41) The method of Example 40, wherein the method comprises intravenously administering methylprednisolone to the subject at a dose of about 2 mg / kg about every 12 hours. (Example 42) The method of Example 40, wherein the method comprises intravenously administering methylprednisolone to the subject at a dose of about 1 to 2 g every 24 hours. (Example 43) The method described in Example 22 or 23, wherein the method includes administering an immune inhibitor to the subject. (Example 44) The method of any one of Examples 1 to 43, wherein the method further comprises treating the subject for neurotoxicity after administering the dose of the T cells. (Example 45) The method described in Example 44, wherein the neurotoxicity includes immune effector cell-associated neurotoxicity syndrome (ICANS), Parkinson's disease, Guillain-Barre syndrome, immune-mediated myelitis, peripheral neuropathy, cranial nerve palsy, or any combination thereof. (Example 46) The method described in Example 45, wherein the neurotoxicity comprises ICANS, and wherein the ICANS comprises encephalopathy, aphasia, headache, decreased level of consciousness, seizures, motor finding, increased intracranial pressure (ICP), cerebral edema, or any combination thereof. (Example 47) The method described in Example 46, wherein the ICANS includes focal or generalized epileptic seizures, non-convulsive epileptic seizures on electroencephalogram (EEG), life-threatening prolonged epileptic seizures, recurrent clinical or electrical epileptic seizures, deep focal motor asthenia, hemiparesis, paraparesis, focal / focal edema on neuroimaging, stupor, unconsciousness, diffuse cerebral edema on neuroimaging, decerebrate or decorticate posturing, cranial nerve VI palsy, optic disc edema, Cushing's syndrome, or any combination thereof. (Example 48) The method of Example 46 or 47, wherein the method comprises administering dexamethasone intravenously to the subject at a dose of about 10 mg every 12 to 24 hours for about 2 to 3 days. (Example 49) The method described in Example 48, wherein the method comprises administering dexamethasone intravenously to the subject at a dose of about 10 mg every 12 hours for about 2 to 3 days or more. (Example 50) The method described in Example 46 or 47, wherein the method comprises administering dexamethasone intravenously to the subject at a dose of about 10 to 20 mg about every 6 hours. (Example 51) The method of any one of Examples 46-50, wherein the method comprises administering methylprednisolone to the subject at a dose of about 1-2 g / day about every 24 hours. (Example 52) The method described in any one of Examples 46 to 50, wherein the ICANS includes cerebral edema. (Example 53) The method described in Example 52, wherein the method includes administering hyperventilation and hypertonicity therapy to the subject. (Example 54) The method of any one of Examples 46 to 53, wherein the method comprises administering to the subject a non-sedative antiepileptic drug. (Example 55) The method described in Example 54, wherein the non-sedating antiepileptic drug is levetiracetam. (Example 56) The method described in Example 45, wherein the neurotoxicity includes Parkinson's disease. (Example 57) The method described in Example 56, wherein the Parkinson's disease includes Parkinsonian symptoms or non-Parkinsonian symptoms. (Example 58) The method described in Example 57, wherein the Parkinson's symptom or the non-Parkinson's symptom comprises tremor, bradykinesia, involuntary movements, stereotypies, loss of spontaneous movement, masked facies, apathy, flatness, fatigue, rigidity, psychomotor slowness, micrographia, dysgraphia, apraxia, lethargy, confusion, drowsiness, loss of consciousness, slow reflexes, hyperreflexia, memory loss, difficulty swallowing, fecal incontinence, falls, stooped posture, bradygait, muscle weakness and wasting, motor dysfunction, motor and sensory loss, akinetic mutism, frontal lobe discharge signs, or any combination thereof. (Example 59) The method of any one of Examples 56-58, wherein the method comprises administering a treatment to the subject to alleviate Parkinson's disease. (Example 60) The method described in Example 45, wherein the neurotoxicity includes Guillain-Barrett syndrome. (Example 61) The method described in Example 60, wherein the Guillain-Barrett syndrome includes symptoms consistent with the Miller-Fisher variant of Guillain-Barrett syndrome, encephalopathy, motor ataxia, speech impairment, polyradiculoneuropathy, or any combination thereof. (Example 62) The method of Example 61, wherein the method comprises administering a treatment to the subject to alleviate Guillain-Barrett's syndrome. (Example 63) The method described in Example 45, wherein the neurotoxicity includes immune-mediated myelitis. (Example 64) The method described in Example 63, wherein the symptoms of immune-mediated myelitis include decreased sensation in the lower extremities or lower abdomen accompanied by impaired sphincter control. (Example 65) The method of Example 63 or 64, wherein the method includes administering a treatment to the subject to alleviate immune-mediated myelitis, optionally wherein the treatment includes a corticosteroid or an immune globulin, and optionally wherein the method includes administering the immune globulin intravenously. (Example 66) The method described in Example 45, wherein the neurotoxicity includes peripheral neuropathy. (Example 67) The method described in Example 66, wherein the peripheral neuropathy includes sensory neuropathy, motor neuropathy, sensory-motor neuropathy, or any combination thereof. (Example 68) The method described in Example 66 or 67, wherein the method comprises administering a treatment to the subject to alleviate peripheral neuropathy. (Example 69) The method described in Example 45, wherein the neurotoxicity includes cranial nerve paralysis. (Example 70) The method described in Example 69, wherein the cranial nerve palsy includes cranial nerve 3 palsy, cranial nerve 6 palsy, cranial nerve 7 palsy, or bilateral cranial nerve 7 palsy. (Example 71) The method described in Example 69 or 70, wherein the method includes administering a treatment to the subject to alleviate cranial nerve palsy. (Example 72) The method described in Example 23, wherein CRS includes hemophagocytic lymphohistiocytosis (HLH) or macrophage activation syndrome (MAS), wherein symptoms of HLH or MAS include hypotension, hypoxia with diffuse alveolar damage, coagulation disorders, cytopenias, multiple organ dysfunction including renal dysfunction, or any combination thereof. (Example 73) The method described in Example 72, wherein the method includes administering a treatment to the subject to alleviate HLH or MAS. (Example 74) The method of any one of Examples 1-73, wherein the method further comprises treating the subject for prolonged or recurrent cytopenia prior to administering to the subject T cells comprising a CAR, after administering to the subject a lymphoablative chemotherapy regimen, or after administering the dose of T cells comprising a CAR. (Example 75) The method described in Example 74, wherein the long-term recurrent cytopenia includes long-term neutropenia, long-term thrombocytopenia, recurrent neutropenia, thrombocytopenia, lymphopenia, anemia, or any combination thereof. (Example 76) The method described in Examples 1 to 75, wherein the method further comprises treating the infection in the subject. (Example 77) The method of Example 76, wherein the infection is due to a viral, bacterial, fungal or unindicated pathogen, optionally wherein the infection comprises lung abscess, sepsis, pneumonia, bronchopulmonary aspergillosis, Pneumocystis jiroveci pneumonia, CMV colitis (with HSV-1 hepatitis), mycotic aneurysm, cerebral aspergillosis, or COVID-19 infection. (Example 78) The method described in Example 76, wherein the infection causes febrile neutropenia or subarachnoid hemorrhage. (Example 79) The method of any one of Examples 76 to 78, wherein the method comprises administering an antimicrobial agent to the subject. (Example 80) The method described in Example 79, wherein the antimicrobial agent is an antibiotic. (Example 81) The method described in Example 80, wherein the antibiotic is a broad-spectrum antibiotic. (Example 82) The method described in Example 77, wherein the infection is viral. (Example 83) The method described in Example 82, wherein the method includes administering an anti-viral therapy or vaccine to the subject. (Example 84) The method of any one of Examples 1 to 83, wherein the method further comprises treating the subject for hypogammaglobulinemia. (Example 85) The method of Example 84, wherein after administering the dose of T cells comprising a CAR, the hypogammaglobulinemia comprises a laboratory IgG level of less than about 500 mg / dL. (Example 86) The method of Example 84 or 85, wherein the method comprises administering to the subject a dose of intravenous immunoglobulin (IVIG) after administering the dose of T cells comprising a CAR. (Example 87) The method of any one of Examples 1 to 86, wherein the method further comprises treating the hypersensitivity reaction in the subject. (Example 88) The method described in Example 87, wherein the hypersensitivity reaction includes flushing, chest discomfort, tachycardia, asthma, tremors, burning sensation, allergic reaction, or any combination thereof. (Example 89) The method described in Example 87 or 88, wherein the method comprises administering a treatment to the subject to reduce the hypersensitivity reaction. (Example 90) The method of any one of Examples 1 to 89, wherein the method further comprises treating a secondary malignancy in the subject. (Example 91) The method described in any one of Examples 1 to 90, wherein the composition further comprises an excipient selected from dimethyl sulfoxide or dextran-40. (Example 92) The method described in Example 91, wherein the excipient is dimethyl sulfoxide. (Example 93) The method described in Example 92, wherein the excipient is about 1% to 10% dimethyl sulfoxide. (Example 94) The method described in Example 93, wherein the excipient is about 5% dimethyl sulfoxide. (Example 95) A pharmaceutical product comprising siltacabtadine autorucel suspension for intravenous infusion, said pharmaceutical product being packaged, and wherein said packaging comprises a label, said label identifying siltacabtadine autorucel suspension as an approved pharmaceutical product for treating adult patients with relapsed or refractory multiple myeloma after receiving four or more prior lines of therapy, said prior lines of therapy comprising a proteasome inhibitor, an immunomodulatory agent, and an anti-CD38 monoclonal antibody. (Example 96) A method for treating relapsed or refractory multiple myeloma in an adult patient in need thereof, comprising administering an approved pharmaceutical product comprising siltacabtadine autorucel suspension in an amount and manner as specified in the pharmaceutical product labeling of the approved pharmaceutical product. Example 97: A method of marketing an approved pharmaceutical product comprising siltacabtadine autorucel suspension, comprising marketing such a pharmaceutical product, wherein the pharmaceutical product label of a reference product for such pharmaceutical product comprises instructions for treating adult patients with relapsed or refractory multiple myeloma after receiving four or more prior lines of therapy, the prior lines of therapy comprising a proteasome inhibitor, an immunomodulatory agent, and an anti-CD38 monoclonal antibody. (Example 98) A method for authorizing the sale of a pharmaceutical product comprising siltacabtadine autorucel suspension, comprising authorizing the sale of such pharmaceutical product, wherein the pharmaceutical product label of a reference product for such pharmaceutical product comprises instructions for treating adult patients with relapsed or refractory multiple myeloma after receiving four or more prior lines of therapy, the prior lines of therapy comprising a proteasome inhibitor, an immunomodulatory agent, and an anti-CD38 monoclonal antibody.

