How to Minimize Neurotoxicity Associated with Chimeric Antigen Receptor (CAR) T-Cell Therapy

Abstract

The present disclosure relates to methods for early identification and detection of neurotoxicity associated with the administration of chimeric antigen receptor (CAR) T cell therapy, as well as mitigation strategies to reduce the occurrence and / or severity of treatment-related neurotoxicity.

Description

[Technical field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims priority to U.S. Provisional Application No. 63 / 186,872, filed May 11, 2021, the entire contents of which are incorporated herein by reference. The present disclosure relates to methods of reducing neurotoxicity associated with chimeric antigen receptor (CAR) T cell therapy. [Background technology]

[0002] Multiple myeloma (MM) is an incurable malignant plasma cell disorder that accounts for approximately 10% of hematological malignancies (Rodriguez-Abreu et al., “Epidemiology of Hematological Malignancies,” Ann. Oncol. 18 Suppl. 1:i3-i8 (2007) and Rajkumar et al., “Consensus Recommendations for the Uniform Reporting of Clinical Trials: Report of the International Myeloma Workshop Consensus Panel 1,” Blood 117(8):4691-4695 (2011)). Multiple myeloma is characterized by the proliferation of neoplastic clones of plasma cells differentiated from B lymphocytes (B cells). These neoplastic clones proliferate within the bone marrow and frequently invade adjacent bones, disrupting both bone homeostasis and hematopoiesis, causing multiple destructive lesions throughout the skeleton that lead to bone pain and fractures (Chung, C., “Role of Immunotherapy in Targeting the Bone Marrow Microenvironment in Multiple Myeloma: An Evolving Therapeutic Strategy,” Pharmacotherapy 37(1):129-143(2017)).

[0003] Worldwide, there were an estimated 80,000 deaths from MM (Ferlay et al., “Cancer Incidence and Mortality Patterns in Europe: Estimates for 40 Countries in 2012,” Eur. J. Cancer 49(6):1374-1403(2013)). The estimated 5-year survival rate for MM patients is approximately 54%. Despite the availability of multiple treatments, the disease recurs in most cases and remains incurable. With each recurrence, symptoms return, quality of life worsens, and the chance and duration of a response usually decreases.

[0004] Standard treatment options for multiple myeloma include immunomodulatory imidazoles, proteasome inhibitors, anti-CD38 antibodies, and autologous stem cell transplantation, but these approaches often fail or the disease becomes refractory, so improved treatment is needed.

[0005] Autologous chimeric antigen receptor (CAR)-T cell therapy is a new form of cancer immunotherapy in which a patient's own T cells are engineered to identify and kill cancer cells within the patient. The use of a patient's own immune cells to eradicate cancer has shown great promise in the treatment of leukemia and lymphoma, and is rapidly gaining popularity in other cancers such as multiple myeloma that require alternative therapies.

[0006] Unfortunately, CAR-T cell therapy can have side effects. CAR-T therapy targeting CD19 antigen in leukemia and lymphoma can be associated with severe and fatal neurotoxicity. Neurotoxicity can occur simultaneously with cytokine release syndrome (CRS) or after CRS resolution (see Yescarta® US Product Insert (USPI) / SmPC, Kymriah® USPI / SmPC, Tecaratus® USPI / SmPC, Breyanzi® USPI, ABECMA® USPI). Immune effector cell-associated neurotoxicity syndrome (ICANS) has been well described in the literature, and symptoms and signs may be progressive and may include aphasia, altered level of consciousness, impaired cognitive abilities, motor paralysis, seizures, cerebral edema, etc. (Lee et al., “ASTCT Consensus Grading for Cytokine Release Syndrome and Neurologic Toxicity Associated with Immune Effector Cells,” Biol. Blood Marrow Transplant 25(4):625-638 (2018); Neelapu et al., “Axicabtagene Ciloleucel CAR T-Cell Therapy in Refractory Large B-Cell Lymphoma,” N. Engl. J. Med. 377:2531-2544 (2017); Santomasso et al., “Clinical and Biological Correlates of Neurotoxicity Associated with CAR T-Cell Therapy in Patients with B-Cell Lymphoma,” N. Engl. J. Med. 377:2531-2544 (2017)). Acute Lymphoblastic Leukemia,”Cancer Discov.8:958-971(2018), and Schuster et al., “Tisagenlecleucel in Adult Relapsed or Refractory Diffuse Large B Cell Lymphoma,”N.Engl.J.Med.380(1):45-56(2019)).In CD19 CAR-T experience, the incidence of ICANS is reported to range from 35% to 87% (Kymriah® USPI / SmPC, Yescarta® USPI / SmPC, Breyanzi® USPI). In BCMA CAR-T experience with idecabtagene vicleucel (also known as bb2121, hereafter referred to as ide-cel [ABECMA® USPI]), the overall incidence of neurotoxic effects ranged from 18% (Munshi et al., “Idecabtagene Vicleucel in Relapsed and Refractory Multiple Myeloma,” N. Engl. J. Med. 384:705-716 (2021) to 42% (Raje et al., “Anti-BCMA CAR T-Cell Therapy bb2121 in Relapsed or Refractory Multiple Myeloma,” N. Engl. J. Med. 384:705-716 (2021)). Myeloma,” N. Engl. J. Med. 380(18):1726-1737(2019)). Specifically, the most frequent CAR-T cell-associated neurotoxicity included encephalopathy (20%), tremor (9%), aphasia (7%), and delirium (6%), with grade 3 Parkinson's disease and grade 4 cerebral edema events also reported. The median time to onset of CAR-T cell-associated neurotoxicity after ide-cel treatment was 2 days (range: 1-42 days), and the median duration of neurotoxic events was 6 days (range: 1-578 days) (ABECMA® USPI).

[0007] Ciltacabtagene autoleucel (cilta-cel) is a genetically modified autologous T-lymphocyte (T-cell) immunotherapy for refractory multiple myeloma that binds to B-cell maturation antigen (BCMA). In clinical studies with cilta-cel, CAR-T neurotoxicity has been observed and classified as ICANS and other neurotoxicity determined to be associated with CAR-T treatment and occurring after resolution of CRS and / or ICANS. To improve treatment outcomes, it is necessary to understand the factors that lead to CAR-T cell neurotoxicity, as well as mitigation strategies to reduce and / or prevent CAR-T cell neurotoxicity.

[0008] It is an object of the present disclosure to overcome this and other deficiencies in the art. Summary of the Invention

[0009] A first aspect of the present disclosure relates to a method of reducing neurotoxicity associated with chimeric antigen receptor (CAR) T-cell therapy, comprising administering CAR-T cell therapy to a subject and determining one or more of: (i) the subject's tumor burden prior to said administration; (ii) the subject's IL-6 level at said administration; (iii) the proliferation of CAR-T cells in the subject after said administration; (iv) the persistence of CAR-T cells in the subject's peripheral blood after said administration; (v) the development of grade 2 or higher cytokine release syndrome (CRS) in the subject after said administration; (vi) the development of immune effector cell-associated neurotoxicity syndrome (ICANS) in the subject after said administration; (vii) the peak peripheral blood level of IL-6 in the subject after said administration; (viii) the peak peripheral blood level of INF-γ in the subject after said administration; and (iv) lymphocyte count in the subject after said administration. The method further comprises administering a palliative therapy to the subject based on said determination to reduce neurotoxicity associated with CAR-T cell therapy.

[0010] Another aspect of the present disclosure relates to a method of treating multiple myeloma in a subject using chimeric antigen receptor (CAR) T-cell therapy while simultaneously reducing neurotoxicity associated with the treatment, comprising administering CAR-T cell therapy to a subject with multiple myeloma, the subject having a tumor burden characterized by bone marrow plasmacytosis less than 80%, serum M protein levels less than 5 g / dL, and serum free light chain levels less than 5000 mg / L.

[0011] Another aspect of the present disclosure relates to a method of treating multiple myeloma in a subject with chimeric antigen receptor (CAR) T-cell therapy while simultaneously reducing neurotoxicity associated with said therapy, comprising administering CAR-T cell therapy to a subject having multiple myeloma and having IL-6 serum levels within the normal reference range, e.g., 0-2 pg / mL.

[0012] A further aspect of the present disclosure relates to a method of reducing neurotoxicity in a subject undergoing chimeric antigen receptor (CAR) T-cell therapy for the treatment of multiple myeloma, comprising administering to a subject undergoing CAR-T cell therapy and having symptoms of CAR-T cell therapy-associated cytokine release syndrome (CRS) or symptoms of immune effector cell-associated neurotoxicity syndrome (ICANS), an anti-inflammatory agent in an amount effective to reduce neurotoxicity in the subject.

[0013] Yet another aspect of the present disclosure relates to a method of reducing neurotoxicity in a subject undergoing chimeric antigen receptor (CAR) T-cell therapy for the treatment of multiple myeloma, the method comprising administering CAR-T cell therapy to a subject and determining whether or not the subject has a maximum CAR-T cell plasma concentration (C ) of 100% or more of the CAR-T cell after administration of the CAR-T cells (e.g., 45-100 days after administration of the CAR-T cells). max ) greater than 1,000 cells / μL and / or a viable CAR-T cell concentration in peripheral blood greater than 300 cells / μL, administering a chemotherapeutic agent to reduce neurotoxicity associated with CAR-T cell therapy.

[0014] Another aspect of the present disclosure relates to a method of reducing neurotoxicity in a subject undergoing chimeric antigen receptor (CAR) T-cell therapy for the treatment of multiple myeloma, comprising administering an IL-6 inhibitor to a subject undergoing CAR-T cell therapy and having a peak peripheral blood IL-6 level above the upper limit of normal peripheral blood IL-6 levels after CAR-T cell administration, to reduce neurotoxicity associated with CAR-T cell therapy.

[0015] Another aspect of the disclosure relates to a method of reducing neurotoxicity in a subject undergoing chimeric antigen receptor (CAR) T-cell therapy for the treatment of multiple myeloma, comprising administering an INF-γ inhibitor to a subject undergoing CAR-T cell therapy and having a peak peripheral blood INF-γ level above the upper limit of normal peripheral blood INF-γ levels after CAR-T cell administration to reduce neurotoxicity associated with the CAR-T cell therapy.

[0016] Cilta-cel is a CAR-T cell therapy using two BCMA-targeting single domain antibodies designed to bind and destroy malignant cells. In phase 1b and 2 clinical trials with cilta-cel, CAR-T neurotoxicity was observed and classified as ICANS and other neurotoxicity associated with CAR-T therapy and occurring after resolution of CRS and / or ICANS. Based on those studies, as described herein, several factors have been identified that correlate with the development of cilta-cel CAR-T cell-associated other neurotoxicity, and mitigation and management strategies have been developed based on those factors. Those strategies were employed in phase 2 and 3 studies evaluating the efficacy of cilta-cel in patients with multiple myeloma. The results indicate that by incorporating mitigation and management strategies, neurotoxic events are generally manageable in patients following treatment with cilta-cel. In fact, of more than 100 patients who received cilta-cel, only one patient experienced a neurotoxic adverse event. Those results indicate that early detection and early management of neurological adverse events leads to better outcomes of CAR-T cell therapy. [Brief description of the drawings]

[0017] [Figure 1] Schematic showing an overview of chimeric antigen receptor T cell (CAR-T) neurotoxicity in the full treatment analysis set of clinical study 68284528MMY2001. AE = adverse event, ICANS = immune effector cell-associated neurotoxicity syndrome, SOC = system organ class, TEAE = treatment-emergent adverse event.

[0018] [Diagram 2] Table showing pharmacokinetic data for subjects with peripheral blood CAR-T cell Cmax >1,000 cells / μL and CAR-T cells >300 cells / μL on day 56. CAR-T = chimeric antigen receptor T cells, Cmax = maximum plasma concentration, ID = identified, ND = not detected, TEAE = treatment-emergent adverse event. Red circle = subject with motor or neurocognitive TEAE. Note: Subject L28US10021005 demonstrated a maximum level of 1,750 cells / μL in subject's CAR+CD3+ T cells on day 14. Concentration at time of symptom onset (day 28) was 362 cells / μL. After day 56, cell counts rapidly decreased to 7 cells / μL.

[0019] [Diagram 3] 1 is a plot showing baseline IL-6 in the all treatment analysis set of clinical study 68284528MMY2001. TEAE = treatment-emergent adverse events. Note: One subject with motor and neurocognitive TEAEs did not report baseline IL-6 levels.

[0020] [Figure 4] Plot of lymphocytes (x109 / L) over time in the full treatment analysis set of clinical study 68284528MMY2001. TEAE = treatment-emergent adverse events.

[0021] [Diagram 5]1 is a plot showing peak IL-6 levels (Cmax) in the full treatment analysis set of clinical study 68284528MMY2001. Cmax = maximum concentration, M&NC = motor and neurocognitive TEAEs, NTX = neurotoxicity, TEAEs = treatment-emergent adverse events.

