Methods of treating lupus with CD19-CD20 dual car-t cells

Abstract

The present disclosure provides methods of treating a systemic lupus erythematosus (e.g. highly active, severe, treatment-refractory systemic lupus erythematosus) by administering a chimeric antigen receptor T cell (CAR-T cell) comprising an anti-CD20 chimeric antigen receptor and an anti-CD19 chimeric antigen receptor. The present disclosure also provides a dose infusion bag comprising a composition comprising CAR-T cells comprising an anti-CD20 chimeric antigen receptor and an anti-CD19 chimeric antigen receptor.

Description

Attorney Docket No.: 000218-0093-W01METHODS OF TREATING LUPUS WITH CD19-CD20 DUAL CAR-T CELLS CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims priority to and benefit of U.S. Provisional Patent Application No. 63 / 720,072, filed November 13, 2024, the contents of which are herein incorporated by reference in their entireties.INCORPORATION-BY-REFERENCE OF SEQUENCE LISTING

[0002] The present application contains a Sequence Listing which has been submitted in XML format via Patent Center and is hereby incorporated by reference in its entirety. Said XML copy, created on November 12, 2025, is named “000218-0093-W01-SL.xml” and is 10,962 bytes in size.TECHNICAL FIELD

[0003] The present disclosure pertains to the field of treatment of lupus, such as systemic lupus erythematosus (SLE), using dual chimeric antigen receptor T (CAR-T) cells comprising a chimeric antigen receptor (CAR) encoding an anti-CD20 antibody and a CAR encoding an anti-CD19 antibody.BACKGROUND

[0004] SLE is a B cell-driven autoantibody-mediated disease in which B-cell depletion leads to clinical efficacy. SLE is characterized by multisystem involvement and immunological abnormalities, including dysfunction of B cells and T cells. Much of the tissue damage is thought to occur through autoantibody formation and immune complex deposition, but the underlying cause remains unknown. Disease manifestations can range from mild to severe and potentially life-threatening, with patients displaying acute, subacute, and chronic patterns of progression.

[0005] Current standard-of-care therapies for SLE include oral corticosteroids, antimalarial therapies (such as hydroxychloroquine), and conventional immunosuppressants (such as mycophenolate mofetil (MMF), azathioprine, methotrexate, and cyclophosphamide (CYC)). In addition, belimumab, an inhibitor of B cell-activating factor (BAFF), and anifrolumab, an anti-interferon (IFN) a / p receptorAttorney Docket No.: 000218-0093-W01mAb, have been approved for patients with SLE who are receiving standard-of-care therapy. Although rituximab, a type I anti-CD20 antibody, is used off-label to treat lupus in clinical practice, the drug failed to meet efficacy endpoints in randomized controlled trials for SLE and LN (Merrill et al. 2010; Rovin et al. 2012). More recently, obinutuzumab, a type II anti-CD20 antibody, demonstrated a clinically meaningful benefit over standard-of-care therapies alone in a Phase II trial in LN (Furie et al. 2022). Obinutuzumab caused more rapid and deeper B-cell depletion than rituximab, suggesting that a higher level of B-cell depletion may be necessary to achieve clinical efficacy in lupus (Gomez Mendez et al. 2018; Furie et al. 2022).

[0006] Despite the use of available therapies, patients with lupus rarely achieve remission. Severe side effects limit long-term use of current treatments, contributing to poor quality of life and significant psychological burden. These patients face double the risk of cardiovascular disease and osteoporosis, and a threefold higher mortality rate compared to the general population. A high unmet need remains for safer and more effective treatments to improve patient outcomes (Hoi et al. 2024; Siegel and Sammaritano 2024).SUMMARY

[0007] In a first aspect, the present disclosure provides methods of treating systemic lupus erythematosus (SLE) in a human subject in need thereof comprising administering a P-CD19CD20-ALLO1 chimeric antigen receptor T cell (CAR-T cell) to the subject.

[0008] In a second aspect, the present disclosure provides the use of a P-CD19CD20-ALLO1 chimeric antigen receptor T cell (CAR-T cell) in the manufacture of a medicament for treating systemic lupus erythematosus (SLE) in a human subject in need thereof.

[0009] In a third aspect, the present disclosure provides a P-CD19CD20-ALLO1 chimeric antigen receptor T cell (CAR-T cell) for use in treating systemic lupus erythematosus (SLE) in a human subject in need thereof.