[0253] example The following examples are provided to further illustrate some of the embodiments disclosed herein. The examples are intended to illustrate, but not to limit, the disclosed embodiments.

[0254] Example 1: Siltacavtadine Autorusel B-cell maturation antigen (BCMA, also called CD269 and TNFRSF17) is a 20 kilodalton type III membrane protein that is part of the tumor necrosis receptor superfamily. BCMA is a cell surface antigen that is expressed at high levels mainly on B-lineage cells. Figure 1 shows the expression of BCMA on various immune-derived cells. Comparative studies have shown that BCMA is deficient in most normal tissues and is not expressed on CD34-positive hematopoietic stem cells. BCMA binds to two ligands that induce B-cell proliferation and plays a vital role in B-cell maturation and subsequent differentiation into plasma cells. Due to its selective expression and biological importance to myeloma cell proliferation and survival, BCMA has become a promising target for CAR-T-based immunotherapy (siltacabtadine autreucel).

[0255] Sirtacabtadine Autorucel is an autologous chimeric antigen receptor T cell (CAR-T) therapy that targets BCMA. The Sirtacabtadine Autorucel chimeric antigen receptor (CAR) is designed to confer avidity and contains two VHH domains that target B cell maturation antigen (BCMA). A diagram of the construct is shown in Figure 2. Sirtacabtadine Autorucel contains a VHH domain that comprises the amino acid sequence set forth in SEQ ID NO: 2 and a VHH domain that comprises the amino acid sequence set forth in SEQ ID NO: 4.

[0256] Example 2: Treatment with Siltacavtadine Autolucel Herein, a stage 1b-2, open-label, multicenter study is described, which aimed to assess the safety and efficacy of siltacabtadine autorucel in adult subjects with relapsed or refractory multiple myeloma. In the stage 1b portion of the study, the recommended stage 2 dose (RP2D) of siltacabtadine autorucel was confirmed. In stage 2, subjects were treated with the RP2D established in stage 1b. The purpose of the stage 2 portion of the study is to further establish the safety and efficacy of siltacabtadine autorucel. In Figure 3, a schematic overview of the study flow chart is described, which includes the lymphodepletion regimen between siltacabtadine autorucel infusions.

[0257] The first analysis was performed approximately 6 months after the last subject received their initial dose of siltacabtadine autorucel. This report was generated by the first analysis designated by regimen. Table 1 lists a summary of the subjects who participated in the study, where percentages were calculated using the number of all subjects who participated in the analysis population as the denominator. In the United States, a total of 113 subjects (stage 1b: 35, stage 2: 78) participated in the study (apheresis), where 101 subjects (stage 1b: 30, stage 2: 71) received the conditioning regimen and 97 subjects (stage 1b: 29, stage 2: 68) received siltacabtadine autorucel infusion and received the infusion under the target RP2D. These 97 subjects constituted the all-treatment analysis group, which was the basis for all efficacy and safety analyses shown below. At the time of clinical cutoff, the median follow-up duration for all treatment analysis groups was 12.4 months according to the limit estimate of the Kaplan-Meier product. Table 2 lists the summary of follow-up periods for the study, which lists follow-up periods relative to the date of the first infusion of siltacabtadine autorucel (Day 1).

[0258] The patient population was screened to include patients with relapsed or refractory multiple myeloma, who had received three previous lines or were incurable with both PI / IMiD, and who had been previously exposed to PI, IMiD, and anti-CD38, where PI was a proteasome inhibitor and IMiD was an immunomodulatory drug. Another possible drug was an alkylating agent (ALKY). Eligible patients were age ≥ 18 years, diagnosed with MM based on International Myeloma Working Group (IMWG) criteria, had measurable disease at baseline, and had an Eastern Cooperative Oncology Group (ECOG) behavioral status score of 0, 1, or 2. The demographic and disease characteristics of the patient population in the stage 1b portion of the study are shown in Figure 8.

[0259] Apheresis was performed on eligible subjects to collect peripheral blood mononuclear cells (PBMCs). Study entry was defined as the day of apheresis. Siltacabtadine Autolucel Drug Product (DP) was produced by selected T cells from the apheresis. Subjects who failed apheresis or production were allowed to reattempt apheresis.

[0260] Bridging therapy (anti-plasma cell-directed therapy between apheresis and the first dose conditioning regimen) was permitted if clinically indicated (i.e., to maintain disease stability while waiting for siltacabtadine auto-repair). Additional cycles of bridging therapy were considered based on the subject's clinical status and time of CAR-T product availability. Bridging therapy was defined as short-term treatment in which at least a disease-stabilizing response had previously occurred for the subject.

[0261] After meeting the safety criteria for treatment, subjects were administered a conditioning regimen to help them achieve lymphodepletion and promote the expansion of CAR-T cells. The lymphodepleting regimen consisted of cyclophosphamide 300 mg / m 2 and fludarabine 30 mg / m 2 , administered intravenously (IV) daily for three days. Cyclophosphamide 300 mg / m before siltacabtadine autolucel injection. 2 and fludarabine 30 mg / m 2 This was consistent with the lymphodepleting regimen used in the commercially available CAR-T products Kymriah and Yescarta.

[0262] Five to seven days after the conditioning regimen began, siltacabtadine autorucel produced from apheresis material by viral transduction as shown in Figure 4 was administered on a day defined as day 1. Subjects were premedicated approximately 1 hour prior to siltacabtadine autorucel infusion. No corticosteroids were used during preinfusion. Table 5 lists the infused prodrugs. After treatment with infused prodrugs, 0.75 x 10 6CAR-positive live T cells / kg (range: 0.5–1.0 × 10 6 Administer siltacabtadine autolysate in a single infusion at a total target dose of 1.0 x 10 CAR-positive live T cells / kg, with a maximum total dose of 1.0 x 10 8 were live CAR-positive T cells.

[0263] One or two frozen patient-specified infusion bags contained a fixed dose of siltacabtadine autorusel. The thawing time of the siltacabtadine autorusel was timed to match the infusion time. The infusion time was confirmed in advance and the start time of thawing was adjusted so that the patient could have the siltacabtadine autorusel for infusion when they were ready. If receiving more than one bag to perform a therapeutic infusion, thaw one bag at a time. The next bag was thawed / infused when it was confirmed that the previous bag had been safely administered.

[0264] The post-infusion phase began on day 1, when siltacabtadine autorucel infusion was completed, and continued until day 100. The post-treatment phase began on day 101 and continued until study completion, defined as 2 years after the last subject received their initial dose of siltacabtadine autorucel. Figure 14 summarizes the amplification and persistence of siltacabtadine autorucel as measured by blood concentrations.

[0265] Example 3: Evaluation of the efficacy of treatment with siltacabtadine autolucel Using the IMWG-based response criteria summarized in Table 6, the study classified responses from best to worst as stringent complete response (sCR), complete response (CR), best partial response (VGPR), partial response (PR), minimal response (MR), disease stabilization, or disease progression. Disease progression was recorded consistently across clinical study sites. The studies performed to assess the IMWG-based response criteria were as follows: · Measurement of myeloma protein in serum and urine: Myeloma protein (M protein) was measured using the following tests from blood and 24-hour urine samples: serum quantitative Ig, serum protein electrophoresis (SPEP), serum immunofixation electrophoresis, serum FLC assay (performed at every disease assessment for subjects suspected of CR / sCR and those with serum FLC disease only), 24-hour urinary M protein electrophoresis quantitative (UPEP), urine immunofixation electrophoresis, serum β2-microglobulin. At least one repeat study demonstrated disease progression based on only one laboratory test. After relapse from CR, disease evaluation continued until disease progression was confirmed. Serum and urine immunofixation and serum free light chain (FLC) assays were performed at screening and thereafter when CR was suspected (if serum or 24-hour urinary M protein electrophoresis [by SPEP or UPEP] was zero or not quantifiable). For subjects with light chain multiple myeloma, serum and urine immunofixation tests were typically performed. Serum calcium corrected for albumin: Collect and analyze blood samples to calculate serum calcium corrected for albumin until proven disease progression develops; development of hypercalcemia (corrected serum calcium >11.5 mg / dL [>2.9 mmol / L]) in the absence of any other cause may indicate disease progression or recurrence. Calcium is bound to albumin, and only unbound (free) calcium has biological activity, therefore serum calcium levels must be adjusted for abnormal albumin levels ("corrected serum calcium"). Bone marrow examination: Bone marrow aspirate or biopsy was performed for clinical evaluation. Bone marrow aspirate was performed for biomarker evaluation. Clinical classification (morphology, cytogenetics, immunohistochemistry or immunofluorescence or flow cytometry) was performed. Immunophenotyping was performed on a portion of the bone marrow aspirate to monitor checkpoint ligand expression in BCMA, CD138 positive multiple myeloma cells, and checkpoint expression in T cells. When possible, bone marrow aspirate was performed to confirm CR and sCR and disease progression. Note that minimal residual disease (MRD) negativity is a potential surrogate for PFS and OS in multiple myeloma treatment, so next generation sequencing (NGS) was used on bone marrow aspirate DNA to monitor MRD in subjects. Baseline bone marrow aspirates were used to define myeloma clones, and post-treatment samples were used to evaluate MRD negativity. Fresh bone marrow aspirates were collected prior to the first dose conditioning regimen (≤7 days). · Bone Survey: During the screening phase, a bone survey (including skull, entire spine, pelvis, thorax, humerus, femur, and any other bones suspected by the investigator to be disease-related) was performed and assessed by radiography ("X-ray") or low-dose computed tomography (CT) scan without the use of IV contrast. If a CT scan was used, it was of diagnostic quality. After siltacabtadine autorucel injection and before confirming disease progression, an X-ray or CT scan was performed focally to document response or progression, with or without clinical indication based on symptoms. Magnetic resonance imaging (MRI) was an acceptable method to assess bone disease and was incorporated as appropriate, however, it did not replace a bone survey. If a radionuclide bone scan was used at screening in addition to a complete bone survey, disease status was documented in two ways. These tests were performed simultaneously. A radionuclide bone scan may not replace a complete bone survey. If a subject showed disease progression manifested by pain symptoms with bone changes, disease progression was documented by bone survey or other radiography depending on the symptoms experienced by the subject. If the radiological survey yielded a clear diagnosis of disease progression, it was not considered necessary to perform a repeat confirmatory x-ray. If changes were not clear, a repeat x-ray was performed within 1 to 3 weeks. Documentation of extramedullary plasmacytoma: Known sites of extramedullary plasmacytoma were documented ≤14 days prior to the first dose conditioning regimen. Clinical examination or MRI was used to document extramedullary site of disease. If there was no contraindication to IV contrast use, CT scan evaluation was considered an acceptable alternative regimen. Positron emission tomography scans or ultrasound studies were not permitted to document the size of extramedullary plasmacytoma. However, if the CT portion of the PET / CT fusion scan was of sufficient diagnostic quality, the PET / CT fusion scan was optional to document extramedullary plasmacytoma. Extramedullary plasmacytoma was assessed by clinical examination or radiological imaging for all subjects with a history of plasmacytoma or if clinically indicated ≤14 days prior to the first dose conditioning regimen. For subjects with a history of plasmacytoma or other subjects with clinical indications during treatment, local assessment, measurement or evaluation of measurable sites of extramedullary disease was performed (for physical examination) every 4 weeks until a confirmed CR or confirmed disease progression occurred. If only radiological assessment was available, evaluation of extramedullary plasmacytoma was performed every 12 weeks. Irradiated or resected lesions were considered non-measurable and were monitored for disease progression only. To qualify for VGPR or PR / minimal response (MR), the sum of the products of the perpendicular diameters of previous extramedullary plasmacytomas had to decrease by more than 90% or at least 50%, respectively, and no new plasmacytomas had to develop. To qualify for disease progression, the sum of the products of the perpendicular diameters of previous extramedullary plasmacytomas had to increase by at least 50%, or the longest diameter of a previous lesion that was >1 cm on the short axis had to increase by at least 50%, or new plasmacytomas had to develop. Although not all extramedullary plasmacytomas present were reported, disease progression criteria were met if the sum of the products of the perpendicular diameters of the reported plasmacytomas increased by at least 50%.