[0022] [Figure 6] 1 is a plot showing peak levels (Cmax) of interferon gamma in the full treatment analysis set of clinical study 68284528MMY2001. Cmax = maximum concentration, M&NC = motor and neurocognitive TEAEs, NTX = neurotoxicity, TEAEs = treatment-emergent adverse events.

[0023] [Figure 7] Frequency distribution of CD4 and CD8 T cells of naive / stem cell memory (Tn / Tscm), central memory (Tcm), effector memory (Tem), and effector memory re-expressed CD45RA (Temra) phenotypes are shown. TEAE = treatment-emergent adverse events, NTX = neurotoxicity, ICANS = immune effector cell-associated neurotoxicity events.

[0024] [Figure 8A] A-B show a comparison of the total number of administered CAR-positive viable T cells (with or without body weight normalization) (Figure 8A), predicted CAR transgene Cmax, AUC0-28d, and Tmax (Figure 8B) between subjects with and without other neurotoxicity in clinical study 68284528MMY2001. AUC0-28d = area under the CAR transgene systemic level-time curve from first dose to day 28, CAR = chimeric antigen receptor, Cmax = maximum CAR transgene systemic level, gDNA = genomic DNA, ICANS = immune effector cell-associated neurotoxicity syndrome, TEAE = treatment-emergent adverse events, Tmax = time to maximum CAR transgene. Other neurotoxicity refers to other events of CAR-T neurotoxicity not defined as ICANS. Red dots represent subjects with motor and neurocognitive TEAEs. [Figure 8B]A-B show a comparison of the total number of administered CAR-positive viable T cells (with or without body weight normalization) (Figure 8A), predicted CAR transgene Cmax, AUC0-28d, and Tmax (Figure 8B) between subjects with and without other neurotoxicity in clinical study 68284528MMY2001. AUC0-28d = area under the CAR transgene systemic level-time curve from first dose to day 28, CAR = chimeric antigen receptor, Cmax = maximum CAR transgene systemic level, gDNA = genomic DNA, ICANS = immune effector cell-associated neurotoxicity syndrome, TEAE = treatment-emergent adverse events, Tmax = time to maximum CAR transgene. Other neurotoxicity refers to other events of CAR-T neurotoxicity not defined as ICANS. Red dots represent subjects with motor and neurocognitive TEAEs.

[0025] [Figure 9] Comparison of predicted Cmax, AUC0-28d and Tmax between subjects without motor and neurocognitive TEAEs and subjects with motor and neurocognitive TEAEs is shown. AUC0-28d = area under the CAR transgene systemic level-time curve from first dose to day 28, CAR = chimeric antigen receptor, Cmax = maximum CAR transgene systemic level, gDNA = genomic DNA, ICANS = immune effector cell-associated neurotoxicity syndrome, TEAE = treatment-emergent adverse event, Tmax = time to maximum CAR transgene.

[0026] [Figure 10] 1 is a table showing a list of tumor burden and bridging therapies in subjects with motor and neurocognitive TAEA in the all-treatment analysis set of clinical study 68284528MMY2001.

[0027] [Figure 11] Forest plot of potential factors associated with motor and neurocognitive TEAEs in the all-treatment analysis set of clinical study 68284528MMY2001. ALC = absolute lymphocyte count, CBC = complete blood count, CRS = cytokine release syndrome, ICANS = immune effector cell-associated neurotoxicity syndrome, OR = odds ratio.

[0028] [Figure 12] 1 is a table listing adverse events occurring under motor and neurocognitive treatment in the full treatment analysis set of clinical study 68284528MMY2001.

[0029] [Figure 13] 1 is a plot showing baseline IL-10 in the full treatment analysis set of clinical study 68284528MMY2001. TEAE = treatment-emergent adverse events.

[0030] [Figure 14] 1 is a plot showing baseline interferon gamma in the all-treatment analysis set of clinical study 68284528MMY2001. TEAE = treatment-emergent adverse events.

[0031] [Figure 15] 1 is a plot showing baseline interleukin-2 receptor subunit alpha in the full treatment analysis set of clinical study 68284528MMY2001. TEAE = treatment-emergent adverse events. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0032] The present disclosure relates to methods for early identification and detection of neurotoxicity associated with the administration of chimeric antigen receptor (CAR) T cell therapy, as well as mitigation strategies to reduce the occurrence and / or severity of treatment-related neurotoxicity.

[0033] CAR-T neurotoxicity is classified as (i) immune effector cell-associated neurotoxicity syndrome (ICANS) and (ii) other neurotoxicity (i.e., non-ICANS). Other neurotoxicity is determined by a medical professional to be associated with CAR-T therapy and occurs after recovery from cytokine release syndrome and / or ICANS. In any embodiment, the methods described herein relate to methods of detecting and mitigating non-ICANS neurotoxicity events associated with CAR-T cell therapy, which are collectively referred to herein as "CAR-T cell-associated neurotoxicity," "CAR-T cell therapy-associated neurotoxicity," and "other neurotoxicity."

[0034] According to the methods described herein, adverse neurotoxicity events associated with administration of CAR-T cell therapy are characterized as performance and motor dysfunction treatment-emergent adverse events (TEAEs), cognitive dysfunction TEAEs, personality change TEAEs, or any combination thereof.

[0035] In any embodiment herein, the movement and motor dysfunction TEAEs specific to CAR-T cell-associated neurotoxicity (i.e., non-ICANS) include, but are not limited to, ataxia, imbalance, bradykinesia, cogwheel rigidity, dysgraphia, dyskinesia, dysmetria, essential tremor, gait disturbance, hand-eye coordination disorder, micrographia, dyskinetic movement disorder, myoclonus, Parkinson's disease, abnormal posture, resting tremor, stereotypy, and tremor. Thus, the methods described herein reduce, minimize, inhibit, or prevent any one or more of the above movement and motor dysfunction adverse events associated with CAR-T cell therapy.

[0036] In any embodiment herein, cognitive impairment TEAEs specific to CAR-T cell-associated neurotoxicity (i.e., non-ICANS) include, but are not limited to, amnesia, apraxia, slackness, cognitive impairment, confusion, decreased level of consciousness, impaired attention, encephalopathy, scattered thinking, leukoencephalopathy, loss of consciousness, memory impairment, mental impairment, altered mental status, non-infectious encephalitis, and psychomotor slowing. Thus, the methods described herein reduce, minimize, inhibit, or prevent any one or more of the above cognitive impairment adverse events associated with CAR-T cell therapy.

[0037] In any embodiment herein, personality change TEAEs specific to CAR-T cell-associated neurotoxicity (i.e., non-ICANS) include, but are not limited to, flat affect, personality changes, or reduced facial expression. Thus, the methods described herein reduce, minimize, inhibit, or prevent any one or more of the above personality adverse events associated with CAR-T cell therapy.

[0038] Accordingly, a first aspect of the present disclosure relates to a method of reducing neurotoxicity associated with chimeric antigen receptor (CAR) T-cell therapy, comprising administering a CAR-T cell therapy to a subject and determining one or more factors associated with the development of CAR-T cell neurotoxicity, including (i) the subject's tumor burden prior to said administration, (ii) the subject's IL-6 level at the time of said administration, (iii) the proliferation of CAR-T cells in the subject following said administration, (iv) the persistence of CAR-T cells in the subject's peripheral blood following said administration, (v) the development of grade 2 or higher cytokine release syndrome (CRS) in the subject following said administration, (vi) the development of immune effector cell-associated neurotoxicity syndrome (ICANS) in the subject following said administration, (vii) the peak peripheral blood level of IL-6 in the subject following said administration, (viii) the peak peripheral blood level of INF-γ in the subject following said administration, and (iv) the lymphocyte count in the subject following said administration.

[0039] CAR-T cell therapy is utilized in the treatment of a variety of conditions, including multiple myeloma, various B-cell lymphomas (e.g., mantle cell lymphoma, follicular lymphoma, high-grade B-cell lymphoma, malignant B-cell lymphoma, large B-cell lymphoma, primary mediastinal large B-cell lymphoma, diffuse large B-cell lymphoma, and B-cell precursor acute lymphoblastic leukemia (ALL). Neurotoxicity is a side effect associated with CAR-T cell administration under the above conditions, and thus the methods described herein are applicable to reduce neurotoxicity, particularly non-ICANS neurotoxicity, in subjects receiving CAR-T cell therapy for any of the above conditions. In any embodiment, the subject treated according to the methods described herein receives CAR-T cell therapy for the treatment of multiple myeloma.

[0040] In any embodiment, the subject treated according to the methods described herein has multiple myeloma and receives CAR-T cell therapy in which the CAR targets B-cell maturation agent (BCMA). In any embodiment, the subject has multiple myeloma and receives idecabutagen bicelucel or cilta-cel. In any embodiment, the subject has multiple myeloma and receives cilta-cel, as described in detail in WO2017 / 025038 (Fan et al.) and WO2018 / 028647 (Fan et al.), the entire contents of which are incorporated herein by reference. Suitable methods of administering CAR-T cell therapy for the treatment of various lymphoid neoplasms are known in the art (see, e.g., Cerrano et al., “The Advent of CAR T-Cell Therapy for Lymphoproliferative Neoplasms: Integrating Research Into Clinical Practice,” Front. Immunol. 11(888) (2020), the entire contents of which are incorporated herein by reference). Methods of administering BCMA CAR-T cell therapy for the treatment of multiple myeloma are also known in the art and described herein (see, e.g., WO2017 / 025038 (Fan et al.) and WO2018 / 028647 (Fan et al.), the entire contents of which are incorporated herein by reference).

[0041] In any embodiment, the method of reducing neurotoxicity associated with CAR-T cell therapy comprises determining at least two of the factors, determining at least three of the factors, determining at least four of the factors, determining at least five of the factors, determining at least six of the factors, determining at least seven of the factors, or determining at least eight of the factors. In some embodiments, the method of reducing neurotoxicity associated with CAR-T cell therapy comprises determining all nine of the factors.

[0042] According to this aspect of the disclosure, depending on the results of the analysis of the one or more factors above, one or more mitigation strategies described herein are used to reduce, prevent, or minimize neurotoxicity associated with CAR-T cell therapy. In some embodiments, a plurality of mitigation strategies described herein are used to reduce neurotoxicity associated with CAR-T cell therapy. In some embodiments, the mitigation strategy comprises administration of a therapeutic agent. In some embodiments, the mitigation strategy comprises further observation and monitoring of the subject to detect early signs of neurotoxicity.

[0043] For purposes of this disclosure, "reducing" neurotoxicity associated with CAR-T cell therapy includes, but is not limited to, reducing motor, cognitive, and / or personality adverse events occurring following CAR-T cell therapy, reducing the magnitude of some or all of the motor, cognitive, and / or personality adverse events associated with the treatment, stabilizing (i.e., not worsening) some or all of the motor, cognitive, and / or personality adverse events, delaying the onset or slowing the progression of some or all of the motor, cognitive, and personality adverse events, or ameliorating some or all of the motor, cognitive, and personality adverse events. In some embodiments, the methods of reducing neurotoxicity described herein are preemptive in nature, i.e., preventing CAR-T cell-associated neurotoxicity. Prevention may include complete protection from CAR-T cell-associated neurotoxicity events or may include prevention of the progression of CAR-T cell-associated neurotoxicity. For example, prevention may not mean completely eliminating any level of neurotoxicity events associated with CAR-T cell therapy, but rather preventing symptoms to a clinically significant or detectable level. Prevention of CAR-T cell-associated neurotoxicity can also mean prevention of progression to a later stage of neurotoxicity compared to the progression experienced by a subject who is not administered the palliative treatment described herein.

[0044] In any embodiment, the method of reducing neurotoxicity associated with CAR-T cell therapy includes determining the subject's tumor burden before administering CAR-T cell therapy. As described herein, high tumor burden in a subject at the time CAR-T cell therapy is administered is associated with the development of CAR-T cell-associated neurotoxicity. The subject's tumor burden can be determined using methods standard in the art for a particular tumor. For example, if the subject has multiple myeloma, the subject's tumor burden can be determined by measuring the subject's bone marrow plasmacytosis level, serum M protein level, and / or serum free light chain level.

[0045] Bone marrow plasmacytosis, i.e., the percentage of bone marrow cells that are plasma cells, may be measured in bone marrow biopsies or aspirates. The number of plasma cells may be determined by immunohistochemistry or flow cytometry techniques using a combination of identified antibodies, including but not limited to antibodies against CD138 or VS38c, Bcl-2, CD79a, and CD20. Plasma cells typically account for about 2%-3% of bone marrow cells. According to the present disclosure, a finding that plasma cells account for 30% or more, 40% or more, 50% or more, 60% or more, 70% or more, or 80% or more of the subject's bone marrow cells is indicative of a high tumor burden in the subject. In any embodiment, a finding that plasma cells account for 80% or more of the subject's bone marrow cells is indicative of a high tumor burden in the subject.