[0010] In some embodiments of any of the above aspects, the SLE is selected from the group consisting of severe SLE; treatment refractory SLE; highly active SLE; severe, treatment-refractory SLE; severe, highly active SLE; highly active, treatmentrefractory SLE; and highly active, severe, treatment-refractory SLE. In someAttorney Docket No.: 000218-0093-W01embodiments, the SLE is highly active, severe, treatment-refractory SLE. In some embodiments, the highly active SLE is identified by (a) British Isles Lupus Assessment Group (BILAG)-2004 Category A disease in one or more organ systems and / or Category B disease in two or more organ systems (excluding constitutional, musculoskeletal, and / or mucocutaneous organ systems for Category B); (b) a Systemic Lupus Erythematosus Disease Activity Index 2000 (SLEDAI-2K) score greater than or equal to 8; and (c) a Physician’s Global Assessment (PGA) score greater than or equal to 1.0 on a 0 to 3 Visual Analog Scale (VAS). In some embodiments of any of the above aspects, the subject suffers from lupus nephritis. In some embodiments of any of the above aspects, the subject does not suffer from lupus nephritis.

[0011] In some embodiments of any of the above aspects, the CAR-T cell is administered to the subject intravenously or is formulated to be administered to the subject intravenously. In some embodiments of any of the above aspects, the CAR-T cell is administered as a single dose or is formulated to be administered as a single dose. In some embodiments, the CAR-T cell is administered in multiple doses or is formulated to be administered in multiple doses. In some embodiments, each of the multiple doses is administered between 3 and 5 years after the previous dose. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 20 x 106cells and about 1.2 x 109cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 20 x 106cells and about 480 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 5 x 106cells and about 1.2 x 109cells.

[0012] In some embodiments of any of the above aspects, administration of the CAR-T cell results in (a) a detectable number of CAR transgene copies in the blood of the subject, (b) a detectable number CAR-T cells in the blood of the subject, (c) the depletion of B-cell levels in the subject by more than 90 percent, (d) immunosuppression-free remission in the subject for at least 6 months, (e) a time to first SLE flare of at least 6 months; (f) a total number of SLE flares less than one per year; (g) reduced glucocorticoid and / or immunosuppressant use as compared to prior to treatment; (h) reduced titers antinuclear antibody (ANA) and double stranded DNA (dsDNA) as compared to prior to treatment; or (i) reduced titers of lupus-associated autoantibodies (e.g., anti-Smith and anti-RNP) as compared to prior to treatment.Attorney Docket No.: 000218-0093-W01

[0013] In some embodiments of any of the above aspects, administration of the CAR-T cell results in (a) an incidence of Grade 2 cytokine release syndrome (CRS) less than 30 percent, (b) an incidence of immune effector cell-associated neurotoxicity syndrome (ICANS) less than 20 percent, (c) an incidence of hemophagocytic lymphohistiocytosis (HLH) less than 10 percent, (d) a risk of serious infection no more than 10% more compared to the incidence of serious infection in patients receiving standard of care at 6 months after treatment, (e) an incidence of immune rejection and graft-versus-host disease (GVHD) less than 10 percent, or (f) an incidence of new T-cell malignancies less than 0.1%.

[0014] In some embodiments of any of the above aspects, the subject has not been previously administered a therapeutic agent to treat SLE. In some embodiments of any of the above aspects, the subject has been previously administered a therapeutic agent to treat SLE. In some embodiments, the subject is refractory to the previously administered therapeutic agent. In some embodiments, the previously administered therapeutic agent is selected from a corticosteroid, an antimalarial therapy, an immunosuppressant, a B-cell depleting therapy, and a cytokine blocker. In some embodiments, the antimalarial therapy is hydroxychloroquine. In some embodiments, the immunosuppressant is selected from the group consisting of cyclophosphamide, azathioprine, mycophenolate mofetil (MMF), mycophenolic acid, and methotrexate. In some embodiments, the B-cell depleting therapy is selected from the group consisting of rituximab, ocrelizumab, ofatumumab, belimumab and obinutuzumab. In some embodiments, the cytokine blocker is anifrolumab.

[0015] In some embodiments of any of the above aspects, the CAR-T cell is administered or formulated to be administered in combination with one or more additional therapeutic agents. In some embodiments, the CAR-T cell is administered or formulated to be administered sequentially with the one or more additional therapeutic agents. In some embodiments, the CAR-T cell is administered or formulated to be administered simultaneously with the one or more additional therapeutic agents. In some embodiments, the one or more additional therapeutic agents is selected from an immunosuppressant, a B-cell depleting therapy, a calcineurin inhibitor, another biologic or targeted therapy, an IL-1 inhibitor, a corticosteroid, a kinase inhibitor, a cytokine blocker, or a combination thereof. In some embodiments, the immunosuppressant is selected from the group consisting of cyclophosphamide, azathioprine, mycophenolate mofetil (MMF), mycophenolic acid, and methotrexate. InAttorney Docket No.: 000218-0093-W01some embodiments, the B-cell depleting therapy is selected from the group consisting of rituximab, ocrelizumab, ofatumumab, belimumab and obinutuzumab. In some embodiments, the calcineurin inhibitor is selected from the group consisting of ciclosporin, tacrolimus, and voclosporin. In some embodiments, the kinase inhibitor is ruxolitinib. In some embodiments, the cytokine blocker is selected from the group consisting of ruxolitinib, etoposide, or emapalumab.