[0266] If the study treatment was identified as interfering with the immunofixation assay, CR was defined as disappearance of the original M protein associated with multiple myeloma at the time of immunofixation, and the assay for CR was unaffected by unrelated M protein secondary to the study treatment. The study endpoints (as assessed by an Independent Review Committee (IRC)) were: Next-generation sequencing (clonoSEQ version 2.0) (Adaptive Biotechnologies, Seattle, WA, USA) was used to assess MRD in patients with suspected complete responses at baseline, day 28, and follow-up at months 6, 12, 18, and 24, and every 12 months until disease progression in patients who remained on study. MRD negativity was assessed in the following samples, which were calibrated or quality controlled and included 10 -5 The persistence of MRD-negativity was assessed by estimating MRD-negativity rates at 6 and 12 months of follow-up. Clinical benefit rate (CBR) was defined as the proportion of subjects who achieved MR or better response (sCR+CR+VGPR+PR+MR) based on IMWG criteria. Overall response rate (ORR) was defined as the proportion of subjects who achieved PR or better response (sCR+CR+VGPR+PR) based on IMWG criteria. VGPR or better response rate was defined as the proportion of subjects who achieved VGPR or better response (sCR+CR+VGPR) based on IMWG criteria. Duration of response (DOR) was calculated from the time from the first documented date of response (PR or better response) to the date of first documented evidence of progressive disease (as defined by IMWG criteria) in responders (those with PR or better response). Relapse from CR with positive immunofixation or trace M protein was not considered to be disease progression. After relapse from CR, disease evaluation continued until disease progression was confirmed. · Duration of response (TTR) was defined as the time from the date of first infusion of siltacabtadine autorucel to the first efficacy assessment at which the subject met all criteria for PR or better response. Progression-free survival (PFS) was defined as the time from the date of first infusion of siltacabtadine autorucel to the date of first documented disease progression (as defined by IMWG criteria) or death from any cause (whichever occurred first). Overall survival (OS) was measured from the date of the first infusion of siltacabtadine autorucel to the date of subject death.

[0267] For ORR, response rates and their 95% exact confidence intervals (CIs) were calculated based on a binomial distribution, and the null hypothesis was rejected if the lower limit of the confidence interval was >30%. Analyses of response rates of VGPR or better response, DOR, PFS, and OS were performed with the same cutoffs as for ORR. The Kaplan-Meier method was used to estimate time-to-event efficacy endpoints (DOR, PFS, and OS). Distributions (medians and Kaplan-Meier curves) of DOR were provided using Kaplan-Meier estimates. Similar analyses were performed for OS, PFS, and TTR.

[0268] Example 4: Safety evaluation of treatment with siltacabtadine autolucel Adverse events, except CRS and CAR-T cell-related neurotoxicity (e.g., ICANS), were tracked, reported, and classified according to the National Cancer Institute Common Terminology Standard for Adverse Events (NCI-CTCAE version 5.0). CRS was graded based on the ASTCT consensus classification and summarized in Table 7. Subjects were immediately admitted to the hospital for evaluation at the first sign of CRS (e.g., fever). If other causes of fever were excluded, tocilizumab intervention was used as needed to treat subjects with symptoms of fever. Tocilizumab was used as needed for early treatment of subjects at high risk of CRS severity (e.g., high baseline tumor burden, early onset of fever, or persistent fever 24 hours after symptomatic treatment). Other monoclonal antibodies targeting cytokines (e.g., anti-IL1 and / or anti-TNFα) were optionally used, especially if CRS did not respond to tocilizumab.

[0269] CAR-T cell-associated neurotoxicity (e.g., ICANS) was classified by the ASTCT consensus classification and summarized in Table 8. All individual symptoms of CRS (e.g., fever, hypotension) and ICANS (e.g., decreased level of consciousness, epilepsy) were obtained as individual adverse events and classified based on the CTCAE criteria. Neurotoxicity not temporally related to CRS or any other neurological adverse event not conforming to ICANS was classified based on the CTCAE criteria. Any adverse event or serious adverse event not risked by NCI CTCAE version 5.0 was classified according to the following criteria grades based on the investigator's clinical judgment. Grade 1: Mild, asymptomatic or mildly symptomatic, clinical or diagnostic observation only, no intervention indicated. Grade 2: Moderate, minimal, focal or non-invasive intervention is indicated, limiting instrumental activities of daily living appropriate for age. Grade 3: Severe or medically significant but not immediately life-threatening, indicating hospitalization or prolonged hospitalization, impairment, or limiting self-care activities of daily living. Grade 4: Life-threatening consequences, emergency intervention indicated. Grade 5: Death related to an adverse event.

[0270] Based on the independent review committee (IRC) assessment, the response and duration of response of responders in the all-treatment analysis population, with a median follow-up period of 12.4 months, are shown in Figure 5. Table 9 summarizes the overall best response of subjects in the all-treatment analysis population. In the all-treatment analysis group, 94 subjects (96.9%) achieved PR or better response, and 65 subjects (67.0%) achieved complete response (CR) or better response, with a CBR of 96.9%, based on the IRC assessment. The following shows the deep and durable response induced by siltacabtadine autorucel. The rate of VGPR or better response was 92.8%, the rate of CR or better response was 67.0%, the median follow-up period of clinical cut-off time was 12.4 months, and the median DOR was not reached. The following summarizes the indicators for evaluating the efficacy of siltacabtadine autorucel. Tumor Burden Reduction: 100% of subjects had a reduction in tumor burden. Figure 10 shows a graph of tumor burden reduction for patients in the Stage 1b-2 study. Overall Response Rate (ORR): 96.9% of subjects had an overall response with 95% exact CI (91.2%, 99.4%). Figure 11 shows a summary of the ORR for patients in the stage 1b-2 study. · VGPR or better: 90 subjects (92.8% of subjects) achieved VGPR (very good partial response) or better. · Duration of response (DOR): With 95% CI (15.9, NE) months, the median DOR was not reached and the probability of responders maintaining their response at 9 and 12 months was 80.2% (95% CI: 70.4%, 87.0%) and 68.2% (95% CI: 54.4%, 78.6%), respectively. Figure 6 shows a Kaplan-Meier plot of the DOR for all responders in the all treatments analysis population and Table 10 summarizes the DOR for all responders in the all treatments analysis population. Figure 13 shows a graph of the DOR for patients in the stage 1b-2 study. Duration of response (TTR): The median duration of first response (PR or better) and best response were 0.95 and 2.56 months, respectively. Progression-free survival (PFS): Median PFS was not reached in months with 95% CI (16.79, NE), and 9- and 12-month PFS rates (95% CI) were 80.3% (70.9%, 87.0%) and 76.6% (66.0%, 84.3%), respectively. Table 11 shows the PFS summary across all treatment analysis arms. Overall survival (OS): At the clinical cutoff time, 14 subjects (14.4%) had died. The 9-month and 12-month overall survival rates (95% CI) were 90.7% (82.8%, 95.0%) and 88.5% (80.2%, 93.5%), respectively. Figure 7 shows the Kaplan-Meier plot of OS based on the all-treatments analysis population, and Table 12 summarizes the OS based on the all-treatments analysis population. · Average residual disease (MRD) negative rate (10 -5 At the sensitivity level of 100%, the MRD-negativity rate was 54.6% (95% CI: 44.2%, 64.8%), with 33 (34.0%) subjects achieving MRD-negative CR / sCR. Table 13 shows the 100% MRD-negativity rate in bone marrow for all subjects in all treatment analysis arms. -5 A summary of the overall MDR-negativity rates in all treatment analysis groups is shown in Table 14. -5 A summary of the overall MDR negativity rates of subjects with evaluable samples in the stage 1b-2 study is shown. Evaluable samples are calibrated, quality controlled samples, and have sufficient cells to be evaluated under the respective test thresholds. A summary of the MRD of patients in the stage 1b-2 study is shown in Figures 12 and 15.

[0271] Siltacabtadine Autorcel was confirmed to have a safety profile consistent with the mechanism of action of CAR-T therapy. Figure 9 shows a summary of adverse events in the stage 1b-2 trial. CRS: CAR-T cell-related adverse events of CRS were common (94.8%) but mostly low grade. CRS of all grades was reported in 92 subjects (94.8%) as assessed by the ASTCT consensus classification system. All CRS events resolved except for one fatal event (1.1%) from a subject with CRS duration of 97 days. Table 15 and Figure 16 provide a summary of CRS events occurring during treatment in the all treatment analysis population. Figures 17-19 provide graphs and summaries of various CRS protein markers observed from all patients. Immune effector cell-associated neurotoxicity (ICANS): ICANS of any grade, as assessed by the ASTCT consensus classification system, was reported in 16 (16.5%) subjects. All events resolved in 24. Table 16 shows a summary of ICANS that occurred after siltacabtadine autorucel infusion in all treatment analysis groups. Cytopenias: Grade 3 or 4 cytopenias were common late in the infusion and included lymphopenia, neutropenia, and thrombocytopenia, but the majority of these events resolved by Day 60. By Day 100 after infusion of siltacabtadine autorucel, 96 (99.0%), 95 (97.9%), and 60 (61.9%) subjects had grade 3 or 4 lymphopenia, neutropenia, and thrombocytopenia, respectively. By Day 60, 88 (90.7%), 85 (87.6%), and 41 (42.3%) subjects had resolved their initial grade 3 or 4 events for lymphopenia, neutropenia, and thrombocytopenia to grade 2 or lower. Table 17 shows a summary of cytopenias after treatment with siltacabtadine autorucel in all treatment analysis groups.