[0046] M protein, also called myeloma protein, monoclonal immunoglobulin, M spike, or paraprotein, is a bone marrow-derived antibody released from myeloma cells and can be detected and quantified using serum or urine electrophoresis, blood or urine immunofixation electrophoresis, or quantitative immunoglobulin testing. A finding that a subject has a serum M protein level of 2 g / dL or greater, 3 g / dL or greater, 4 g / dL or greater, 5 g / dL or greater is indicative of a high tumor burden in the subject. In any embodiment, a finding that a subject has a serum M protein level of 5 g / dL or greater is indicative of a high tumor burden in the subject.

[0047] In some cases, myeloma cells do not produce the entire M protein (i.e., intact immunoglobulin) but instead release only light chains, i.e., the kappa (κ) and lambda (λ) free immunoglobulin light chains. These free light chains, also called Bence Jones proteins, can be detected in the blood or urine by light chain-specific antibodies (see, e.g., Bradwell et al., Highly Sensitive, Automated Immunoassay for Immunoglobulin Free Light Chains in Serum and Urine,” Clin. Chem. 47(4):673-80 (2001), the entire contents of which are incorporated herein by reference). The normal ranges for the concentrations of kappa and lambda chains in serum in healthy individuals are about 3.3-19.4 mg / L and about 5.7-26.6 mg / L for kappa and lambda chains, respectively, with a kappa / lambda ratio of 0.26-1.65. A finding that the subject has a serum free light chain level greater than 100 mg / L, greater than 500 mg / L, greater than 1000 mg / L, greater than 2000 mg / L, greater than 3000 mg / L, greater than 4000 mg / L, greater than 5000 mg / L or greater is indicative of a high tumor burden in the subject. In any embodiment, a finding that the subject has a serum free light chain level greater than 5000 mg / L is indicative of a high tumor burden in the subject.

[0048] If a subject is determined to have a high tumor burden at the time CAR-T cell therapy is administered, the risk of developing neurotoxicity associated with CAR-T cell therapy can be mitigated by administering a bridge therapy to reduce the subject's tumor burden prior to administration of CAR-T cell therapy. A "bridge therapy" is any therapy suitable for reducing the tumor burden between leukapheresis, i.e., when the subject's T cells are harvested, and conditioning, i.e., when conditioning chemotherapy is administered to the subject in anticipation of receiving CAR-T cell therapy. In some embodiments, the bridge therapy is administered in an amount effective to reduce the subject's tumor burden to achieve a tumor burden characterized by bone marrow plasmacytosis of less than 80%, serum M protein levels of less than 5 g / dL, and serum free light chain levels of less than 5000 mg / L prior to administration of CAR-T cell therapy. In some embodiments, the bridging therapy is administered in an amount effective to reduce the subject's tumor burden to achieve bone marrow plasmacytosis less than 50%, serum M protein levels less than 3 g / dL, and serum free light chain levels less than 3000 mg / L prior to administration of the CAR-T cell therapy.

[0049] Suitable bridging therapies include, but are not limited to, chemotherapeutic agents, immunomodulatory agents, proteasome inhibitors, or any combination thereof.

[0050] In any embodiment, the bridge therapy is chemotherapy. Suitable chemotherapeutic agents include alkylating agents such as cyclophosphamide (Cytoxan), melphalan, melflufen (Pepaxto®), and bendamustine (Treanda®), as well as topoisomerase inhibitors such as etoposide (VP-16) and doxorubicin (adriamycin, Doxil).

[0051] In any embodiment, the bridge therapy is a proteasome inhibitor. Proteasome inhibitors suitable for administration as bridge therapy include, but are not limited to, bortezomib (Velcade®), carfilzomib (Kyprolis®), and ixazomib (Ninlaro®). In any embodiment, the proteasome inhibitor may be administered with dexamethasone, a combination of dexamethasone and lenalidomide, or a combination of dexamethasone and cyclophosphamide.

[0052] In any embodiment, the bridge therapy is an immunomodulator. Suitable immunomodulators for subjects with multiple myeloma include, but are not limited to, CD38 inhibitors and SLAMF7 inhibitors. Suitable CD38 inhibitors include anti-CD38 monoclonal antibodies, such as daratumumab (Darzalex®) and isatuximab (Sarclisa®). Suitable SLAMF7 inhibitors include the anti-SLAMF7 monoclonal antibody, elotuzumab (Empliciti®). Other suitable immunomodulators include, but are not limited to, lenalidomide (Revlimid®), pomalidomide (Pomalyst®), thalidomide, and combinations thereof. In any embodiment, the aforementioned immunomodulators may be administered in combination with dexamethasone.

[0053] Other suitable bridging therapies include, but are not limited to, histone deacetylase (HDAC) inhibitors such as panobinostat (Farydak®), nuclear export inhibitors such as selinexor (Xpovio®), and antibody drug conjugates such as belantamab mafodotin-blmf (Blenrep).

[0054] In some embodiments, the bridge therapy comprises any combination of the aforementioned chemotherapeutic agents, proteasome inhibitors, or immunomodulators. Suitable combination therapies include lenalidomide (or pomalidomide or thalidomide) and dexamethasone; carfilzomib (or ixazomib or bortezomib), lenalidomide, and dexamethasone; bortezomib (or carfilzomib), cyclophosphamide, and dexamethasone; elotuzumab, lenalidomide, and dexamethasone; daratumumab, lenalidomide, and dexamethasone; isatuximab, lenalidomide, and dexamethasone; bortezomib, liposomal doxorubicin, and dexamethasone; panobinostat, bortezomib, and dexamethasone; elotuzumab, bortezomib, and dexamethasone. including, but not limited to, zomib, and dexamethasone; melphalan and prednisone (MP) (with or without thalidomide or bortezomib); vincristine, doxorubicin (adriamycin), and dexamethasone (referred to as VAD); dexamethasone, cyclophosphamide, etoposide, and cisplatin (referred to as DCEP); dexamethasone, thalidomide, cisplatin, doxorubicin, cyclophosphamide, and etoposide (referred to as DT-PACE) (with or without bortezomib); and selinexor, bortezomib, and dexamethasone.

[0055] In any embodiment, the method of reducing neurotoxicity associated with CAR-T cell therapy includes determining the subject's blood IL-6 level at the time of administration of the CAR-T cell therapy. If the subject's blood IL-6 level is above the upper limit of normal, an IL-6 inhibitor is administered as a palliative to reduce IL-6 levels prior to administration of the CAR-T therapy. Suitable IL-6 inhibitors include, but are not limited to, tocilizumab (Actemra®), sarulumab (Kevzara®), siltuximab (Sylvant®), and clazakizumab (Atal and Fatima, "IL-6 Inhibitors in the Treatment of Serious COVID-19: A Promising Therapy," Pharmaceutical Medicine 34:223-231 (2020), the entire contents of which are incorporated herein by reference). The normal reference range for IL-6 is 0-2 pg / ml (see, e.g., Wang et al., IL-6 Signaling in Peripheral Blood T Cells Predicts Clinical Outcome in Breast Cancer, Cancer Research 77(5):1119-1126 (2016), the entire contents of which are incorporated herein by reference). Thus, if a subject's blood IL-6 level exceeds 2 pg / mL, tocilizumab or a similarly effective IL-6 inhibitor is administered to the subject to reduce IL-6 levels to within the reference range of 0-2 pg / mL prior to administration of CAR-T cell therapy.

[0056] In any embodiment, the method of reducing neurotoxicity associated with CAR-T cell therapy includes determining the subject's peak peripheral blood level of IL-6 after administration of CAR-T cell therapy. If the subject's blood IL-6 level exceeds the upper limit of normal, an IL-6 inhibitor, such as tocilizumab, saluliumab, siltuximab, or clazakizumab, is administered as a palliative to reduce IL-6 levels. As noted above, the reference range for normal blood IL-6 levels is 0-2 pg / ml. Thus, if the subject's peak peripheral blood IL-6 level exceeds 2 pg / mL, tocilizumab or a similarly effective IL-6 inhibitor is administered to the subject in an amount effective to reduce IL-6 levels to within the reference range of 0-2 pg / mL.

[0057] In any embodiment, the method of reducing neurotoxicity associated with CAR-T cell therapy includes determining the peak peripheral blood INF-γ level of the subject after administration of the CAR-T cell therapy. If the subject's blood INF-γ level is above the upper limit of normal, an INF-γ inhibitor is administered as a palliative agent to reduce the INF-γ level to an acceptable level. Suitable INF-γ inhibitors, such as IFN-γ monoclonal antibodies known in the art, emapalumab (Gamifant®) (Vallurupalli and Berliner, "Emapalumab for the Treatment of Relapsed / Refractory Hemophagocytic Lymphohistiocytosis," Blood 134(21):1783-1786 (2019), the entire contents of which are incorporated herein by reference), are suitable for use in accordance with the methods described herein. Normal blood INF-γ levels are less than about 2.0 pg / mL. Therefore, if a subject's INF-γ peak value in peripheral blood exceeds 2 pg / mL, emapalumab or a similarly effective INF-γ inhibitor is administered to the subject to reduce the INF-γ level to the reference range of less than 2 pg / mL.

[0058] In any embodiment, the method of reducing neurotoxicity associated with CAR-T cell therapy includes determining the lymphocyte count of the subject after administration of CAR-T cell therapy. If the lymphocyte count of the subject is above the upper limit of normal about 2 weeks, about 3 weeks, about 4 weeks, about 5 weeks, about 6 weeks, about 7 weeks, about 8 weeks, about 9 weeks, or about 10 weeks after administration of CAR-T therapy, an anti-inflammatory agent is administered as a palliative to reduce the lymphocyte count. Suitable anti-inflammatory agents include, but are not limited to, IL-6 inhibitors, IL-1 receptor antagonists (e.g., anakinra), tyrosine kinase inhibitors (e.g., dasatinib), steroids, and methotrexate. A normal lymphocyte count is between 0.8 and 3.0×10 9 / L. Therefore, the subject's lymphocyte count is in the range of 3.0 × 10 9 / L, lymphocyte count is 0.8–3.0 × 10 9 An anti-inflammatory agent is administered to the subject to reduce the lymphocyte count to approach or within the reference range of 1 / L.

[0059] In any embodiment, the method of reducing neurotoxicity associated with CAR-T cell therapy includes determining the proliferation and persistence of CAR-T cells in a subject following administration of CAR-T cell therapy. When proliferation of CAR-T cells in a subject is high (maximum plasma concentration of CAR-T cells (C maxIf the subject has high CAR-T cell proliferation (defined as CAR-T cell levels in peripheral blood greater than 1,000 cells / μL), a palliative therapy is administered to reduce the number of CAR-T cells in the subject. Similarly, if CAR-T cell persistence is high (defined as CAR-T cell levels in peripheral blood greater than 300 cells / μL about 45-100 days after treatment), a palliative therapy is administered to reduce the number of CAR-T cells remaining in the subject. Suitable palliative therapies for reducing the number of CAR-T cells in the subject include, but are not limited to, chemotherapeutic agents and anti-inflammatory agents. Suitable chemotherapeutic agents are described above and include alkylating agents and topoisomerase inhibitors. Suitable anti-inflammatory agents include, but are not limited to, IL-6 inhibitors, IL-1 receptor antagonists (e.g., anakinra), tyrosine kinase inhibitors (e.g., dasatinib), steroids, and methotrexate. If the subject has high CAR-T cell proliferation, a palliative agent is administered in an amount and for a duration effective to reduce the plasma concentration of CAR-T cells to less than 1,000 cells / μL. If the subject's CAR-T cells have high peripheral persistence (e.g., about 45-100 days after treatment), the palliative agent is administered in an amount and for a duration effective to reduce the number of CAR-T cells in the peripheral blood to less than 300 cells / μL.

[0060] In any embodiment, the method of reducing neurotoxicity associated with CAR-T cell therapy includes monitoring the subject for the onset of cytokine release syndrome (CRS). As used herein, the term "cytokine release syndrome" or "CRS" refers to a supraphysiological response following immunotherapy that causes activation or engagement of endogenous or infused T cells and / or other immune effector cells (e.g., Lee et al., "ASTCT Consensus Grading for Cytokine Release Syndrome and Neurologic Toxicity Associated with Immune Effector Cells," Biol. Blood Marrow Transplant. 25(4):625-638 (2019), the entire contents of which are incorporated herein by reference). As described herein, antecedent or concurrent CRS correlates with the onset of CAR-T cell-associated neurotoxicity. In particular, subjects who develop grade 2 or higher CRS are predisposed to developing CAR-T cell therapy-associated neurotoxicity and need to be aggressively treated with anti-inflammatory agents to resolve CRS. Suitable therapeutic approaches for the treatment of CRS are disclosed below.