[0016] In some embodiments of any of the above aspects, the subject receives a lymphodepleting chemotherapy (LDC) regimen prior to the administration of CAR-T cell. In some embodiments, the LDC regimen comprises cyclophosphamide (300-750 mg / m2IV) and fludarabine (0-30 mg / m2IV) for 3 consecutive days. In some embodiments, the LDC regimen is administered on Days -5, -4, and -3, and the CAR-T cells is administered or to be administered on Day 1.BRIEF DESCRIPTION OF THE DRAWINGS

[0017] Figure 1 provides a diagram showing the anti-CD19 and anti-CD20 CAR components of the P-CD19CD20-ALLO1 Transposon. [CAR = chimeric antigen receptor; cHS4 = chicken hypersensitive site 4; DHFR = dihydrofolate reductase; EEF1A1 = Eukaryotic translation elongation factor 1 alpha 1; HBB = hemoglobin subunit beta; iCasp9 = inducible caspase 9; ITR = inverted terminal repeat; nt = nucleotides; SV40 poly(A) = simian virus 40 polyadenylation; T2A = Thosea asigna virus 2 A, UTR = untranslated region; VH = variable heavy chain],

[0018] Figure 2 provides an illustration of P-CD19CD20-ALLO1 chimeric antigen receptor (CAR) on the surface of the T cell membrane.

[0019] Figures 3A and 3B provide graphs showing the % lysis of antigen positive (Raji) tumor cells (Figure 3 A) and antigen negative (K562) tumor cells (Figure 3B) by P-CD19CD20-ALLO1 across Effector to Target (E:T) ratios of 20:1-0.039:1. Each data point shows average of triplicates for Raji cell killing and average of duplicates for K562 cell killing.

[0020] Figures 4A-4D provide graphs showing IFN-y (Figures 4A and 4B) and Granzyme B (Figures 4C and 4D) levels in supernatants from coculture assays of P-CD19CD20-ALLO1 CAR-T cells (“Lot X (CAR-T)”) and mock transfected controlsAttorney Docket No.: 000218-0093-W01(“Lot X (mock)”) with either antigen-positive cells (Raji) (Figures 4A and 4C) or antigen-negative cells (Jurkat) (Figures 4B and 4D).

[0021] Figure 5 provides the results of a flow cytometry -based cytotoxicity assay against PBMC from SLE patients. Cytotoxic activity of 3 lots of P-CD19CD20-ALLO1 CAR-T cells against peripheral blood mononuclear cell (PBMC) samples isolated from SLE patients at E:T ratios of 1 : 1 and 5:1 was evaluated by flow cytometry after 48 hours co-culture. Each cell population in the PBMC sample was quantified using Countbright™ beads and numbers were normalized to the PBMC samples alone. Abbreviations: CAR-T = chimeric antigen receptor T cell; E:T = Effector-to-Target ratio; NK = natural killer; PBMC = peripheral blood mononuclear cell; SLE = systemic lupus erythematosus.

[0022] Figures 6A-6C provide flow cytometry-based cytotoxicity assay with PBMCs and isolated T and B cells from healthy donors. Cytotoxic activity of 3 lots of P-CD19CD20-ALLO1 CAR-T cells against PBMC samples (Figure 6A) as well as isolated T cells (Figure 6B) and B cells (Figure 6C) from three healthy donors at E:T ratios of 1:1 and 5:1 was evaluated by flow cytometry after 48 hours co-culture. Each cell population in the PBMC sample was quantified using Countbright™ beads and numbers were normalized to the PBMC samples alone without CAR-T cells. Abbreviations: CAR-T = chimeric antigen receptor T cell; E:T = Effector-to-Target ratio; HD = Healthy donor; NK = natural killer; PBMC = peripheral blood mononuclear cell; SLE = systemic lupus erythematosus.

[0023] Figure 7 provides flow cytometry-based cytotoxicity assays of CD20-positive T cells present in PBMC samples from SLE patients. Cytotoxic activity of 3 lots of P-CD19CD20-ALLO1 CAR-T cells against CD20-positive T cells in PBMC samples from SLE patients at E:T ratios of 1 : 1 and 5 : 1 was evaluated by flow cytometry after 48 hours co-culture. CD20-positive T cells within PBMC samples after treatment with the indicated CAR-T cells were quantified using Countbright™ beads and numbers were normalized to PBMC samples alone (no CAR-T) to express data as % of surviving cells after treatment with CAR-T. Only patient PBMC samples containing at least 1000 CD20-positive T cells per well were included in this assessment. Abbreviations: CAR = chimeric antigen receptor; E:T = Effector to target; SLE = Systemic lupus erythematosus.Attorney Docket No.: 000218-0093-W01

[0024] Figures 8A and 8B provide graphs showing the relative fold change of viable P-CD19CD20-ALLO1 cells after 24 and 48 hours of treatment with rimiducid (API 903) relative to the untreated control (0 nM). Data are the average of triplicate wells with error bars presented by mean ± standard deviation.