[0272] In summary, single-agent, single-infusion siltacabtadine autorucel demonstrated unprecedented clinical activity in a heavily pretreated patient population, including an ORR of 96.9% and rapid onset of response within 1 month.

[0273] Example 5: Evaluation of treatment with siltacabtadine autorucel at a median follow-up period of 18 months Up to the time of this analysis, 97 patients had received siltacabtadine autorucel injection (median dose, 0.71 × 10 6 and the range is 0.51×10 6 ~0.95×10 6 Patients received 100 CAR-positive live T cells / kg. In 91 patients with baseline cytogenetic data, 23 patients (23.7%) had high-risk cytogenetic features with at least one abnormal chromosome, including del17p (19.6%), t(4;14) (3.1%), and / or t(14;16) (2.1%). In 96 patients with evaluable bone marrow biopsy and / or aspirate samples, more than half of patients (60.4%) had low disease burden (≤30 plasma cells), 21.9% had high disease burden (≥60 plasma cells), and 17.7% had intermediate disease burden (30-60 plasma cells). 19.6% of patients had plasmacytoma detected at screening.

[0274] Tables 18–26 and Figures 20–23 detail various safety / efficacy parameters over a median follow-up period of 18 months, and these parameters are also summarized below. effect With a median follow-up of 18 months, the ORR was 97.9% (95% CI, 92.7–99.7), with an sCR rate of 80.4%, a VGPR rate of 14.4%, and a PR rate of 3.1% (Table 1). The median duration of first response was 1 month (range, 0.9-10.7), the median duration of best response was 2.6 months (range, 0.9-15.2), and the median duration of CR or better response was 2.6 months (range, 0.9-15.2). The median DOR for the population was 21.8 months (95% CI, 21.8 to not estimable) and was not reached in patients with sCR. Median PFS for all patients was 22.8 months (95% CI, 22.8-not estimable) and was not reached in sCR patients (Figure 20). The 18-month PFS rates for all patients and sCR patients were 66.0% (95% CI, 54.9-75.0) and 75.9% (95% CI, 63.6-84.5), respectively. · The 18-month OS rate for all patients was 80.9% (95% CI, 71.4-87.6) (Figure 20). Among 61 patients with MRD assessable, 91.8% had a 10 -5 MRD-negativity was achieved at a threshold of 1. 44.3% (27 / 61) of patients maintained MRD-negativity for ≥6 months and 18% (11 / 61) maintained MRD-negativity for ≥12 months. o The 18-month PFS rates in patients with MRD persisting for ≥6 months and ≥12 months were 96.3% (95% CI, 76.5-99.5) and 100%, respectively. ORR across all subgroups was consistently high (range, 95.1%–100%), including patients treated with three prior lines of therapy (100% [95% CI, 80.5–100]), patients with high-risk cytogenetic features (100% [95% CI, 85.2–100]), patients with high disease burden (≥60% bone marrow plasma cells, 95.2% [95% CI, 76.2–99.9]), and patients with plasmacytoma (100.0% [95% CI, 82.4–100]) (Table 2). MRD negativity across all subgroups of MRD-evaluable patients (threshold, 10 -5 ) was between 80% and 100%. · The negative benefits of ORR and MRD were consistent, but other efficacy outcomes were not sustained in specific subgroups. Patients with stage ISS III disease had a lower median DOR (13.8 months [95% CI, 5.1 to not estimable]), 18-month PFS rate (34.3% [95% CI, 9.4 to 61.6]), and 18-month OS rate (48.2% [95% CI, 20.8 to 71.2]), whereas patients with baseline plasmacytoma had a lower median DOR (6.8 months [4.0 to not estimable]), 18-month PFS rate (46.8% [95% CI, 23.7 to 67.0]), and 18-month OS rate (64.5% [95% CI, 35.6 to 83.0]). · Patients with high tumor burden (≥60% bone marrow plasma cells) had lower 18-month PFS rate (50.6% [95% CI, 27.5-69.9]) and 18-month OS rate (71.4% [95% CI, 47.2-86.0]). safety No new safety signals were observed in patients treated with siltacabtadine autorucel with long follow-up duration. The most common (≥25%) grade 3 / 4 treatment-emergent AEs (TEAEs) were neutropenia (94.8%), anemia (68.0%), leukopenia (60.8%), thrombocytopenia (59.8%), and lymphopenia (49.5%) (Table 3). The most common grade 3 / 4 non-hematologic TEAEs were hypophosphatemia (7.2%), fatigue (5.2%) and elevated aspartate aminotransferase (5.2%) (Table 3). CRS was reported in 94.8% (n=92) of patients (94.6% grade 1 / 2) with a median time to onset of 7 days (range: 1-12) and median duration of 4 days (range: 1-97). 91 of 92 patients had CRS in remission within 14 days, and one patient with grade 5 CRS and hemophagocytic lymphohistiocytosis died on day 99 after prolonged grade 4 CRS sequelae. Compared to the initial analysis, no new neurotoxicity events were reported and no new cases of neurocognitive TEAEs were identified during the extended follow-up. One patient with progressive disease was re-treated with siltacabtadine autorucel and the patient had stable disease (by computer algorithm) after re-treatment and did not experience treatment-related neurotoxicity.

[0275] This extended follow-up of the siltacabtadine autorucel trial described herein demonstrated that siltacabtadine autorucel maintained clinical benefit for 18 months in RRMM patients who had received a median of six prior therapies, and that such patients experienced a manageable side effect profile and no new safety signals. At an additional 6 months of follow-up, the high ORR (97.9%) was maintained. Deep and durable responses were observed, with 80.4% of patients exhibiting sCR. MRD negativity was also maintained in 91.8% of evaluable patients, lasting for 6 to 12 months. Given evidence showing the prognostic value of MRD negativity for improved long-term survival outcomes across multiple disease settings, including patients exposed to several prior lines of therapy, it is of particular interest to assess the persistence of MRD-negativity.

[0276] After 18 months of follow-up, no new safety signals were identified for siltacabtadine autreucel. The most commonly reported TEAEs were consistent with the side effect profile of siltacabtadine autreucel reported in the initial analysis. No new neurotoxicity events were reported, and no new motor or neurocognitive TEAE cases were observed.

[0277] In summary, the data from the siltacabtadine autorucel trial with longer follow-up period described herein demonstrated significant clinical benefit of siltacabtadine autorucel in triple-exposed RRMM patients.

[0278] Example 6: Evaluation of the siltacabtadine autorucel treatment regimen in patient subgroups We then evaluated the efficacy and safety of siltacabtadine autorucel in different patient subgroups in a stage 1b-2 siltacabtadine autorucel clinical trial. Eligible patients had multiple myeloma (MM) and had received at least three prior regimens or had disease refractory to both proteasome inhibitors (PIs) and immunomodulatory drugs (IMiDs), and had received PIs, IMiDs, and anti-CD38 antibodies. After apheresis, bridging therapy was allowed to be performed. Patients underwent lymphodepletion (300 mg / m 2 Cyclophosphamide, 30 mg / m 2 Fludarabine, daily, 3 days total) followed by a single siltacabtadine auto-infusion (target dose: 0.75 × 10 6 CAR+ viable T cells / kg, range 0.5-1.0 x 10 6 ) was received. The primary objectives were to characterize the safety and efficacy of siltacabtadine autorucel, confirm the recommended stage 2 dose (stage 1b), and evaluate the efficacy (stage 2). Lee et al. (Blood 2014) classified cytokine release syndrome (CRS) and neurotoxicity based on the Common Terminology Standard for Adverse Events (CTCAE) v5.0 (stage 1b). CRS and immune effector cell-associated neurotoxicity (ICANS) were classified based on the American Society for Transplantation and Cellular Therapy (ASTCT) criteria (stage 2). Herein, Lee et al. and CTCAE v5.0 were mapped to ASTCT for CRS and ICANS, respectively. Efficacy and safety were assessed in the following subgroups based on baseline (BL) characteristics: age ≥65 years, Black / African American, three prior lines of therapy (LOT), ≥4 prior LOT, incurable with three, incurable with five drugs, standard and high-risk cytogenetics, International Classification System stage III, bone marrow plasma cells (≤30%, >30 to <60%, and ≥60%), BCMA tumor expression (<median, ≥median), and presence of plasmacytoma (bone-based and extramedullary).

[0279] Tables 28-32 and Figures 24-27 detail various safety / efficacy parameters of different subgroups analyzed with a median follow-up of 18 months. These parameters are summarized as follows:

[0280] By the time of the subgroup analysis, at a median follow-up of 18 months, 97 patients in the population (58.8% were male, median age was 61 years [range 43-78], median time from diagnosis to study entry was 5.9 years [1.6-18.2]) had received siltacabtadine autorucel. Efficacy results were comparable to the population, with ORR consistently high across all evaluated subgroups (range 95.1%-100%, Table), including subgroups with high-risk cytogenetics, ISS stage IIIMM, ≥60% BL myeloid cells, and BL plasmacytoma. Median DOR and median PFS were consistent with the population or not reached in the majority of subgroups, but were even lower in patients with high-risk disease such as ISS stage III and BL plasmacytoma (Table). In all subgroups, the ORR was consistently higher than 10% (range 95.1%-100%, Table). -5 Below the threshold of 1, the majority of MRD-evaluable patients (80%-100%) achieved MRD negativity. 18-month PFS and OS rates were consistent with the population in most subgroups. These improvements were also achieved in patients with high-risk disease, albeit at a lower incidence (Table). In the different subgroups, the incidence of CRS, ICANS, and other CAR T-cell neurotoxicity (events not reported as ICANS [i.e., occurring at some time after recovery from CRS and ICANS]) was consistent with the population, with no new safety signals.

[0281] At a median follow-up of 18 months, a single infusion of siltacabtadine autorucel induced deep and durable responses in all evaluated poor-risk patient subgroups described herein. Across the different subgroups, 90%-100% of patients achieved ORR, including patients with high-risk cytogenetics, ISS stage III MM, BL bone marrow ≥ 60% and BL plasmacytoma. The safety profile of siltacabtadine autorucel across subgroups was consistent with the population and there were no new safety signals.