[0061] Symptoms of CRS include, but are not limited to, fever, fatigue, myalgia, arthralgia, headache, nausea / vomiting, diarrhea, skin rash, tachypnea, hypoxia, pulmonary edema, elevated D-dimer levels, hypofibrinogenemia, renal dysfunction (e.g., azotemia), hepatic dysfunction (e.g., transaminitis and / or hyperbilirubinemia), and cardiovascular dysfunction (e.g., tachycardia, hypotension, capillary leak, pulse pressure dilation, cardiac output regulation). Grading of CRS may be performed using any of the known and acceptable rating scales described in Riegler et al., “Current Approaches in the Grading and Management of Cytokine Release Syndrome after Chimeric Antigen Receptor T-cell Therapy,” Ther. Clin. Risk Manag. 15:323-335 (2019), the entire contents of which are incorporated herein by reference.

[0062] In any embodiment, the method of reducing neurotoxicity associated with CAR-T cell therapy includes monitoring the subject for the onset of immune effector cell-associated neurotoxicity syndrome (ICANS) following administration of CAR-T cell therapy. As used herein, the term "immune effector cell-associated neurotoxicity syndrome" or "ICANS" refers to a disease characterized by a pathological process involving the central nervous system following immunotherapy that results in the activation or engagement of endogenous or infused T cells and / or other immune effector cells (see, e.g., Lee et al., "ASTCT Consensus Grading for Cytokine Release Syndrome and Neurologic Toxicity Associated with Immune Effector Cells," Biol. Blood Marrow Transplant. 25(4):625-638 (2019), the entire contents of which are incorporated herein by reference). As described herein, pre-onset or concurrent ICANS correlates with the onset of neurotoxicity associated with CAR-T cells. In particular, subjects who develop any level of ICANS are prone to developing neurotoxicity associated with CAR-T cell therapy and need to be aggressively treated with anti-inflammatory agents to resolve ICANS. Suitable therapeutic agents for the treatment of ICANS are disclosed below.

[0063] Symptoms of ICANS include, but are not limited to, delirium, encephalopathy, aphasia (progressing to expressive aphasia), fatigue, poor concentration, agitation, tremors, seizures, dysgraphia, mild difficulties with expressive language, apraxia, and cerebral edema. ICANS grading can be performed using any of the known and acceptable rating scales described in Lee et al., “ASTCT Consensus Grading for Cytokine Release Syndrome and Neurologic Toxicity Associated with Immune Effector Cells,” Biol. Blood Marrow Transplant 25:625-638 (2019), the entire contents of which are incorporated herein by reference. In any embodiment, ICANS is graded according to the Immune Effector Cell-Associated Encephalopathy (ICE) Assessment Tool (ICE-Tool) described herein and in Lee et al. Biol. Blood Marrow Transplant 25:625-638 (2019), the contents of which are incorporated herein by reference in their entirety.

[0064] Suitable methods and therapies for treating CRS and ICANS known in the art (see, e.g., Riegler et al., “Current Approaches in the Grading and Management of Cytokine Release Syndrome after Chimeric Antigen Receptor T-cell Therapy,” Ther. Clin. Risk Manag. 15:323-335 (2019), the entire contents of which are incorporated herein by reference) are suitable for use in accordance with the methods described herein. The primary therapeutic agents administered to alleviate CRS are anti-inflammatory agents, including, but not limited to, IL-6 inhibitors (e.g., tocilizumab, saluliumab, siltuximab, or clazakizumab), IL-1 inhibitors (e.g., anakinra), Janus kinase 1 / 2 inhibitors (e.g., ruxolitinib), and corticosteroids such as dexamethasone, methylprednisolone, and hydrocortisone. In some embodiments, a combination of the above-mentioned therapies, such as an IL-6 inhibitor and a steroid, is administered to the subject to actively treat and resolve CRS. Other therapeutic agents suitable for treating CRS include antithymocyte globulin and cyclophosphamide. Treatment of ICANS primarily involves administration of corticosteroids (dexamethasone, methylprednisolone, hydrocortisone), but may also include administration of anti-inflammatory agents (i.e., IL-6 inhibitors) if CRS occurs concomitantly.

[0065] The method disclosed herein for reducing neurotoxicity associated with CAR-T cell therapy includes determining one or more of the factors associated with the onset of CAR-T cell neurotoxicity described above. However, the method further includes monitoring the subject for symptoms of agraphia, micrographia, dysgraphia, or any combination thereof after administration of CAR-T cell therapy. The monitoring provides a means for early detection and early identification of neurotoxicity associated with CAR-T cell therapy. Symptoms of agraphia (i.e., loss of ability to write), micrographia (i.e., a disorder characterized by abnormally small, irregular writing, or increasingly small writing), and dysgraphia (i.e., characterized by difficulty or inconsistency in spacing between letters and words, spelling errors, incomplete words, missing words or letters), or any combination thereof, can be monitored by providing the subject with periodic writing assessments. Writing assessments are performed prior to administration of CAR-T cell therapy and periodically after administration of CAR-T cell therapy to assess and detect changes as early as possible. Continued evaluation may be performed at 1 month, 2 months, 3 months, 4 months, 5 months, 6 months, 7 months, 8 months, 9 months, 10 months, 11 months, 12 months, or more than 12 months after administration of the CAR-T cell therapy.

[0066] If the changes in the writing assessment indicate the onset of agraphia, micrographia, and / or dysgraphia, the individual is subject to further neurological evaluation to assess neurotoxicity and identify / rule out non-CAR-T cell therapy causes of the observed neurotoxicity (e.g., infection). The neurological evaluation should include any one or more of the following: (i) evaluation of a cerebrospinal fluid sample from the subject to analyze for the presence of infection, leptomeningeal disease, paraneoplastic syndrome, or a combination thereof; (ii) measuring the subject's serum levels of human herpesvirus (HHV)-6, HHV-7, or both; (iii) measuring the subject's serum thiamine levels; (iv) imaging the subject's brain by positron emission tomography or magnetic resonance imaging; and (v) performing an electroencephalogram (EEG).

[0067] Another aspect of the present application relates to a method of treating multiple myeloma in a subject using chimeric antigen receptor (CAR) T-cell therapy while minimizing neurotoxicity associated with said therapy, comprising administering CAR-T cell therapy to a subject with multiple myeloma, wherein the subject has a tumor burden characterized by bone marrow plasmacytosis less than 80%, serum M protein levels less than 5 g / dL, and serum free light chain levels less than 5000 mg / L.

[0068] In some embodiments, the subject has a tumor burden characterized by bone marrow plasmacytosis less than 50%, serum M protein levels less than 3 g / dL, and serum free light chain levels less than 3000 mg / L.

[0069] If the subject's tumor burden does not meet the necessary criteria for receiving CAR-T cell therapy (e.g., because the subject's tumor burden is characterized by bone marrow plasmacytosis greater than 80%, serum M protein level greater than 5 g / dL, or serum free light chain level greater than 5000 mg / L), a bridge therapy may be administered to the subject in an amount effective to reduce the subject's tumor burden to a level acceptable for receiving CAR-T cell therapy. For example, the bridge therapy is administered in an amount and for a duration effective to reduce the subject's tumor burden such that the subject's bone marrow plasmacytosis is less than 80%, serum M protein level is less than 5 g / dL, and serum free light chain level is less than 5000 mg / L. Suitable bridge therapies are described above and include, but are not limited to, chemotherapeutic agents (i.e., alkylating agents and topoisomerase inhibitors), immunomodulatory agents, proteosome inhibitors, and any combination thereof.

[0070] Another aspect of the present disclosure relates to a method of treating multiple myeloma in a subject with CAR-T cell therapy, and simultaneously reducing neurotoxicity associated with said therapy, comprising administering CAR-T cell therapy to a subject having multiple myeloma and having an IL-6 serum level within the normal reference range of 0-2 pg / mL.

[0071] If the subject's IL-6 serum levels are not within the normal reference range of IL-6 levels, IL-6 inhibitor therapy is administered in an amount and for a duration effective to reduce IL-6 levels to normal reference levels prior to administration of CAR-T cell therapy. Suitable IL-6 inhibitors are described above.

[0072] Another aspect of the present disclosure relates to a method for reducing the neurotoxicity associated with CAR-T cell therapy in a subject undergoing CAR-T cell therapy for the treatment of multiple myeloma. The method comprises administering an anti-inflammatory agent to a subject undergoing CAR-T cell therapy and having symptoms of CAR-T cell therapy-associated cytokine release syndrome (CRS) or immune effector cell-associated neurotoxicity syndrome (ICANS) in an amount effective to reduce the neurotoxicity in the subject. Suitable anti-inflammatory agents include, but are not limited to, IL-6 inhibitors (e.g., tocilizumab, saruliumab, siltuximab, or clazakizumab), IL-1 inhibitors (e.g., anakinra), Janus kinase 1 / 2 inhibitors (e.g., ruxolitinib), and corticosteroids such as dexamethasone, methylprednisolone, and hydrocortisone mentioned above.

[0073] Another aspect of the present disclosure relates to a method of reducing neurotoxicity associated with CAR-T cell therapy in a subject undergoing CAR-T cell therapy for the treatment of multiple myeloma, comprising administering a chemotherapeutic agent to a subject undergoing CAR-T cell therapy and having a high level of CAR-T cell proliferation, thereby reducing the level of CAR-T cells and associated neurotoxicity in the subject. As described above, a high level of CAR-T cell proliferation is associated with a high maximum plasma concentration (C max ) is defined as a peripheral blood concentration of greater than 1,000 cells / μL. Similarly, if a subject receives CAR-T cell therapy and has high levels of persistence of CAR-T cells, a chemotherapeutic agent is administered to the subject to reduce the persistence of CAR-T cell levels and associated neurotoxicity. As described above, high levels of persistence of CAR-T cells are defined as a peripheral blood concentration of greater than 300 cells / μL about 45-65 days after receiving CAR-T cell therapy.

[0074] Another aspect of the present disclosure relates to a method for reducing the neurotoxicity associated with CAR-T cell therapy in a subject undergoing CAR-T cell therapy for the treatment of multiple myeloma, comprising administering an IL-6 inhibitor to a subject undergoing CAR-T cell therapy and experiencing an increase in IL-6 levels in peripheral blood that exceeds the upper limit of normal IL-6 levels, to reduce the neurotoxicity associated with CAR-T cell therapy.Suitable IL-6 inhibitors are described above.

[0075] Another aspect of the present disclosure relates to a method for reducing the neurotoxicity associated with CAR-T cell therapy in a subject undergoing CAR-T cell therapy for the treatment of multiple myeloma, comprising administering an INF-γ inhibitor to a subject undergoing CAR-T cell therapy and experiencing an increase in INF-γ levels in peripheral blood that exceeds the upper limit of normal INF-γ levels, to reduce the neurotoxicity associated with CAR-T cell therapy. Suitable INF-γ inhibitors are described above. EXAMPLES

[0076] Example 1 - Clinical Study 68284528MMY2001 Study design Study 68284528MMY2001 was a Phase 1b-2, open-label, multicenter study designed to evaluate the safety and efficacy of cilta-cel in adult subjects with relapsed or refractory multiple myeloma (RRMM). The study consisted of two parts: Phase 1b and Phase 2. In the Phase 1b part, a staggered enrollment strategy was used to confirm the dose levels recommended for study in Phase 2 (RP2D). In the Phase 2 part, additional subjects were treated with cilta-cel using RP2D to further characterize safety and efficacy.

[0077] Enrolled subjects underwent apheresis to obtain peripheral blood mononuclear cells (PBMCs), and selected subject T cells from the apheresis product were used to prepare cilta-cel. After cilta-cel production and product release, subjects underwent a 3-day lymphodepleting conditioning regimen with cyclophosphamide and fludarabine, followed by cilta-cel infusion 5-7 days after conditioning initiation. Some study subjects received bridge therapy between apheresis and initiation of lymphodepleting chemotherapy to maintain disease stability. Subjects were closely monitored for safety and disease assessments during the post-cilta-cel infusion period (days 1-100). Assessments were less frequent during the post-treatment period (day 101 through study completion), with safety and disease assessments occurring every 28 days. Survival status and subsequent anticancer therapy information were collected every 16 weeks after disease progression. The study will be completed 2 years after the last subject received their first dose of cilta-cel. Subjects treated with cilta-cel will then be enrolled in a long-term follow-up study (study 68284528MMY4002) for continued monitoring for up to 15 years.

[0078] Safety Analysis Set The primary analysis population for all safety analyses was the overall treatment population, which included all 97 subjects who received an infusion of cilta-cel as of the clinical cut-off date.