[0282] Example 7: Comparison of Siltacabtadine Autorusel Treatment Method with Physician's Choice of Treatment As described in Examples 2-4 (ibid.), siltacabtadine autreucel is effective and safe in patients with relapsed or refractory multiple myeloma ("RRMM") who have been triple-exposed (exposed to an immunomodulatory drug, a proteasome inhibitor, and an anti-CD38 monoclonal antibody). In the absence of a clear standard of care for the indication and a lack of head-to-head trials evaluating siltacabtadine autreucel and other related therapies, an indirect treatment comparison (ITC) was performed between siltacabtadine autreucel and the treatments used in current clinical practice (i.e., physician's choice of treatment ("PCT")). Briefly, a meta-analysis was performed to evaluate siltacabtadine autreucel and PCT in triple-exposed RRMM patients by integrating ITCs to obtain a single ensemble effect estimate for overall survival ("OS") and progression-free survival ("PFS").

[0283] An ITC examining the comparative efficacy of siltacabtadine autorucel and PCT on OS and PFS is included. Figure 28 summarizes the selection of comparison groups for the ITC analysis. Data on PCT are derived from the following sources: (i) the Flatiron database, primarily based on US community multiple myeloma registries; (ii) long-term follow-up results of three global RRMM randomized clinical trials of daratumumab (POLLUX [NCT02076009], CASTOR [NCT02136134] and EQUULEUS [NCT01998971]); (iii) the US-based retrospective MAMMOTH study; (iv) a representative German patient registry maintained by the OncologyInformationService (OIs); and (v) a prospective, non-interventional, multinational study of real-life standard of care efficacy and safety in patients with RRMM who had received at least three prior lines of therapy (including at least one PI, an IMiD and an anti-CD38 antibody) in the LocoMMotion study [NCT04035226]. In each ITC, the PCT group consisted of patients who met the essential eligibility criteria for the stage 1b-2 clinical trial of siltacabtadine autreusell described in Examples 2-4 (the "siltacabtadine autreusell trial") and made it equivalent to the siltacabtadine autreusell trial using an inverse treatment probability weighting method. Therefore, the ITCs were considered suitable for meta-analysis. The meta-analysis used robust variance estimators to interpret the use of the siltacabtadine autreusell trial in each pair of ITCs.

[0284] A meta-analysis was performed on two populations: (1) the "ITT" (intention to treat) population, which included all participants enrolled in the siltacabtadine autorusel studies (apheresis) and all patients fulfilling the criteria in the PCT group, and (2) the "mITT" (modified intention to treat) population, which included all participants who infused siltacabtadine autorusel in the siltacabtadine autorusel studies and all patients fulfilling the criteria of no progression or death within 47 days after starting treatment (OIs and LocoMMotion is 52 days) in the PCT group, and the median (mean) time from apheresis to infusion in the siltacabtadine autorusel studies. The meta-analysis was performed in two forms. 1. The main meta-analyses considered all index dates for the modified intention to treat ("mITT") and intention to treat ("ITT") populations and used all participants treated with siltacabtadine autreuxel in the siltacabtadine autreuxel trials compared to PCT. For these meta-analyses, the start of each eligible LOT was used as the index date. Patients who met the criteria in the PCT group who received multiple subsequent therapies contributed multiple LOTs (as independent observations) to the analysis if they also met the criteria at the LOT start. ITCs with all index dates are not available for MAMMOTH and LocoMMotion. 2. Additional analyses were performed on the ITC using the first index date of the mITT and ITT populations. For these meta-analyses, the start of the first qualifying LOT was used as the index date, and each patient in the PCT group contributed only the first qualifying LOT to the analysis.

[0285] Sensitivity analyses considered ITC effect estimates based on all trial participants in the siltacabtadine Autorxel trial. Ensemble pooled effect estimates were presented as hazard ratios (HRs) and corresponding 95% confidence intervals (CIs).

[0286] Based on data availability, the main meta-analysis included four ITCs for OS and three ITCs for PFS. Sensitivity analyses including all participants in the trials verified the results of the main meta-analysis. Figures 29-32 show meta-analysis comparisons for overall survival (OS) or progression-free survival (PFS) of study-entered / treated patients and those who received physician's choice of treatment in the siltacabtadine Autorxel trials using all index dates or the first index date.

[0287] Siltacabtadine autorucel demonstrated significant superiority over PCT in ORR for OS, PFS, and TTNT, highlighting its potential as an effective therapy in triple-exposed RRMM patients. Conclusions were consistent across populations (mITT and ITT) and available index dates.

[0288] In summary, the meta-analysis demonstrated that siltacabtadine autreuil has significant advantages over PCT in OS and PFS, highlighting its potential as an effective therapy in triple-exposed RRMM patients.Without a head-to-head comparison of siltacabtadine autreuil to treatments used in real-world clinical practice, this meta-analysis for ITC demonstrated that the clinical benefit provided by siltacabtadine autreuil was much greater than that of PCT for triple-exposed RRMM patients. [Table 1] [Table 2] [Table 3] [Table 4] [Table 5] [Table 6] TIFF2025507678000008.tif158170 [Table 7] [Table 8] a: Based on the toxicity classification by Lee et al. 2019 b: ICANS grade is determined by the most severe event (ICE score, level of consciousness, epilepsy, motor manifestations, elevated ICP / cerebral edema) that cannot be attributed to any other cause. Note: All other neurological adverse events (unrelated to ICANS) during the two studies should continue to be classified using CTCAE version 5.0. [Table 9] [Table 10] [Table 11] [Table 12] [Table 13] [Table 14] [Table 15] [Table 16] [Table 17] [Table 18] [Table 19] [Table 20] [Table 21] [Table 22] [Table 23] [Table 24] [Table 25] [Table 26] [Table 27] [Table 28] [Table 29] [Table 30] [Table 31] [Table 32] [Table 33] TIFF2025507678000036.tif106170 [Table 34]