[0079] Chimeric antigen receptor T cell (CAR-T) neurotoxicity CAR-T neurotoxicity was classified as immune effector cell-associated neurotoxicity syndrome (ICANS) and / or other neurotoxicity associated with CAR-T treatment and occurring after recovery from ICANS. Other neurotoxicity was classified as motor and neurocognitive adverse events.

[0080] In study 68284528MMY2001, 20 subjects (20.6%) experienced treatment-emergent CAR-T neurotoxicity events. Nine subjects (9.3%) experienced grade 3 or 4 events, and one subject (1.0%) experienced a grade 5 event. Note that ICANS and other neurotoxicity are not mutually exclusive, as eight subjects (8.2%) experienced both ICANS and other neurotoxicity of any grade, as shown in Figure 1.

[0081] Immune effector cell-associated neurotoxicity Sixteen subjects (16.5%) experienced ICANS (Table 1). Ten subjects (10.3%) had a maximum grade 1 event, four subjects (4.1%) had a maximum grade 2 event, and one subject (1.0%) had a maximum grade 3 or grade 4 event, respectively. No subjects experienced a grade 5 ICANS event. [Table 1] TIFF2024518549000003.tif136165TIFF2024518549000004.tif135165

[0082] For two subjects in Phase 1b, the term reported was CAR-T-associated encephalopathy syndrome (CRES) and was coded to ICANS according to the Medical Dictionary for Regulatory Activities (MedDRA v 23) (Table 2). Because the events were reported before the publication of the American Society for Transplantation and Cellular Therapy (ASTCT) consensus criteria, they were graded according to the National Cancer Institute's Common Terminology Criteria for Adverse Events (NCI-CTCAE v 5.0) and are included as ICANS in Table 1. The highest grade for those two subjects was grade 1 (one subject) and grade 3 (one subject) according to NCI-CTCAE v 5.0. [Table 2] TIFF2024518549000006.tif94165

[0083] The median time from cilta-cel infusion to the first onset of ICANS was 8.0 days (range: 3-12 days), and the median duration was 4 days (range: 1-12 days). Treatment-emergent clinical manifestations of ICANS included aphasia, bradyslexia, dysgraphia, encephalopathy, decreased level of consciousness, and confusional state.

[0084] At clinical cutoff, all 16 subjects who experienced ICANS had recovered. All subjects who experienced ICANS also experienced CRS. Fifteen subjects experienced ICANS simultaneously with CRS, and one subject experienced ICANS 4 days after CRS recovery.

[0085] Other neurotoxicities Twelve subjects (12.4%) experienced other CAR-T neurotoxicity not defined as investigator-assessed ICANS, either due to symptoms or time of onset (i.e., onset after the recovery period from CRS and / or ICANS). These events included a variety of symptoms of varying severity, including impaired consciousness, coordination and balance, movement and motor function disorders, mental impairment TEAEs, cranial neuropathy, and peripheral neuropathy.

[0086] Three subjects (3.1%) experienced maximum toxicity of grade 2. Eight subjects (8.2%) experienced grade 3 or 4 toxicity, and one subject (1.0%) experienced grade 5 toxicity. The events occurred with a median onset of 26.5 days (range, 11-108 days) and a median time to resolution of 70.0 days (range, 2-159 days) from cilta-cel infusion. At the time of clinical cutoff, five of the 12 patients (41.7%) had recovered, five (41.7%) had not recovered, one (8.3%) was recovering / resolving, and one (8.3%) had died from grade 5 neurotoxicity. The events are summarized in Table 2.

[0087] Other neurotoxicities characterized by adverse events occurring under exercise and neurocognitive treatments Symptoms associated with other CAR-T neurotoxicity events varied widely across the 12 subjects, although 5 of the 12 subjects (subjects L28US10002023, L28US10003011, L28US10017023, L28US10021005, and L28US10025003) experienced similar motor and neurocognitive treatment-emergent adverse event (TEAE) symptoms, including a range of motor (e.g., micrographia, tremor, etc.), cognitive (e.g., memory loss, attention problems, etc.), and personality changes (e.g., reduced facial expression, flat affect, etc.) TEAEs, which in some cases progressed to an inability to work or care for oneself.

[0088] One subject (1.0%) experienced a maximum toxicity of grade 2. Three subjects (3.1%) experienced grade 3 toxicity, and one subject (1.0%) experienced grade 5 toxicity. The events occurred with a median onset of 27 days from cilta-cel infusion (range: 14-108 days) (Table 3). Subjects were treated with steroids, systemic chemotherapy (cyclophosphamide), intrathecal chemotherapy (methotrexate, cytarabine), IL-1 receptor antagonists (anakinra), tyrosine kinase inhibitors (dasatinib), anti-IL-6 antibodies (siltuximab), and other agents (e.g., carbidopa / levodopa, levetiracetam, etc.), with limited or no improvement in symptoms.

[0089] At the time of clinical cutoff, one patient (8.3%) was recovering / resolving, one patient (8.3%) had died from grade 5 neurotoxicity, and three patients (25.0%) had not recovered / resolved (two of whom died from other causes [lung abscess and septic shock, both confirmed by autopsy]) (Table 3 ). [Table 3] TIFF2024518549000008.tif100165

[0090] The occurrence of motor, neurocognitive, and personality change TEAEs in those five subjects was potentially associated with a combination of two or more factors, such as high tumor burden, antecedent grade ≥2 CRS, antecedent ICANS, and high CAR-T cell proliferation and persistence. To minimize the risk of neurotoxicity in subjects in the ongoing cilta-cel clinical development program, the following monitoring and mitigation strategies were implemented: 1) intensive bridging therapy to reduce baseline tumor burden, 2) early aggressive treatment of CRS and ICANS, 3) written evaluation for early detection of neurotoxicity symptoms, and 4) extending the monitoring and reporting period for neurotoxicity up to 1 year after cilta-cel infusion. Subjects treated with cilta-cel were then enrolled in a long-term follow-up study (study 68284528MMY4002) and will be continuously monitored for up to 15 years.

[0091] Example 2 - Clinical Study 68284528MMY2003 Study 68284528MMY2003 is a phase 2, multi-cohort, open-label, multicenter study to determine whether treatment with cilta-cel results in minimal residual disease (MRD) negativity in adult subjects with multiple myeloma (MM). Cohorts of approximately 20 subjects each are planned, representing unique patient populations with MM and unmet medical need. Cohort A = Progressive disease refractory to lenalidomide with 1-3 prior lines of therapy (including proteasome inhibitors (PI) and immunomodulatory agents (IMiDs)). Cohort B = Early relapse defined as disease progression <12 months after autologous stem cell transplant (ASCT) or <12 months after initiation of first-line therapy for subjects not undergoing ASCT, with one prior line of therapy including a PI and an IMiD. Cohort C = relapsed or refractory disease in subjects previously treated with PI, IMiD, anti-CD38 monoclonal antibody (mAb), and BCMA-directed therapy (excluding immunotherapy). Cohort D = Cilta-cel + lenalidomide. MM patients who have not achieved complete response (CR) after a total of 4-8 cycles of initial therapy including induction therapy, high-dose chemotherapy, and ASCT with or without consolidation therapy. Cohort E = Daratumumab, bortezomib, lenalidomide, and dexamethasone (D-VRd) induction, Cilta-cel, followed by Daratumumab and lenalidomide (DR). High-risk newly diagnosed and untreated MM (hr-NDMM) subjects not planned for ASCT as initial therapy.

[0092] Eligible subjects will undergo apheresis, PBMC collection, and CAR-T generation as described in Study 8284528MMY2001 (Example 1). Based on the results of Study 68284528MMY2001, strategies to mitigate motor and neurocognitive TEAEs were implemented in Study 68284528MMY2003.

[0093] Other neurotoxic events not defined as ICANS (assessed by symptoms or time to onset) were reported in two subjects (11.1%). These other neurotoxic events included bradyslurred speech, facial palsy, gait disturbance, and pain, each reported at an incidence of 5.6%. The severity of these events was grade 1 or 2, and none were considered serious. The median time to onset was 20.0 days (range: 11-29 days). In one subject, the event resolved within 4 days, while in the other subject it was persistent.

[0094] After the clinical cutoff of Study 68284528MMY2003, another case of neurotoxicity was reported for one subject in Cohort B; adverse events included motor and neurocognitive TEAEs. The subject was a 44-year-old male. The male received a cilta-cel infusion on Study Day 1. The subject experienced CRS on Study Days 6 through 10, with a maximum severity of Grade 3. The subject was treated with tocilizumab, dexamethasone, dopamine, norepinephrine, and antibiotics. CRS resolved and the subject was discharged on Study Day 16. The subject did not experience ICANS.

[0095] On study day 55, the subject experienced encephalopathy with frontal lobe symptoms (grade 3) and bradykinesia (grade 3). Flat affect, akinetic mutism, lethargy, release signs (mild to moderate), symmetric rigidity, and parkinsonian gait and posture (no tremor) were observed. The subject was hospitalized. Evaluation included brain MRI with subtle abnormalities in the bilateral caudate nuclei (i.e., hyperintensity on fluid-attenuated inversion recovery (FLAIR) sequences), EEG with bilateral temporal lobe depression, PET / CT brain scan pending results, CSF evaluation for infection, and negative paraneoplastic antibody panel. The subject was treated with high-dose methylprednisolone, plasmapheresis, and IV immunoglobulin. The event is ongoing and is believed to be related to cilta-cel.

[0096] Example 3 - Clinical Study 68284528MMY3002 Study 68284528MMY3002 is a phase 3 randomized study comparing cilta-cel to pomalidomide, bortezomib, and dexamethasone or daratumumab, pomalidomide, and dexamethasone in patients with relapsed lenalidomide-refractory MM. Approximately 400 subjects are planned (200 subjects per treatment group). Based on the findings of Study 68284528MMY2001 (Example 1), a strategy to mitigate motor and neurocognitive TEAEs was implemented in Study 68284528MMY3002.

[0097] At the time of clinical cutoff, one of 16 subjects treated with cilta-cel experienced other neurotoxicity. The subject experienced left facial palsy (study day 18) followed by diplopia in the left eye (study day 36). The severity of both events was grade 2 and both resolved (study day 56 and study day 53, respectively). No subjects in the study reported motor or neurocognitive TEAEs.

[0098] Two other cases of neurotoxicity were reported after the clinical cutoff for Study 68284528MMY3002. One subject experienced left Bell's palsy (day 24 post-infusion). The severity of the event was grade 2 and is currently ongoing. The patient was treated with 60 mg of prednisone daily for 3 days, tapered over an additional 8 days. The patient also experienced word naming problems and long-term memory problems beginning on days 34 and 42 post-infusion, respectively. Both of those grade 1 events are currently ongoing. One subject experienced bilateral facial paralysis (day 25 post-infusion). The severity of the events was grade 2 and is currently ongoing. An MRI of the brain showed no acute intracranial abnormalities. It demonstrated that CSF cytology showed mature lymphocytes and monocytes. CSF flow cytometry showed >95% CD5 positive presumptive T cells and no evidence of malignancy. CSF meningitis / encephalitis panel was negative. CSF cultures, cryptococcal antigen, and AFB smears were negative. Patients were treated with 60 mg prednisone daily for 2 days, tapered over an additional 8 days. No motor or neurocognitive TEAEs have been reported in the study to date.

[0099] Example 4 - Evaluation of factors that lead to CAR-T cell neurotoxicity Neurotoxicity after CRS resolution is reported in the literature for CD19-targeted chimeric antigen receptor T cell (CAR T) therapy. Additionally, a grade 3 Parkinson's disease case is referenced in the USPI for ide-cel (BCMA-targeted CAR-T therapy). Study 68284528MMY2001 noted that there were other cases of neurotoxicity characterized by motor and neurocognitive TEAEs that occurred after a period of recovery from cytokine release syndrome (CRS) or immune effector cell-associated neurotoxicity syndrome (ICANS), which appeared to share common features in terms of clinical symptoms and CAR-T correlation data. Similar to what has been observed with approved CD19 and B-cell maturation antigen (BCMA)-directed CAR-T therapy, the exact mechanism of said neurotoxicity is unclear at this time. Thus far, brain magnetic resonance imaging (MRI) and virology workup have yielded essentially negative results in those cases. Additionally, comprehensive evaluation with paraneoplastic and autoimmune antibody panels has been negative when performed.

[0100] Objective: The primary objective of this analysis was to investigate clinical data related to the occurrence of motor and neurocognitive TEAEs in patients with multiple myeloma receiving cilta-cel infusions.

[0101] Methods: Analyses were performed to examine clinical data, including demographics, baseline disease characteristics, baseline laboratory values, exposure to cilta-cel, CRS or ICANS adverse events (including grade), lymphocyte counts, neutrophil counts, and platelet counts, that correlated with the occurrence of motor and neurocognitive TEAEs.