[0289] Example 8: Approved Pharmaceutical Product Label Approved Pharmaceutical Product Label: [Table 35] TIFF2025507678000039.tif245170TIFF2025507678000040.tif11170 [Table 36] 1. Indications and Usage CARVYKTI is a genetically modified autologous T-cell immunotherapy directed against the B-cell maturation antigen (BCMA) indicated for the treatment of adult patients with relapsed or refractory multiple myeloma after receiving four or more prior lines of therapy, including a proteasome inhibitor, an immunomodulatory agent, and an anti-CD38 monoclonal antibody. 2 Dosage and Administration For autologous use only. For intravenous use only. 2.1 Dosage CARVYKTI is delivered as a single-dose infusion and contains a suspension of live chimeric antigen receptor (CAR)-positive T cells in a single infusion bag. The recommended dose range is 0.5 to 1.0 × 10 6 The maximum dose of CAR-positive live T cells per injection is 1 x 10 8 These are live CAR-positive T cells. 2.2 Administration CARVYKTI is for autologous use only. Patient identity must match the patient identifier on the CARVYKTI case and infusion bag. Do not inject CARVYKTI if the information on the patient-specific label does not match the expected patient. Preparing the CARVYKTI Injection for the Patient Confirm availability of CARVYKTI prior to initiating a lymphodepleting chemotherapy regimen. Pretreatment Administration of lymphodepleting chemotherapy regimen: Cyclophosphamide 300 mg / m per day 2 Intravenous (IV) and fludarabine 30 mg / m 2 The IV was continued for 3 days. Refer to the prescribing information for cyclophosphamide and fludarabine for information on dose adjustments for renal impairment. The lymphodepleting regimen should be delayed if the patient experiences severe adverse reactions from prior bridging therapy (including clinically significant active infection, cardiac toxicity, and pulmonary toxicity) or active graft anti-host disease in patients with prior allogeneic stem cell transplantation. If the delay in CARVYKTI administration exceeds 14 days and the patient has recovered from the toxicities of the first lymphodepleting regimen, consider repeating the lymphodepleting regimen. CARVYKTI infusions are administered 2 to 4 days after completion of the lymphodepleting chemotherapy regimen. CARVYKTI infusion should be delayed if a patient has any of the following conditions: - Clinically significant active infection or inflammatory disorder. Cyclophosphamide- and fludarabine-conditioned ≥ Grade 3 non-hematologic toxicity, excluding Grade 3 nausea, vomiting, diarrhea, or constipation. CARVYKTI infusion should be delayed until these events have resolved ≤ Grade 1. Premedication Administer all patients the following infusion prodrugs 30-60 minutes prior to the CARVYKTI infusion: Antipyretics (acetaminophen 650 to 1000 mg orally or intravenously). Antihistamines (diphenhydramine 25 to 50 mg or equivalent given orally or intravenously). Prophylactic use of systemic corticosteroids should be avoided as they may inhibit the activity of CARVYKTI. Acceptance of CARVYKTI All locations approved for use for injection will support the required storage conditions for liquid nitrogen vapor. CARVYKTI will be shipped in a liquid nitrogen vapor transport box directly to the cell lab or clinical pharmacy associated with the infusion center. Verify patient identity using patient identifiers on the shipping container. If it is anticipated that the patient will not be ready to administer that day, transfer CARVYKTI to on-site liquid nitrogen vapor storage prior to the expiration of the shipping container. 2. Preparing CARVYKTI for Injection Do not thaw the product until you are ready to use it. The thawing time for CARVYKTI has been timed to match the infusion time. The infusion time was confirmed in advance and the start of thawing was timed to allow CARVYKTI infusion when the patient was ready. Once thawed, CARVYKTI infusion should be completed within 2.5 hours at room / ambient temperature (20°C-25°C). Before thawing the product, ensure that tocilizumab and emergency equipment are available prior to infusion and during recovery. 1. Verify Patient Identification: Before preparing CARVYKTI, match the patient identity to the patient identifier on the CARVYKTI case. If the information on the patient specific label does not match the expected patient, do not remove the CARVYKTI infusion bag from the case. If there are any differences between the label and the patient identifier, contact Janssen Biotech, Inc. by calling 1-800-526-7736. 2. After patient identity is verified, remove the CARVYKTI product bag from the case and check that the patient information on the case label matches the patient information on the bag label. 3. Check product bag prior to thawing for signs that may compromise the integrity of the container, such as ruptures or cracks. If bag is damaged, do not administer and contact Janssen Biotech, Inc. at 1-800-526-7736. 4. Place the infusion bag in a sealable plastic bag (preferably sterile) before thawing. 5. Thaw CARVYKTI using a water bath or dry thawing method at 37°C ± 2°C until no ice is visible in the infusion bag. Total time from start of thawing to completion of thawing should not exceed 15 minutes. 6. Remove the infusion bag from the sealable plastic bag and wipe. Gently mix the contents of the bag to disperse any clumps of cellular material. If cell clumps are still visible, continue to gently mix the contents of the bag. Small clumps of cellular material should be dispersed with gentle manual mixing. Do not pre-filter, wash, spin, or resuspend CARVYKTI in new medium into a different container prior to infusion. 7. Do not use products that have been refrozen or thawed in the refrigerator. Administration · For autologous injection only. Do not use leukocyte depletion filters. Ensure that at least two doses of tocilizumab and emergency facilities are available before infusion and during recovery. Central venous access can be used for the infusion of CARVYKTI and is encouraged in patients with poor peripheral access. 1. Verify patient identity using the patient identifier on the infusion bag. Do not inject CARVYKTI if the information on the patient specific label does not match the expected patient. 2. Irrigate the infusion set tubing with saline prior to infusion. 3. After thawing, administer the entire contents of the CARVYKTI bag via intravenous infusion within 2.5 hours using an infusion device equipped with an online filter. 4. Gently mix the contents of the bag during CARVYKTI infusion to disperse any clumps of cellular material. 5. After injection of all of the contents of the product bag, flush the administration lines (including the on-line filter) with saline at a volume equal to or greater than the total capacity of the primary administration device, including the infusion tubing, to ensure that all product is delivered. CARVYKTI contains human blood cells that have been genetically modified with a copy-dead, self-inactivating slow viral vector. Handle and dispose of CARVYKTI according to general precautions and local biosafety guidelines to avoid potential transmission of infectious diseases. Post-injection monitoring CARVYKTI will be administered at a REMS-certified healthcare facility. Monitor patients for signs and symptoms of Cytokine Release Syndrome (CRS) and neurotoxicity at least daily within 10 days of receiving CARVYKTI in a qualified medical center. Monitor periodically for signs and symptoms of delayed neurotoxicity for 4 weeks. Patients will be instructed to remain near a certified medical facility for at least four weeks after the infusion. Patients are instructed to avoid driving or risky activities for at least 8 weeks after the injection. 2.3 Management of Serious Adverse Reactions Cytokine release syndrome Identify CRS based on clinical symptoms [see Warnings and Precautions (5.1)]. Assess and treat other etiologies of fever, hypoxia, and hypotension. Consider monitoring laboratory tests for disseminated intravascular coagulation, hematological parameters, and pulmonary, cardiac, renal, and hepatic function. If CRS is suspected, manage based on suggestions in Table 1. Cardiac and other organ function in patients who experience CRS should be closely monitored until symptoms resolve. Consider antiepileptic prophylaxis with levetiracetam in patients who experience CRS. Patients experiencing grade 2 or higher CRS (e.g., hypotension unresponsive to fluids, or oxygen deprivation requiring supplemental oxygenation) should be monitored with continuous telemetry and pulse oximetry. For severe or life-threatening CRS, consider intensive care unit-level monitoring and supportive care. For CRS that is refractory to frontline interventions (e.g., tocilizumab or tocilizumab plus corticosteroids), consider alternative treatment regimens (i.e., higher corticosteroid doses, alternative anti-cytokine agents such as anti-IL1 and / or anti-TNFα, anti-T cell therapy). Refractory CRS is characterized by failure to improve fever, end-organ toxicity (e.g., hypoxia, hypotension) within 12 hours of frontline intervention or development of HLH / MAS. If neurotoxicity is suspected during CRS, administer the following: Corticosteroids due to more invasive interventions for CRS and neurotoxicity levels in Tables 1 and 2 Tocilizumab according to CRS level in Table 1 Neurotoxic antiepileptic drugs in Table 2 [Table 37] TIFF2025507678000043.tif166170Neurotoxicity Monitor patients for signs and symptoms of neurotoxicity (ICANS and other neurotoxicity) (Table 2). Rule out other etiologies of neurological signs or symptoms. Provide intensive and supportive care for severe or life-threatening neurotoxicity. For non-ICANS neurotoxicity, see Section 5.2. If ICANS is suspected, manage based on suggestions in Table 2. If CRS is suspected to occur during a neurotoxic event, administer the following: Corticosteroids due to more invasive interventions for CRS and neurotoxicity levels in Tables 1 and 2 Tocilizumab according to CRS grade in Table 1 Neurotoxic antiepileptic drugs in Table 2 [Table 38] TIFF2025507678000045.tif1661703 Dosage form and strength CARVYKTI is a cell suspension for intravenous infusion. A single dose of CARVYKTI is 0.5-1.0 × 10 6 Cell suspension of CAR-positive live T cells / kg body weight, at most 1 x 10 8 contains live CAR-positive T cells [see how to supply / store and handle (16)]. 4 Contraindications none. 5. Warnings and Precautions 5.1 Cytokine release syndrome Cytokine release syndrome, including fatal or life-threatening reactions, occurs following CARVYKTI treatment. CRS occurred in 95% (92 / 97) of patients who received siltacavtadine autorucel. 5% (5 / 97) of patients had Grade 3 or higher CRS (2019 ASTCT grade) 1 Grade 5 CRS was reported in one patient. The median duration of CRS was 7 days (range: 1-12 days). Median duration of CRS was 4 days (range: 1-40 days) in all patients except one patient who had a fatal outcome and had a 97 day duration of CRS. Among patients who experienced CRS, the most common symptoms of CRS included fever (100%), hypotension (43%), elevated aspartate aminotransferase (AST) (22%), chills (15%), elevated alanine aminotransferase (14%), and sinus tachycardia (11%). Grade ≥3 events related to CRS included elevated AST and ALT, hyperbilirubinemia, hypotension, pyrexia, hypoxia, respiratory failure, acute kidney injury, diffuse intravascular coagulation, HLH / MAS, angina, supraventricular and ventricular tachycardia, malaise, myalgia, increased C-reactive protein, ferritin, blood alkaline phosphatase, and gamma-glutamyltransferase [see Adverse Reactions (6.1)]. Identify CRS based on clinical symptoms. Assess and treat other etiologies of fever, hypoxia, and hypotension. CRS has reportedly been linked to the onset of HLH / MAS, and the physiology of the syndromes may overlap. HLH / MAS is a potentially life-threatening condition. Assess for evidence of HLH / MAS in patients with CRS who continue to experience progressive symptoms or are incurable after treatment. See Section 5.3, Hemophagocytic Lymphohistiocytosis (HLH) / Macrophage Activation Syndrome (MAS). Of the 97 patients, 69 (71%) patients received treatment for CRS with tocilizumab and / or corticosteroids after siltacabtadine autorucel injection: 44 (45%) patients received tocilizumab without corticosteroids, where 33 (34%) received a single agent, 11 (11%) received more than one dose, 24 patients (25%) received tocilizumab and corticosteroids, and 1 patient (1%) received corticosteroids only. Ensure that at least two tocilizumab drugs are available prior to CARVYKTI infusion. Monitor patients for signs and symptoms of CRS at least daily for 10 days after CARVYKTI infusion at a REMS-certified site. Monitor patients for signs or symptoms of CRS for at least 4 weeks after infusion. If the first sign of CRS appears, initiate supportive care, tocilizumab or tocilizumab and corticosteroid treatment as shown in Table 1 [see Medication and Administration (2.3)]. Advise patients to seek prompt medical attention if they experience any signs or symptoms of CRS [see Patient Advice Information (17)]. 5.2 Neurotoxicity Neurotoxicity may occur following CARVYKTI treatment and may be severe, life-threatening, or fatal. Neurotoxicity includes ICANS, neurotoxicity with Parkinsonian signs and symptoms, Guillain-Barrett syndrome, peripheral neuropathy, and cranial nerve palsies. Patients should be questioned about signs and symptoms of these neurotoxicities, as well as the delayed onset nature of some of these toxicities. Patients should be advised to seek prompt medical attention if any of these signs or symptoms of neurotoxicity occur at any time for further evaluation and management [see Patient Advice (17)]. Overall, 26% (25 / 97) of patients experienced one or more of the following neurotoxicity subtypes after siltacabtadine autocervical infusion: 2 have occurred, with 11% (11 / 97) of patients experiencing grade 3 or higher events. These neurotoxicity subtypes have also been observed in two ongoing studies [see Adverse Reactions (6.1)]. Immune effector cell-associated neurotoxicity syndrome (ICANS) Patients receiving CARVYKTI may experience fatal or life-threatening ICANS following CARVYKTI treatment, including before the onset of CRS, concurrent with CRS, after resolution of CRS, or in the absence of CRS. Among patients receiving siltacabutadine autorucel, ICANS occurred in 23% (22 / 97), including 3% (3 / 97) grade 3 or grade 4 events and 2% (2 / 97) grade 5 (fatal) events. Two patients developed persistent grade 3 and grade 1 ICANS on the last known day of life, and one patient developed persistent grade 1 ICANS at the time of death from neurotoxicity with parkinsonian features. The median duration of ICANS onset was 8 days (range: 1-28 days). Of the 22 patients, 17 (77%) ICANS resolved, with a median resolution time of 6 days (range: 2-143 days). The median duration of ICANS for all patients, including those with fatal ICANS, those with persistent ICANS at the time of death from other causes, or those with persistent ICANS on the last known day of life, was 7.5 days (range: 2-927 days). All 22 ICANS patients had CRS. ICANS onset occurred during CRS in 16 patients, before CRS onset in 3 patients, and after a CRS event in 3 patients. The most common (≥5%) symptoms of ICANS include encephalopathy (23%), aphasia (8%) and headache (6%). Monitor patients for signs and symptoms of ICANS at least daily within 10 days of CARVYKTI infusion at a REMS-certified healthcare facility. Rule out other causes of ICANS symptoms. Monitor patients for signs or symptoms of ICANS within at least 4 weeks of infusion and treat as appropriate. Neurotoxicity should be managed with supportive care and / or corticosteroids as needed [see Dosage and Administration (2.3)]. Parkinson's disease Of the 25 patients who experienced any neurotoxicity in the CARTITUDE-1 study, 5 male patients had neurotoxicity with some Parkinson's signs and symptoms different from ICANS. Neurotoxicity with Parkinson's disease has been reported in other ongoing trials of siltacabtadine autorucel. Patients have Parkinson's and non-Parkinson's symptoms including tremor, bradykinesia, involuntary movements, stereotypies, loss of locomotor activity, masked facies, apathy, flatness, fatigue, rigidity, psychomotor slowness, micrographia, dysgraphia, apraxia, lethargy, confusion, drowsiness, loss of consciousness, slow reflexes, hyperreflexia, memory loss, dysphagia, fecal incontinence, falls, stooped posture, bradygait, muscle weakness and wasting, motor dysfunction, motor and sensory loss, akinetic mutism, and frontal discharge signs. Symptoms were refractory to one or more of the following treatments attempted in one or more patients: systemic chemotherapy, intrathecal chemotherapy and steroids, dopamine agonists, systemic corticosteroids, plasma replacement, and intravenous immunoglobulin and dasatinib. One patient experienced partial resolution without Parkinsonian treatment, immunosuppressants, or chemotherapy, with residual gait disturbance. In CARTITUDE-1, the median duration of Parkinsonian symptoms in five patients was 43 days (range: 15 to 108 days) after siltacabtadine autorucel injection. One patient died of Parkinsonian neurotoxicity 247 days after siltacabtadine autorucel injection, two patients with persistent Parkinsonian disease died of infectious causes 162 and 119 days after siltacabtadine autorucel injection, and in the remaining two patients, Parkinsonian symptoms persisted for up to 530 days after siltacabtadine autorucel injection. The maximum toxicity grade in 1, 2, 1 and 1 patients was 2, 3, 4 and 5, respectively. All 5 patients had a history of previous CRS (n=grade 4, 2; n=grade 1, 3) and 4 of the 5 patients had previous ICANS (n=grade 3, 1; n=grade 1, 2). Patients should be monitored for Parkinson's signs and symptoms, which may be delayed and managed with supportive care measures. There is limited information available on the efficacy of medications to treat Parkinson's disease in improving or resolving Parkinson's symptoms following treatment with CARVYKTI. Guillain-Barrett syndrome Fatal outcomes following Guillain-Barrett syndrome (GBS) have occurred in separate ongoing studies of siltacabtadine autoreceptor using intravenous immunoglobulin therapy. Reported symptoms included encephalopathy, motor asthenia, speech impairment, and polyradiculoneuritis, consistent with the Miller-Fisher variant of GBS. Monitor for GBS. Assess patients for GBS peripheral neuropathy. Based on severity of GBS, employ supportive care measures and consider treating GBS with immunoglobulins in combination with plasma replacement. Peripheral neuropathy In CARTITUDE-1, six patients developed peripheral neuropathy. These neuropathy manifested as sensory, motor, or sensorimotor neuropathy. The median time to symptom onset was 62 days (range: 4 to 136 days), and the median duration of peripheral neuropathy was 256 days (range: 2 to 465 days), including patients with persistent neuropathy. Of these six patients, two patients developed grade 3 peripheral neuropathy and three patients experienced neuropathy resolution. The use of corticosteroid treatment in one patient was not associated with improvement in peripheral neuropathy. In other ongoing siltacabtadine autorusel trials, patients who experienced peripheral neuropathy also experienced cranial nerve palsies or GBS. Cranial nerve palsy In CARTITUDE-1, three patients (3.1%) experienced cranial nerve palsy. All three patients had VII cranial nerve palsy, and one patient had concomitant VII cranial nerve palsy. The median time to onset was 26 days (range: 21-101 days) after injection siltacabtadine autorusel. All three patients received systemic corticosteroids and resolved symptoms, and one patient received valacyclovir in addition to corticosteroids. The median time to resolution was 70 days (range: 1-79 days) after symptom onset. In the ongoing siltacabtadine autorusel trial, third and sixth cranial nerve palsies in patients with cranial nerve palsy, the occurrence of bilateral VII cranial nerve palsy, worsening of cranial nerve palsy after improvement, and the occurrence of peripheral neuropathy have also been reported. Monitor patients for signs and symptoms of cranial nerve palsy. Consider management with systemic corticosteroids based on the severity and progression of signs and symptoms. 5.3 Hemophagocytic lymphohistiocytosis (HLH) / macrophage activation syndrome (MAS) One patient (1%) developed fatal HLH 99 days after siltacabtadine autorucel infusion. The HLH event was preceded by prolonged CRS lasting 97 days. Symptoms of HLH / MAS include hypotension, hypoxia with diffuse alveolar damage, coagulopathy, cytopenias, and multiple organ dysfunction, including renal dysfunction. HLH is a life-threatening disease with high mortality if not detected and treated early. Treatment of HLH / MAS should be performed according to JAEA standards. 5.4 CARVYKTI REMS Due to the risk of CRS and neurotoxicity, CARVYKTI is only available under a restricted provision of a Risk Evaluation and Remission Strategy (REMS) (referred to as the CARVYKTI REMS program) [see Black Box Warning, Warnings, and Precautions (5.1, 5.2)]. Required Components of the CARVYKTI REMS: Healthcare facilities that dispense and administer CARVYKTI must register and comply with the requirements of the REMS. Qualified healthcare facilities must have tocilizumab readily available on-site. If CRS needs to be treated, ensure that each patient has at least two doses of troponin anti-infusion available within two hours of CARVYKTI infusion. Qualified healthcare facilities must ensure that health care providers who prescribe, dispense or administer CARVYKTI have received training in CRS and neurotoxicity management. For more information, please visit www.carvyktirems.com or call 1-844-672-0067. 5.5 Prolonged and recurrent cytopenias After lymphodepleting chemotherapy and CARVYKTI infusion, patients may exhibit prolonged and recurrent cytopenias. In the CARTITUDE-1 (N=97) study, 30% (29 / 97) patients experienced prolonged grade 3 or grade 4 neutropenia and 41% (40 / 97) patients experienced prolonged grade 3 or grade 4 thrombocytopenia, which still did not resolve by day 30 after infusion siltacabtadine autorucel. 31% (29 / 95) patients recovered from grade 3 or grade 4 neutropenia after 1 month, with a median recovery time of 1.8 months (range: 1.0 to 3.7 months) after infusion siltacabtadine autorucel. 52% (32 / 61) patients recovered from grade 3 or grade 4 thrombocytopenia after 1 month, with a median recovery time of 1.9 months (range: 1.1 to 8.5 months) after infusion siltacabtadine autorucel. One patient received autologous stem cells for hematopoietic reconstitution due to long-term thrombocytopenia. After infusion of siltacabtadine autorucel, 63% (61 / 97), 18% (17 / 97), 60% (58 / 97), and 37% (36 / 97) of patients experienced recurrent grade 3 or grade 4 neutropenia, thrombocytopenia, lymphopenia, and anemia after recovery from initial grade 3 or grade 4 cytopenia. At 60 days after infusion of siltacabtadine autorucel, 31%, 12%, and 6% of patients experienced recurrent grade 3 or higher lymphopenia, neutropenia, and thrombocytopenia, respectively, after initial recovery from grade 3 or grade 4 cytopenia [see Adverse Reactions (6.1)]. 87% (84 / 97) of patients experienced 1, 2, or 3 or more recurrent grade 3 or grade 4 cytopenias after initial recovery from grade 3 or grade 4 cytopenia. Six and 11 patients had grade 3 or grade 4 neutropenia and thrombocytopenia, respectively, at the time of death. Blood counts will be monitored before and after CARVYKTI infusion. Transfusions with growth factors and blood products will assist in managing cytopenias according to local institutional guidelines. 5.6 Infection CARVYKTI should not be administered to patients with an active infection or inflammatory disorder. Patients have developed a severe, life-threatening, or fatal infection after CARVYKTI infusion [see Adverse Reactions (6.1)]. Infections (all grades) occurred in 57 (59%) patients: 23% (22 / 97) patients had grade 3 or grade 4 infections, 17% had grade 3 or grade 4 infections with unspecified pathogens, 7% had viral infections, 1% had bacterial infections, and 1% had fungal infections. Overall, four patients had grade 5 infections: lung abscess (n=1), sepsis (n=2), and pneumonia (n=1). Monitor patients for signs and symptoms of infection before and after CARVYKTI infusion and treat patients appropriately. Administer prophylactic, advanced and / or therapeutic antimicrobials per standard institutional guidelines. Febrile ne...

Claims

1. 1. A method of treating a subject comprising administering to the subject a single infusion of a dose of a composition comprising T cells containing a chimeric antigen receptor (CAR); wherein the CAR comprises the amino acid sequence of SEQ ID NO: 17; Wherein the dose is 0.5 x 10 ６ From 1.0 x 10 ６ T cells / kg subject body weight, and wherein the method comprises completing administration of the dose of T cells to the subject within about 2.5 hours at a temperature of about 20° C. to 25° C.

2. 2. The method of claim 1, wherein the subject has relapsed or refractory multiple myeloma, and the subject has received multiple prior lines of therapy, and optionally, the subject has received three or more prior lines of therapy.

3. 3. The method of claim 1 or 2, wherein the subject has received four or more prior lines of therapy.

4. 4. The method of claim 2 or 3, wherein the prior lines of therapy include a proteasome inhibitor, an immunomodulatory agent, and an anti-CD38 monoclonal antibody.

5. The method of any one of claims 1 to 4, wherein the T cells are autologous T cells.

6. (1) administering to the subject a lymphodepleting chemotherapy regimen prior to administering the T cells to the subject, optionally comprising: (a) the lymphodepleting chemotherapy regimen comprises administering to the subject cyclophosphamide and fludarabine; (b) the lymphodepleting chemotherapy regimen comprises intravenously administering to the subject cyclophosphamide and fludarabine; (c) the lymphodepleting chemotherapy regimen comprises administering to the subject about 300 mg / m ２ and cyclophosphamide at a dose of 30 mg / m ２ daily intravenously administering fludarabine at a dose of (d) the lymphodepleting chemotherapy regimen is about 3 days; (e) the method comprises administering to the subject the lymphodepleting chemotherapy regimen for at least about 2-4 days prior to administering the T cells to the subject; or (2) administering a premedication to the subject at most 60 minutes prior to administering the T cells to the subject, and wherein the premedication includes an antipyretic and an antihistamine, optionally comprising: (a) the method comprises administering the premedication to the subject about 30-60 minutes prior to administering the T cells to the subject; (b) the antipyretic comprises paracetamol or acetaminophen; (c) the antipyretic comprises acetaminophen in a dosage of about 650 mg to 1000 mg; (d) the antihistamine comprises diphenhydramine, (e) the dose of diphenhydramine is about 25 mg to 50 mg, or equivalent; (f) the premedication is administered orally or intravenously; or (g) The method of any one of claims 1 to 5, further comprising: the premedication being free of systemic corticosteroids.

7. 7. The method of any one of claims 1-6, wherein the method comprises thawing the dose of T cells prior to administration, wherein the thawing is completed within about 15 minutes, and wherein optionally thawing the dose of T cells at a temperature of about 37°C ± 2°C.