[0102] Subjects and Database. The analysis included 97 subjects who received a cilta-cel infusion from the main cohort of study 68284528MMY2001. Five subjects were considered to have motor and neurocognitive TEAEs.

[0103] Elapsed days and baseline. Study day 1 refers to the start of the first dose of cilta-cel. Baseline values ​​are defined as the closest non-missing value prior to the first dose of cilta-cel (including time, if any), except for parameters related to disease-related efficacy assessments where baseline values ​​are defined as the closest non-missing value at the start of conditioning therapy and prior to cilta-cel infusion. Depending on the time and event schedule of the protocol, baseline for non-efficacy variables may be at screening, prior to conditioning, or prior to cilta-cel infusion.

[0104] Motor and Neurocognitive TEAEs. Subjects with other neurotoxicity characterized by motor and motor dysfunction TEAEs and cognitive dysfunction TEAEs were identified by time of onset of CAR-T-related neurotoxicity (i.e., after resolution of CRS and / or ICANS) and reporting of MedDRA Preferred Terms (Table 4) from at least two of the following categories: [Table 4]

[0105] Statistical methods. Descriptive statistics and frequency distributions with number and percentage of subjects in each category were included, where appropriate. Differences in clinical variables between subjects with and without motor and neurocognitive TEAEs were assessed using Wilcoxon rank sum tests for continuous variables and Fisher's exact tests for categorical variables. Logistic regression models with motor and neurocognitive TEAEs as dependent variables and each clinical variable as an independent variable were performed to evaluate each clinical variable.

[0106] Given the exploratory nature of the analyses with multiplexity and small number of events (i.e., motor and neurocognitive TEAEs), results are hypothesis generating and should be interpreted with caution.

[0107] Data processing: Cytokines: FRONTAGE central laboratory data was used. For results "below the lower limit of quantification (LLOQ)", the LLOQ was used in the analysis.

[0108] result Variables examined in exploring common characteristics among subjects who presented with motor or neurocognitive TEAEs included: Baseline variables: Demographics: age, gender, race, ethnicity, Baseline tumor burden: bone marrow plasmacytosis, LDH, soft tissue plasmacytoma (extramedullary (yes / no, number, SPD)), serum M spike / serum free light chain, Baseline disease characteristics: disease tempo (percentage change in tumor burden from screening to baseline), type of multiple myeloma, type of measurable disease, treatment history, time since MM diagnosis, ECOG PS, tumor BCMA expression, time since last apheresis, Use of bridge therapy, bridge use of daratumumab / lenalidomide, Viral infection before apheresis, prior radiation therapy to the brain, Baseline laboratory values ​​(fibrinogen, C-reactive protein, ferritin, platelet count, beta-2 microglobulin, IL-6, IL-10, INF-gamma, IL-2 receptor alpha, estimated glomerular filtration rate), and Exposure data: dose, duration of infusion. Post-baseline variables: CRS, maximum CRS grade (<2 vs. ≥2), ICANS, maximum ICANS grade (<2 vs. ≥2), concomitant CRS and ICANS therapy (i.e., steroids, tocilizumab, anakinra), · Absolute lymphocyte count (ALC), absolute neutrophil count (ANC), and platelet count for the first 30 days from local complete blood count (CBC). Maximum ferritin levels during the first 2 / 3 weeks, ferritin at day 28, and -CAR-T cell proliferation, CAR-T cell survival. Other variables investigated included the pattern of BCMA expression in normal (non-diseased) human brain sections, as well as the dose and production regimen of cilta-cel.

[0109] The analysis of those variables is described in detail below. Contingency tables and box plots are displayed to evaluate the correlation between clinical variables and the occurrence of motor and neurocognitive TEAEs. In summary, the following variables are associated with the occurrence of motor and neurocognitive TEAEs: high tumor burden at baseline, baseline IL-6, maximum grade of CRS, ICANS, lymphocytes on days 14, 21, and 28, and proliferation / persistence of high-density cells. Forest plots of estimated odds ratios (OR) and confidence intervals (CI) of those variables associated with the occurrence of motor and neurocognitive TEAEs are provided (Figure 11).

[0110] demographics Possible relationships between subject demographics and the occurrence of observed motor and neurocognitive TEAEs were explored. Of the five subjects in study 68284528MMY2001 who met the criteria defining motor and neurocognitive TEAEs including personality changes, all five were Caucasian males (Table 5 and Figure 12). [Table 5]

[0111] Baseline disease characteristics-disease burden A possible causal relationship between the disease burden level and the occurrence of observed neurotoxicity was investigated. As there are no publicly available criteria to define high and low disease burden, in an exploratory analysis, disease burden categories were defined as follows: subjects were classified as having high tumor burden if any of the following parameters were met: (i) plasmacytosis in bone marrow ≥80%, (ii) serum M spike ≥5 g / dL, (iii) serum free light chain ≥5000 mg / L. Subjects were classified as having low tumor burden if all of the following parameters (applicable to the subject) were met: (i) plasmacytosis <50%, (ii) serum M spike <3 g / dL, (iii) serum free light chain <3000 mg / L. Subjects not meeting either criterion were considered to have moderate disease burden.

[0112] Applying the aforementioned criteria, higher baseline disease burden was associated with higher CAR-T proliferation and neurotoxicity (Table 6). Lower baseline disease burden was associated with lower incidence of neurotoxicity. [Table 6] TIFF2024518549000012.tif83165

[0113] Upfront treatment of multiple myeloma and bridge therapy for the treatment of multiple myeloma No association was identified between prior treatment and bridging therapy for MM and the occurrence of observed performance / motor and cognitive dysfunction TEAEs (Table 7). All five subjects received bridging therapy between apheresis and the start of conditioning therapy. After bridging therapy, four of five subjects (80%) experienced an increase in tumor burden (three had increasing paraproteins and one had plasmacytoma enlargement despite improvement in paraproteins). [Table 7]

[0114] Proliferation of chimeric antigen receptor T cells and persistence of chimeric antigen receptor T cells In study 68284528MMY2001, CAR-T proliferation and persistence were associated with motor and neurocognitive TEAEs (Table 8), with subjects L28US10003011, L28US10025003, L28US10017023, and L28US10002023 being among those with the highest levels of CAR-T in peripheral blood (Figure 2) (subject L28US10021005 was not among those who showed high cell proliferation). Additionally, subjects with motor or neurocognitive TEAEs (subjects L28US10002023, L28US10003011, L28US10017023, L28US10021005, and L28US10025003 (Figure 12)) notably had substantially longer persistence of CAR-T in peripheral blood than subjects who did not develop other symptoms of neurotoxicity (Figure 2). Cerebrospinal fluid (CSF) examination of two subjects with motor or neurocognitive TEAEs (for whom CSF was available) demonstrated that CAR-T cells comprised a major portion of T cells that were primarily of the effector memory phenotype.

[0115] These data suggest that high levels of CAR-T cell proliferation and CAR-T cell persistence are associated with an increased risk of developing motor and neurocognitive TEAEs after resolution of CRS, which may lead to the development of these other neurotoxicities. [Table 8]

[0116] Preceding or concurrent occurrence of cytokine release syndrome and immune effector cell-associated neurotoxicity syndrome The correlation between the presence of preceding or concurrent CRS and ICANS and the occurrence of motor and neurocognitive TEAEs was investigated (Table 9). All subjects with motor or neurocognitive TEAEs had CRS of grade 2 or higher, whereas subjects with CRS of grade 1 or lower reported no other neurotoxicity of this type. Subjects with ICANS (any grade) were more likely to develop motor and neurocognitive TEAEs than those without ICANS (80.0% and 20.0%, respectively). Thus, higher grade CRS (grade 2 or higher) and any grade ICANS appear to be associated with motor and neurocognitive TEAEs. [Table 9]

[0117] Clinical Laboratory Evaluation Early in the time course of cilta-cel infusion, laboratory values ​​associated with risk and occurrence of motor and neurocognitive TEAEs include high baseline (i.e., pre-cilta-cel infusion) IL-6 (Figure 3) and high lymphocyte counts on study days 14, 21, and 28 after cilta-cel infusion (Figure 4). Data for additional baseline variables (IL-10, INF-γ, and IL2Ra) are provided in Figures 13-15.

[0118] Biomarker analysis – peak cytokine levels Peak levels of several proinflammatory cytokines in peripheral blood, including IL-6 (Figure 5) and INF-γ (Figure 6), were significantly increased (C max ) was elevated in subjects with motor and neurocognitive TEAEs compared with subjects with other neurotoxicity or no neurotoxic TEAEs. Thus, both biomarkers appear to be associated with motor and neurocognitive TEAEs.

[0119] Biomarker analysis-T cell memory phenotype at the time of apheresis Peripheral T cell memory phenotype was assessed at the time of apheresis using standard flow cytometry methods. Figure 7 shows the frequency distribution of CD4 and CD8 T cells of naive / stem cell memory (Tn / Tscm), central memory (Tcm), effector memory (Tem), and effector memory re-expressed CD45RA (Temra) phenotypes. The frequency of T cell memory subsets in subjects with motor and neurocognitive TEAEs overlapped with that of other subjects with other neurotoxicities, ICANS, etc.

[0120] Neuropathological findings at autopsy In clinical study 68284528MMY2001, three of five subjects experienced other neurotoxicity characterized by motor and neurocognitive TEAEs. Causes of death were neurotoxicity in one subject, lung abscess in one subject (autopsy performed), and septic shock in the last subject (autopsy performed). Neuropathology reports for two subjects who underwent autopsy showed focal gliosis and T cell infiltration (CD8 + >CD4 + ) was shown. These T lymphocytes were + It is unclear whether these abnormalities are cellular. No abnormalities were reported in other brain regions (e.g., cerebellum, substantia nigra) that may be associated with motor TEAEs in either subject. Preservation of pigmentation in the substantia nigra was reported in both subjects.

[0121] Immunohistochemical evaluation of B cell maturation antigen expression To determine whether motor and neurocognitive TEAEs are related to target expression, the expression of B cell migration antigen (BCMA) in normal (non-diseased) human brain was evaluated. The study concluded that BCMA expression was not detectable in normal (non-diseased) adult human brain.

[0122] An immunohistochemistry (IHC) assay using a commercially available mAb (clone E6D7B) from Cell Signaling Technology, Inc. was developed for application to formalin-fixed, paraffin-embedded (FFPE) brain samples. Two additional IHC assays using a second commercially available mAb (clone D6) from Santa Cruz Biotechnology, Inc. were developed at a third-party molecular pathology laboratory, also for application to FFPE samples. Both antibody clones (E6D7B and D6) sensitively and specifically detected BCMA in FFPE tissues and cell line controls (EDMS-RIM-367752, EDMS-RIM-367755, EDMS-RIM-387220).

[0123] Immunohistochemistry was performed internally on a total of 107 commercially available FFPE human brain samples covering all brain regions across 63 individual donors (EDMS-RIM-387220). All samples included in the study were quality checked to confirm location and suitability for IHC analysis. The internal assay used the E6D7B clone. Sporadic immunoreactivity was detected in the striatum and brainstem, and to a lesser extent in the thalamus, midbrain, hippocampus, and cerebellum. Immunoreactivity appeared as fibrils and aggregates within neuronal cell bodies in gray matter, or as short, thin threads along glial processes. The immunoreactivity was not reproduced when IHC was repeated using the D6 clone in a third-party laboratory.

[0124] Given the conflicting results seen with the two mAb clones, further investigation was undertaken, and based on the results outlined below, it was determined that the immunoreactivity observed with the E6D7B clone represents nonspecific cross-reactivity and does not reflect true BCMA expression.

[0125] In situ hybridization using a BCMA-specific ribonucleic acid (RNA) probe from RNAscope (ACD Bio) was performed on 25 randomly selected brain samples pre-stained with the E6D7B clone. B cell maturation antigen RNA was not detected in areas / neurons corresponding to E6D7B-mediated immunoreactivity (EDMS-RIM-387220).

[0126] The subcellular localization of neuronal immunoreactivity seen with E6D7B clones is not consistent with the current understanding of BCMA expression biology. In plasma and MM cells, where BCMA is expressed, BCMA protein is detected on the cell membrane and within the Golgi apparatus (Gras 1995 et al., “BCMAp: An Integral Membrane Protein in the Golgi Apparatus of Human Mature B Lymphocytes,” Internat. Immunol. 7:1093-1106 (1995), the entire contents of which are incorporated herein by reference). No membrane immunoreactivity was detected in brain samples. Furthermore, confocal microscopy with E6D7B clones revealed that immunoreactivity within neuronal cell bodies did not colocalize with a Golgi-specific marker (EDMS-RIM-387220).