8. The method further comprises treating cytokine release syndrome (CRS) in the subject after administering the dose of T cells, where optionally, treating CRS in the subject comprises administering to the subject an anti-cytokine agent or a corticosteroid, where optionally, (1) the anti-cytokine agent comprises a cytokine-targeting monoclonal antibody, optionally wherein the cytokine-targeting monoclonal antibody is an IL-6R inhibitor, optionally wherein the IL-6R inhibitor is tocilizumab, optionally wherein the method comprises intravenously administering tocilizumab at a dose of about 8 mg / kg in about 1 hour, optionally wherein the dose of tocilizumab does not exceed about 800 mg, optionally wherein the dose of tocilizumab does not exceed 3 doses or does not exceed 4 doses total in a 24 hour period, further optionally wherein the anti-cytokine agent further comprises an anti-cytokine agent other than tocilizumab, further optionally wherein the anti-cytokine agent further comprises a cytokine-targeting monoclonal antibody other than tocilizumab; (2) the corticosteroid comprises dexamethasone or methylprednisolone, optionally wherein the corticosteroid is dexamethasone, and further optionally wherein the method comprises administering intravenously to the subject a dose of about 10 mg of dexamethasone about every 12-24 hours, and further optionally wherein the method comprises administering intravenously to the subject a dose of about 10 mg of dexamethasone about every 12 hours, and further optionally wherein the method comprises administering intravenously to the subject a dose of about 20 mg of dexamethasone about every 6-12 hours. wherein, further optionally, the method comprises administering intravenously to the subject a dose of about 20 mg of dexamethasone about every 6 hours, wherein, optionally, the corticosteroid is methylprednisolone, wherein, further optionally, the method comprises administering intravenously to the subject a dose of about 2 mg / kg of methylprednisolone about every 12 hours, wherein, further optionally, the method comprises administering intravenously to the subject a dose of about 1-2 g of methylprednisolone about every 24 hours; or (3) The method according to any one of claims 1 to 7, wherein the method comprises administering to the subject an immune inhibitor.

9. 9. The method of claim 8, wherein CRS comprises fever, fever, hypotension, elevated aspartate aminotransferase, chills, elevated alanine aminotransferase, sinus tachycardia, hyperbilirubinemia, hypoxia, respiratory failure, acute kidney injury, disseminated intravascular coagulation and hemorrhage (e.g., retroperitoneal, intracerebral or gastrointestinal hemorrhage), hemophagocytic lymphohistiocytosis (HLH), macrophage activation syndrome (MAS), angina, supraventricular and ventricular tachycardia, dysphoria, myalgia, elevated C-reactive protein, ferritin, blood alkaline phosphatase, gamma-glutamyltransferase, organ toxicity, or any combination thereof.

10. 10. The method of claim 9, wherein CRS comprises hemophagocytic lymphohistiocytosis (HLH) or macrophage activation syndrome (MAS), wherein symptoms of HLH or MAS comprise hypotension, hypoxia with diffuse alveolar damage, coagulopathy, cytopenias, multi-organ dysfunction including renal dysfunction, or any combination thereof, and optionally, the method comprises administering a treatment to the subject to alleviate HLH or MAS.

11. 11. The method of any one of claims 1 to 10, wherein the method further comprises treating the subject for neurotoxicity after administering the dose of the T cells.

12. 12. The method of claim 11, wherein the neurotoxicity comprises immune effector cell-associated neurotoxicity syndrome (ICANS), Parkinson's disease, Guillain-Barrett's syndrome, immune-mediated myelitis, peripheral neuropathy, cranial nerve palsy, or any combination thereof.

13. The neurotoxicity comprises ICANS, and wherein the ICANS comprises encephalopathy, aphasia, headache, decreased level of consciousness, seizures, motor episodes, increased intracranial pressure (ICP), cerebral edema, or any combination thereof, and optionally, the ICANS comprises focal or generalized epileptic seizures, non-convulsive epileptic seizures on electroencephalogram (EEG), life-threatening prolonged epileptic seizures, recurrent clinical or electroepileptic seizures, deep focal motor asthenia, hemiparesis, paraparesis, focal / focal edema on neuroimaging, stupor, unconsciousness, diffuse cerebral edema on neuroimaging, decerebrate or decorticate posturing, cranial nerve VI palsy, optic disc edema, Cushing's syndrome, or any combination thereof, and optionally, (1) The method comprises administering dexamethasone intravenously to the subject at a dose of about 10 mg every about 12-24 hours for about 2-3 days; (2) The method comprises administering dexamethasone intravenously to the subject at a dose of about 10 mg about every 12 hours for about 2-3 days or more; (3) The method comprises administering dexamethasone intravenously to the subject at a dose of about 10-20 mg about every 6 hours; or (4) The method includes administering to the subject about every 24 hours a dose of about 1-2 g / day of methylprednisolone; 13. The method of claim 12, wherein, further optionally, the ICANS comprises cerebral edema, wherein, further optionally, the method comprises administering to the subject hyperventilation and hypertonicity therapy, wherein, further optionally, the method comprises administering to the subject a non-sedating anti-epileptic drug, and wherein, further optionally, the non-sedating anti-epileptic drug is levetiracetam.

14. 13. The method of claim 12, wherein the neurotoxicity comprises Parkinson's disease, optionally wherein the Parkinson's disease comprises parkinsonian symptoms or non-parkinsonian symptoms, optionally wherein the Parkinsonian symptoms or non-parkinsonian symptoms comprise tremor, bradykinesia, involuntary movements, stereotypies, loss of locomotor activity, masked facies, apathy, flatness, fatigue, rigidity, psychomotor slowness, micrographia, dysgraphia, apraxia, lethargy, confusion, drowsiness, loss of consciousness, slowed reflexes, hyperreflexia, memory loss, dysphagia, fecal incontinence, falls, hunched posture, bradygasm, muscle weakness and weakness, motor dysfunction, motor and sensory loss, akinetic mutism, frontal discharge signs, or any combination thereof, optionally wherein the method comprises administering a treatment to the subject to alleviate Parkinson's disease.

15. 13. The method of claim 12, wherein the neurotoxicity comprises Guillain-Barrett's syndrome, optionally comprising symptoms consistent with Miller-Fisher variant of Guillain-Barrett's syndrome, encephalopathy, motor asthenia, speech impairment, polyradiculoneuritis, or any combination thereof, optionally comprising administering a treatment to the subject to alleviate Guillain-Barrett's syndrome.

16. 13. The method of claim 12, wherein the neurotoxicity comprises immune-mediated myelitis, optionally wherein symptoms of immune-mediated myelitis comprise lower extremity or abdominal sensation loss with impaired sphincter control, optionally wherein the method comprises administering a treatment to the subject to alleviate the immune-mediated myelitis, optionally wherein the treatment comprises a corticosteroid or immune globulin, and optionally wherein the method comprises administering the immune globulin intravenously.

17. 13. The method of claim 12, wherein the neurotoxicity comprises peripheral neuropathy, optionally wherein the peripheral neuropathy comprises sensory neuropathy, motor neuropathy, sensory-motor neuropathy, or any combination thereof, and optionally wherein the method comprises administering a treatment to the subject to alleviate the peripheral neuropathy.

18. 13. The method of claim 12, wherein the neurotoxicity comprises cranial nerve palsy, optionally comprising cranial nerve palsy third, sixth, seventh or bilateral seventh cranial nerve palsy, and optionally wherein the method comprises administering a treatment to the subject to alleviate the cranial nerve palsy.

19. 19. The method of any one of claims 1-18, wherein the method further comprises treating prolonged or recurrent cytopenia in the subject prior to administering T cells comprising a CAR to the subject, after administering a lympho-depleting chemotherapy regimen to the subject, or after administering the dose of T cells comprising a CAR, wherein optionally the prolonged recurrent cytopenia comprises prolonged neutropenia, prolonged thrombocytopenia, recurrent neutropenia, thrombocytopenia, lymphopenia, anemia, or any combination thereof.

20. 20. The method of any one of claims 1 to 19, wherein the method further comprises treating an infection in the subject, optionally wherein the infection is viral, bacterial, fungal or due to an unindicated pathogen, optionally wherein the infection comprises lung abscess, sepsis, pneumonia, bronchopulmonary aspergillosis, Pneumocystis jirovecii pneumonia, CMV colitis (with HSV-1 hepatitis), mycotic aneurysm, cerebral aspergillosis or COVID-19 infection, optionally wherein the infection causes febrile neutropenia or subarachnoid hemorrhage, optionally wherein the method comprises administering to the subject an antimicrobial agent, optionally wherein the antimicrobial agent is an antibiotic, further optionally wherein the antibiotic is a broad spectrum antibiotic, optionally wherein the infection is viral, and further optionally wherein the method comprises administering to the subject an antiviral therapy or vaccine.

21. 21. The method of any one of claims 1-20, wherein the method further comprises treating hypogammaglobulinemia in the subject, where, optionally, after administering the dose of T cells comprising a CAR, hypogammaglobulinemia comprises a laboratory IgG level of less than about 500 mg / dL, and where, optionally, the method comprises administering a dose of intravenous immunoglobulin (IVIG) to the subject after administering the dose of T cells comprising a CAR.

22. 22. The method of any one of claims 1 to 21, wherein the method further comprises treating a hypersensitivity reaction in the subject, optionally comprising flushing, chest discomfort, tachycardia, asthma, tremors, burning sensation, an allergic reaction, or any combination thereof, and optionally wherein the method comprises administering a treatment to the subject to alleviate the hypersensitivity reaction.

23. The method of any one of claims 1 to 22, wherein the method further comprises treating a secondary malignancy in the subject.

24. 24. The method of any one of claims 1 to 23, wherein the composition further comprises an excipient selected from dimethylsulfoxide or dextran-40, optionally wherein the excipient is dimethylsulfoxide, optionally wherein the excipient is about 1-10% dimethylsulfoxide, and further optionally wherein the excipient is about 5% dimethylsulfoxide.

25. A pharmaceutical product comprising siltacabtadine autorucel suspension for intravenous infusion, said pharmaceutical product being packaged, and wherein said package contains a label, said label identifying siltacabtadine autorucel suspension as an approved pharmaceutical product for treating adult patients with relapsed or refractory multiple myeloma after receiving four or more prior lines of therapy, said prior lines of therapy including a proteasome inhibitor, an immunomodulatory agent, and an anti-CD38 monoclonal antibody.

26. A method for treating relapsed or refractory multiple myeloma in an adult patient in need thereof, comprising administering an approved pharmaceutical product comprising siltacabtadine autorucel suspension in an amount and manner indicated in the pharmaceutical product labeling of the approved pharmaceutical product.

27. A method of marketing an approved pharmaceutical product comprising siltacabtadine autorucel suspension, the method comprising marketing a pharmaceutical product in which the pharmaceutical product label of a reference product comprises instructions for treating adult patients with relapsed or refractory multiple myeloma after receiving four or more prior lines of therapy, the prior lines of therapy comprising a proteasome inhibitor, an immunomodulatory agent, and an anti-CD38 monoclonal antibody.

28. A method for authorizing the sale of a pharmaceutical product comprising siltacabtadine autorucel suspension, the pharmaceutical product label of a reference product comprising instructions for treating adult patients with relapsed or refractory multiple myeloma after receiving four or more prior lines of therapy, the prior lines of therapy comprising a proteasome inhibitor, an immunomodulatory agent, and an anti-CD38 monoclonal antibody.

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