[0127] The immunoreactivity pattern seen in the E6D7B clone does not correlate with previously reported BCMA expression data. A literature review and examination of publicly available BCMA expression data was performed. Low levels of BCMA RNA are detectable in the striatum during fetal development, but levels decline gradually throughout adolescence. After the age of 30 years, BCMA-RNA expression is negligible (Brainspan.org, "Atlas of the Developing Human Brain", available at https: / / www.brainspan.org, accessed March 17, 2021; GTExPortal. Broad Institute of MIT and Harvard. Available at https: / / www.gtexportal / home, accessed March 17, 2021, the contents of which are incorporated herein by reference in their entirety).No BCMA RNA or protein was detected in the cerebrum, cerebellum, or brainstem of normal adults at any age (Brainspan.org, “Atlas of the Developing Human Brain,” available at https: / / www.brainspan.org, accessed March 17, 2021; Bu et al., “Pre-Clinical Validation of B Cell Maturation Antigen (BCMA) as a Target for T Cell Immunotherapy of Multiple Myeloma,” Oncotarget 9(40):25764-25780 (2018); GTExPortal. Broad Institute of MIT and Harvard. Available at https: / / www.gtexportal / home. Accessed March 17, 2021; Carpenter et al., “B-Cell Maturation Antigen is a Promising Target for Adoptive T-Cell Therapy of Multiple Myeloma,” Clin. Cancer 2019, 10:131–135 (2019)). Res. 19(8):2048-2460 (2013), and Krumbholz et al., “BAFF is Produced by Astrocytes and Up-Regulated in Multiple Sclerosis Lesions and Primary Central Nervous System Lymphoma,” J. Exp. Med. 201(2):195-200 (2005), the contents of which are incorporated herein by reference in their entireties.) In the current IHC study, immunoreactivity for the E6D7B clone was found in multiple brain regions and in the striatum in donors ranging in age from 39 to 85 years.

[0128] The E6D7B clone is used to perform immunohistochemistry on FFPE brain samples from four cynomolgus monkeys. Although no immunoreactivity is detected in the brain, the antibody functions as an IHC reagent on a cynomolgus monkey FFPE tissue control (EDMS-RIM-387220).

[0129] Immunohistochemistry stained tissues were examined by an outside neuropathologist who ensured that appropriate control samples were included in the IHC assays and confirmed that both mAb clones performed similarly against tissue and cell line controls, but only the E6D7B clone showed immunoreactivity in the brain. The neuropathologist suggests that the immunoreactivity seen with the E6D7B clone is likely nonspecific (EDMS-RIM-387220).

[0130] Pharmacokinetics Exposure-response relationships for safety endpoints in Study 68284528MMY2001 are shown in Figures 8A-8B. For other neurotoxicities (including motor and neurocognitive TEAEs), no clear trends were observed with respect to the total dose of cilta-cel infused (Figure 8A). In clinical study 68284528MMY2001, one target dose level of cilta-cel (0.75 [range: 0.5-1.0] x 10 6 This is expected, since only 100 CAR-positive viable T cells / kg were investigated.

[0131] Median systemic CAR transgene levels (C) in subjects with other neurotoxicity (including motor and neurocognitive TEAEs) or motor and neurocognitive TEAEs max and the area under the time curve from the first dose to the 28th day (AUC 0-28d ) were generally higher than subjects with other neurotoxicity (including motor and neurocognitive TEAEs) ( FIG. 8B ) or without motor and neurocognitive TEAEs ( FIG. 9 ), respectively, although there is overlap in the range of systemic exposure of the CAR transgene between subjects with and without different grades of these TEAEs.

[0132] C max and AUC 0-28d As well as the time to maximum proliferation (T max) ranges overlapped (Figure 8B). Five subjects with motor and neurocognitive TEAEs appeared to have a delayed peak in expansion, although the small number of subjects precluded definitive conclusions (Figure 9).

[0133] Chemistry, manufacturing, and controls evaluation Five subjects in Study 68284528MMY2001 experienced motor and neurocognitive TEAEs (Subjects L28US10003011, L28US10025003, L28US1002023, L28US10017023, L2810021005). A thorough review of batch documentation was performed for all five subject batches. Key drug quality attributes met the release specifications outlined in Table 10 below, with the exception of Batch 19HC0096, which was shipped per exceptional shipping procedures (Reference Nos. IND18080, S / Ns 0080 and 0085). [Table 10]

[0134] Manufacturing of Cilta-cel drug product in clinical study 68284528MMY2001 is performed at Cincinnati Children's Hospital Medical Center (CCHMC), Janssen, Spring House, Pennsylvania and Janssen, Raritan, New Jersey. All five study batches (Table 10) were manufactured at Raritan facilities using vector batches LV-LICAR2SINV8008 (19GC0067 and 19HC0096) and LICAR2SINV8010 (19KC0177, 19JC0127, and 19KC0165). A total of 97 batches were manufactured in study 68284528MMY2001, of which 68 batches were manufactured at Raritan facilities.

[0135] The five target batches listed in Table 10 were investigated for events or deviations that may have occurred during manufacturing. With the exception of batch 19KC0165, no significant events or deviations were reported during the manufacturing of those batches. In-process control testing for all five target batches met the acceptance criteria, and with the exception of the dosage specifications for batch 19HC0096, all release results also met the acceptance criteria. The release data are distributed across the range of manufacturing experience. Cell clumps were observed in two batches of the formulation and are recorded per the visual inspection standard operating procedure. Cell clumps are not unexpected in the CAR-T process.

[0136] During the processing of batch 19KC0165 on day 10, a malfunction in the reheat system from a water supply line accidentally disconnected by a contractor working in the facility caused the room temperature to drop below the alarm limit of 55°F. The reheat system is responsible for adding heat to the pre-chilled air in the facility and is accomplished by heating hot water heating elements in the air terminal. During the 10-day processing, cells are collected, formulated into cryopreservation media, and stored at 2-8°C (35.6-46.4°F). During this step, the temperature fluctuations observed in the room are within the normal processing range of 2°C from ambient temperature. Additionally, a second deviation occurred during the controlled rate freezing (CRF) step of the batch, where a sensor failure error was observed when loading samples into the chamber. The CRF cycle was stopped, restarted, and continued as scheduled. Both deviations were deemed to have no impact on the resulting drug product.

[0137] Process characteristic data (%CAR) of T cell expansion after transduction of five control batches of cilta-cel (Table 10) + CD4 + , %CAR + CD8 + , %CAR + / CD45 + , %CAR + / CCR7 + ) was compared to data sets from all three pharmaceutical manufacturing sites and assessed for any trends. Batch 19GC0067 showed T cell expansion after transduction and CD4 + / CD8 +The ratio indicates that the ratio reached the high end of the manufacturing range, however it was observed in other batches manufactured at CCHMC and Raritan, and there is no correlation with motor or neurocognitive events. The other four subject batches in the study showed no observable trends, and the data are spread across the entire manufacturing experience range for those attributes. %CAR + The CD45, CCR7 attributes show no trend in any of the five batches studied. The vector copy number in all five batches is below 0.5 copies per cell.

[0138] There were no trends identified in the in-analysis or shipping data for the five batches of cilta-cel drug product used to treat the five subjects with subsequent motor and neurocognitive TEAEs, as well as the occurrence of these adverse events (Table 10). Process characteristic attributes of all batches in production are continuously monitored in ongoing and planned clinical studies.

[0139] overview As more safety data is generated on approved and new CAR-T therapies, the definition of CAR-T-related neurotoxicity will continue to evolve beyond ICANS to include other neurotoxicities. This document focused on other neurotoxicities categorized by cluster of motor and neurocognitive adverse events in ongoing studies of cilta-cel. To date, factors potentially associated with motor and neurocognitive adverse events include: High tumor burden at baseline (high tumor burden is defined as - 80% or more of bone marrow plasmacytosis or - Serum M spike of 5 g / dL or more, or - Serum free light chains ≥ 5000mg / L. High baseline IL-6 levels, High lymphocyte counts on days 14, 21, and 28 of the study (after Cilta-cel infusion); High peak levels of IL-6 and INF-γ in peripheral blood (C max ), -Preceding CRS (grade ≥2) or ICANS (any grade), - CAR-T cell proliferation with high levels of cell proliferation (CAR-T cell C max defined as >1,000 cells / μL), and · CAR-T cell persistence with high levels of cell persistence (defined as >300 CAR-T cells / μL on study day 56).

[0140] Figure 11 summarizes the odds ratios and confidence intervals for each of these possible factors.

[0141] Example 5 - Mitigation strategies to reduce CAR-T-associated neurotoxicity After receiving the first cases of motor and neurocognitive TEAEs, an ad hoc Safety Management Team was convened, a notification was sent to the United States (US) Food and Drug Administration (FDA), Dear Investigator Letters (DIL) were distributed to ongoing study sites (Studies 68274528MMY2001, 68284528MMY2002, and 68274528MMY2003), a neurotoxicity working group was established, and regular investigator conference calls were conducted (Study 68274528MMY2001). In accordance with the DIL, investigators were asked to alert the medical monitor for any neurotoxicity (including ICANS) following cilta-cel infusion. Investigators were advised to aggressively treat cases of ICANS with steroids according to the ASTCT consensus criteria (included in the protocol).

[0142] Given the cumulative cases of motor and neurocognitive TEAEs observed in clinical study 68274528MMY2001, their rapidly progressive severity (including inability to work or care for oneself after the initial insidious subtle symptom onset), and the first fatal outcome, immediate changes in the conduct of the ongoing study were implemented for safety reasons, followed by amendments to the protocol and informed consent forms. The clinical summary and decision were communicated to the US FDA, and DILs were sent to all ongoing sites (Studies 68274528MMY2001, 68284528MMY2002, 68274528MMY2003, and 68284528MMY3002). Based on the new data, a program-wide protocol amendment was issued with a monitoring and mitigation strategy for motor and neurocognitive TEAEs.

[0143] The status of studies conducted in the United States (Clinical Studies 68284528MMY2001, 68284528MMY2003, and 68284528MMY3002) is summarized in Table 12. [Table 11]

[0144] The following mitigation procedures were identified and implemented: 1. Dear Investigator Letters will be sent to the principal investigators of Studies 68284528MMY2001, 68284528MMY2003, and 68284528MMY3002. 2. The 68284528MMY2001, 68284528MMY2003, and 68284528MMY3002 protocols were modified to include descriptions of key features of patterns of motor and neurocognitive TEAEs and the likelihood of severe disability outcomes. 3. Updated informed consent to include recent motor and neurocognitive TEAEs and a provision to send a CSF sample to the sponsor if a lumbar puncture was performed. Modified informed consent forms for Study 68284528MMY2003 and Study 68284528MMY3002 to include provisions for autopsy samples. 4. For subjects with a history of relevant neurological disease (stroke, encephalitis, etc.), we have modified both the 68284528MMY2003 and 68284528MMY3002 protocols to recommend a baseline MRI and EEG of the brain. 5. 68284528MMY2003 protocol amended to allow for bridge therapy (based on investigator choice) with the goal of reducing tumor burden prior to CAR-T infusion. Previously, only drugs to which the subject had been previously exposed were allowed as part of bridge therapy. 68284528MMY2001 protocol amended to allow for bridge therapy in case of retreatment. 68284528MMY3002 protocol amended to allow additional cycles of bridge therapy based on the subject's clinical status, also with the goal of reducing tumor burden prior to CAR-T infusion. 6. Subjects with high baseline disease burden, especially those with disease progression despite bridging therapy, are potentially at high risk of developing severe neurotoxicity characterized by TEAEs of motor, neurocognitive, and personality changes and therefore a risk-benefit discussion should be conducted prior to CAR-T infusion. 7. Amended the 68284528MMY2001, 68284528MMY2003, and 68284528MMY3002 protocols to include a recommendation for extended use of prophylactic antibiotics (up to 6 months or longer per institutional guidelines) or to be consistent with consensus guidelines after ASCT. 8. The 68284528MMY2001, 68284528MMY2003, and 68284528MMY3002 protocols were amended to incorporate a recommendation for early and aggressive steroid treatment for ICANS of any grade to evaluate whether this intervention, at the investigator's discretion, could reduce the risk of motor and neurocognitive TEAEs occurring after resolution of CRS. 9. Monitoring and reporting of neurological events (new events or worsening of existing events) will be extended to one year post-infusion (instead of 100 days) with instructions to notify the medical monitor if motor or neurocognitive symptoms are observed. 10. The 68284528MMY2001, 68284528MMY2003, and 68284528MMY3002 protocols have been modified to incorporate routine writing assessments that will be performed on subjects periodically, pre- and post-infusion, throughout the study period to investigate writing changes as a potential early indicator of other neurotoxicity. This new writing tool was developed by Janssen and consists of a writing log and scoring criteria (Grade 1 and Grade 2) for three preferred terms (agraphia, micrographia, and dysgraphia). Sites are instructed to report them as TEAEs in the electronic database and to notify the medical monitor immediately if changes are observed. If a subject develops a severe adverse event (SAE) of neurotoxicity after CRS resolution, a copy of the log should be added as part of the source documentation. Writing assessments will be in addition to the current Immune Effector Cell-Associated Encephalopathy Assessment Tool (ICE). 11. Added recommended workup for subjects who develop other neurotoxic events, including guidance to rule out specific viral infections (CSF analysis to rule out human herpesvirus [HHV]-6, HHV-7, John Cunningham virus [JCV], herpes simplex virus [HSV]-1, 2, and serology for HHV-6, HHV-7, and JCV), CSF flow cytometry to rule out leptomeningeal disease and paraneoplastic etiologies, serum thiamine levels (consider supplementation), and imaging studies (i.e., PET scan or MRI perfusion, EEG). 12. The 68284528MMY2001, 68284528MMY2003, and 68284528MMY3002 protocols have been amended, in consultation with the sponsor, to include consideration of treatments aimed at mitigating or eliminating CAR-T when motor and neurocognitive TEAEs are unresponsive to other interventions. 13. In addition to the mitigation procedures above, a Data Monitoring Committee (DMC) will be established for study 68284528MMY2003 to review all subjects receiving treatment regardless of cohort. 14. In cohort D of study 68284528MMY2003, the following additional mitigation steps were implemented: a. Dosing of the first 5 subjects will be spaced at least 4 weeks apart b. The first 5 subjects will start without lenalidomide c. The DMC will decide whether it is safe to initiate lenalidomide after reviewing the data from the first 5 subjects. d. The first 5 subjects (subjects 6-10) to start lenalidomide should be spaced at least 4 weeks apart.

[0145] Additionally, multiple assessments were performed to evaluate possible predictors and underlying pathology of motor and neurocognitive TEAEs (see Example 4), the results of which are summarized herein.

[0146] Planned monitoring In addition to a long-term follow-up study (study 68284528MMY4002) to continue monitoring subjects treated with cilta-cel for up to 15 years, a post-authorization safety observational study using a registry is planned to further characterize identified risks and further evaluate potential risks and unknowns, with a particular focus on long-term safety.

[0147] Guidance for doctors Risk monitoring and mitigation strategies outlined in the Investigator Brochure will be followed (see, e.g., Table 13 below). Additionally, in the ongoing cilta-cel clinical development program, monitoring and mitigation strategies such as intensive bridging therapy to reduce baseline tumor burden, early aggressive treatment of CRS and ICANS, written assessment for early detection of neurotoxicity symptoms, and extended monitoring and reporting periods for neurotoxicity beyond 100 days after cilta-cel infusion have been implemented to minimize the risk of motor and neurocognitive TEAEs in subjects. With the exception of the five subjects in Clinical Study 68284528MMY2001 and one subject in Clinical Study 68284528MMY2003 mentioned above, no additional cases of other neurotoxicity characterized by motor and neurocognitive TEAEs have been reported to date in any study in the cilta-cel development program. [Table 12] TIFF2024518549000019.tif135165

[0148] If any neurological or psychiatric symptoms are observed (see below), the medical monitor should be contacted and the subject should be immediately referred to a neurologist for full evaluation. Following cilta-cel infusion, subjects should be monitored for neurotoxicity for the duration of the study. Particular attention should be paid to any of the following situations: Motor and Neurocognitive TEAS (often with subtle onset) Motor dysfunction (e.g. micrographia or changes in writing, tremor, bradykinesia, rigidity, hindlimb twisting, balance and coordination problems, writing problems, difficulty with activities of daily living such as dressing and eating), Cognitive impairment (e.g. memory loss or forgetfulness, attention problems, mental slowness or amnesia, difficulty speaking or slurred speech, difficulty reading or understanding words), Personality changes (e.g., decreased facial expression, flat affect, decreased ability to express emotions, decreased social interaction, disinterest in activities). Early detection, workup, and intervention may be important to prevent worsening neurotoxicity. Below is a list of potential diagnostic procedures to consider in subjects with new neurological symptoms: · Brain PET / Computed Tomography (CT) and / or brain MRI with perfusion and EEG. Lumbar puncture to rule out infectious diseases (especially JCV, varicella-zoster virus (HZV), HSV-1 / 2, HHV-6, HHV-7, Epstein-Barr virus (EBV), and cytomegalovirus (CMV)). · Serological testing for HHV-6 and HHV-7 by polymerase chain reaction for viremia. CSF flow cytometry and cytology should be considered to rule out leptomeningeal disease. CSF analysis should be considered to exclude paraneoplastic syndromes. · Thiamine levels (consider empirical thiamine replacement while waiting for results). For subjects who develop neurotoxicity unresponsive to other interventions, therapies aimed at reducing or eliminating CAR-T, including chemotherapy, may be considered in consultation with the sponsor.

[0149] conclusion The examples presented herein demonstrate the use of the following strategies used to mitigate and further understand the pathology of motor and neurocognitive adverse events: (i) baseline magnetic resonance imaging (MRI) and baseline electroencephalography (EEG) of the brain in subjects with a history of relevant neurological disease (e.g., stroke, encephalitis), (ii) bridging therapy to reduce tumor burden prior to administration of immune effector cell therapy (e.g., cilta-cel infusion). 1 (iii) antibiotic prophylaxis for up to 6 months or more (per institutional guidelines or per post-autologous stem cell transplant [ASCT] consensus guidelines); (iv) early and aggressive steroid treatment for any grade of ICANS; (v) extension of the monitoring and reporting period for neurological adverse events to more than 100 days after administration of immune effector cell therapy; (vi) writing assessment at baseline (e.g., before cilta-cel infusion) and during treatment to investigate writing changes as a potential early indicator of motor and neurocognitive adverse events; (vii) virological workup to rule out infection, cerebrospinal fluid (CSF) flow cytometry to rule out leptomeningeal disease and paraneoplastic etiologies, serum thiamine levels (considering supplementation), and brain imaging (e.g., positron emission tomography (PET) scan or MRI perfusion, EEG); and (viii) treatment to reduce or eliminate CAR-T cells in the event of motor and neurocognitive adverse events unresponsive to other interventions. 1 For subjects with high baseline disease burden despite bridging therapy, risk / benefit discussions are warranted as they are at higher risk of developing severe motor, neurocognitive, and personality-changing adverse events.

[0150] The above mitigation strategies have been initiated for all ongoing cilta-cel studies (Urgent Safety Measure (USM)). Since implementation of the mitigation strategies, the incidence of motor and neurocognitive adverse events has decreased from 5% to 1%. Motor and neurocognitive adverse events are continually monitored, and exploratory evaluations of possible predictors and contributors to these adverse events are continually performed.

[0151] As more safety data are generated for approved and new CAR-T therapies, the definition of CAR-T-associated neurotoxicity will continue to evolve beyond ICANS to include other neurotoxicities as motor and neurocognitive adverse events. Indeed, parkinsonism has been reported with ide-cel, another BCMA-targeted CAR-T therapy, in patients with MM. Motor and neurocognitive adverse events will be monitored on an ongoing basis, and exploratory evaluations of possible predictors and underlying pathologies for these adverse events will be performed on an ongoing basis.

[0152] While preferred embodiments have been shown and described in detail herein, it will be apparent to those skilled in the art that various modifications, additions, substitutions, and the like may be made therein without departing from the spirit of the invention, and therefore are deemed to be within the scope of the invention as defined in the claims.

Claims

1. 1. A pharmaceutical composition comprising chimeric antigen receptor (CAR)-T cells for use in a method for treating multiple myeloma in a subject and simultaneously reducing non-immune effector cell-associated neurotoxicity syndrome (non-ICANS) neurotoxicity associated with CAR-T cell therapy, comprising: the non-ICANS neurotoxicity occurs after resolution of ICANS and / or cytokine release syndrome (CRS); The method comprises: (i) administering said pharmaceutical composition to said subject having multiple myeloma and a tumor burden characterized by bone marrow plasmacytosis less than 80%, serum M protein levels less than 5 g / dL, and serum free light chain levels less than 5000 mg / L; (ii) administering the pharmaceutical composition to the subject; and a. administering an anti-inflammatory agent to the subject receiving the pharmaceutical composition and having symptoms of CAR-T cell therapy-associated CRS or ICANS, wherein the CRS is Grade 2 or greater; b. administering a chemotherapeutic or anti-inflammatory agent to said subject receiving said pharmaceutical composition and having a CAR-T cell maximum plasma concentration (Cmax) of greater than 1,000 cells / μL and / or a persisting CAR-T cell concentration in peripheral blood of greater than 300 cells / μL following administration of said CAR-T cells; c. administering to said subject receiving said pharmaceutical composition and having a peak peripheral blood IL-6 level above the upper limit of normal following administration of CAR-T cells an IL-6 inhibitor; d. administering an IFNγ inhibitor to said subject receiving the pharmaceutical composition and having a peak peripheral blood IFNγ level above the upper limit of normal following administration of CAR-T cells; and / or e. administering an anti-inflammatory agent to the subject receiving the pharmaceutical composition and having a lymphocyte count above the upper limit of normal about 2 weeks, about 3 weeks, about 4 weeks, about 5 weeks, about 6 weeks, about 7 weeks, about 8 weeks, about 9 weeks, or about 10 weeks after administration of the CAR-T cells; and / or (iii) administering said pharmaceutical composition to said subject having multiple myeloma and having an IL-6 serum level within the normal reference range of 0-2 pg / mL. The pharmaceutical composition comprising:

2. the method further comprises administering to the subject a bridge therapy to achieve a tumor burden characterized by bone marrow plasmacytosis less than 80%, serum M protein levels less than 5 g / dL, and serum free light chain levels less than 5000 mg / L prior to administering the pharmaceutical composition; Optionally, the bridge therapy comprises: (a) a chemotherapeutic agent, such as an alkylating agent or a topoisomerase inhibitor; (b) an immunomodulatory agent such as a CD38 inhibitor, preferably, the CD38 inhibitor is daratumumab; (c) an immunomodulatory agent, such as lenalidomide, pomalidomide, thalidomide, or a combination thereof; (d) a proteasome inhibitor, such as bortezomib, carfilzomib, ixazomib, or a combination thereof; or (e) Any combination of (a) to (d).

2. The pharmaceutical composition of claim 1 (i), comprising:

3. the anti-inflammatory agent comprises an IL-6 inhibitor; Preferably, the IL-6 inhibitor comprises tocilizumab, or steroids such as dexamethasone and methylprednisolone. A pharmaceutical composition according to claim 1(ii)a.

4. 4. The pharmaceutical composition of claim 1(ii)a or claim 3, wherein the ICANS is grade 1 or higher.

5. The pharmaceutical composition of claim 1(ii)b, wherein the chemotherapeutic agent comprises an alkylating agent or a topoisomerase inhibitor.

6. The pharmaceutical composition of claim 1(ii)b or claim 5, wherein the chemotherapeutic or anti-inflammatory agent is administered to reduce the number of CAR-T cells in the subject.

7. 2. The pharmaceutical composition of claim 1(ii)c, wherein the IL-6 inhibitor comprises tocilizumab, saluliumab, siltuximab, or clazakizumab.

8. The pharmaceutical composition of claim 1(ii)c or claim 7, wherein the IL-6 inhibitor is administered to reduce IL-6 levels to within the normal reference range of 0 to 2 pg / ml.

9. The pharmaceutical composition of claim 1(ii)d, wherein the IFNγ inhibitor is emapalumab.

10. 10. The pharmaceutical composition of claim 1(ii)d or claim 9, wherein the IFNγ inhibitor is administered to reduce IFNγ levels to within the normal reference range of less than about 2.0 pg / mL.

11. 2. The pharmaceutical composition of claim 1(ii)e, wherein the anti-inflammatory agent comprises an IL-6 inhibitor, such as tocilizumab, or a steroid.

12. The anti-inflammatory agent reduces lymphocyte count by 0.8 to 3.0×10 ９ 12. The pharmaceutical composition of claim 1(ii)e or claim 11, administered to reduce serum erythrocyte sedimentation rate (SSE) to within the normal reference range.

13. 2. The pharmaceutical composition of claim 1(i), wherein the subject has a tumor burden characterized by bone marrow plasmacytosis less than 50%, serum M protein levels less than 3 g / dL, and serum free light chain levels less than 3000 mg / L.

14. 2. The pharmaceutical composition of claim 1(iii), wherein the method further comprises administering to the subject an IL-6 inhibitor to achieve the normal reference range of IL-6 in the subject prior to administering the pharmaceutical composition.

15. the CAR-T cell therapy is a B cell maturation agent (BCMA) CAR-T cell therapy; 2. The pharmaceutical composition of claim 1, wherein the BCMA CAR-T cell therapy is siltacabtadine autolucel (cilta-cel).

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