[0025] Figures 9A and 9B provide graphs showing CD 19 and CD20 expression on target cells assessed by flow cytometry (Figure 9A) and specific cytotoxicity of P-CD19CD20-ALLO1 CAR-T cells against CD19-expressing cells and CD20-expressing cells after 72-hour coculture at 1:1 Effector to Target (E:T) ratio (Figure 9B).

[0026] Figures 10A and 10B provide graphs showing CAR expression (Figure 10A) and CAR density (Figure 10B) in dual CAR constructs compared with single CAR constructs in rested and phorbol ester + ionomycin (PMA / I)-stimulated cells evaluated by flow cytometry. Abbreviations: CAR = chimeric antigen receptor; gMFI = geometric mean fluorescence intensity.

[0027] Figures 11A-11J provide graphs showing the in vitro potency of P-CD19CD20-ALLO1 compared to its CD 19- and CD20-single targeting counterparts using serial restimulation against the CD 19- and CD20-positive (WT) RAJI cell line, as well as RAJI cells engineered to express only CD19 or CD20. Figures 11A-11C show RAJI cell killing in WT, CD19KO or CD20KO RAJI cell lines, respectively. Figures 1 ID - 11 J show the induction of secretion of the cytokines IL-2 (Figure 1 ID), TNFa (Figure HE), IFNy (Figure 1 IF), Granzyme A (Figure 11G), Granzyme B (Figure 11H), perforin (Figure 111) and Granulysin (Figure 11 J). Abbreviations: E:T = Effector to target.

[0028] Figure 12 provides a schema for a Phase I clinical trial to evaluate the safety, tolerability, pharmacokinetics (PK), cellular kinetics (CK), pharmacodynamics (PD), and efficacy of CD19CD20-ALLO1 CAR- T cells, showing patients enrolled in two stages: a dose-escalation stage and an expansion stage and details on these two stages. The dose-escalation stage will have a 3 + 3 design per the dose-escalation rules. Up to two optional cohorts may be opened based on emerging data. The initial participant receiving P-CD19CD20-ALLO1 in each cohort will be observed for a minimum of 28 days before additional participants are treated in the same cohort. Subsequent participants will be staggered such that their P-CD19CD20-ALLO1 isAttorney Docket No.: 000218-0093-W01administered at least 14 days apart. There is a dose-limiting toxicity (DLT) window of 28 days.

[0029] Figure 13 provides the dosing and screen schedule for the CD19CD20-ALLO1 CAR- T clinical trial.aAfter discharge, participants must remain within a 1-hour travel radius from the site through Day 29.bAfter 5 years, participants will be transitioned to a separate protocol for continued monitoring for up to 15 years after P-CD19CD20-ALLO1 infusion. [DLT = dose-limiting toxicity; E = enrollment],DETAILED DESCRIPTIONGeneral

[0030] Practice of the methods, as well as preparation and use of the compositions disclosed herein employ, unless otherwise indicated, conventional techniques in molecular biology, biochemistry, chromatin structure and analysis, computational chemistry, cell culture, recombinant DNA and related fields as are within the skill of the art. These techniques are fully explained in the literature.

[0031] The term “herein” means the entire application.

[0032] It should be understood that any of the embodiments described herein, including those described under different aspects of the disclosure and different parts of the specification (including embodiments described only in the Examples) can be combined with one or more other embodiments disclosed herein, unless explicitly disclaimed or improper. Combination of embodiments are not limited to those specific combinations claimed via the multiple dependent claims.

[0033] Any publications, patents and published patent applications referred to in this application are specifically incorporated by reference herein. In case of conflict, the present specification, including its specific definitions, will control.

[0034] Throughout this specification, the word “comprise,” or variations such as “comprises” or “comprising,” which is synonymous with “including,” “containing,” or “characterized by,” is inclusive or open-ended and does not exclude additional, unrecited elements or method steps.

[0035] Throughout the specification, where compositions are described as having, including, or comprising (or variations thereof), specific components, it is contemplated that compositions also may consist essentially of, or consist of, the recited components. Similarly, where methods or processes are described as having, including,Attorney Docket No.: 000218-0093-W01or comprising specific process steps, the processes also may consist essentially of, or consist of, the recited processing steps. Further, it should be understood that the order of steps or order for performing certain actions is immaterial so long as the compositions and methods described herein remains operable. Moreover, two or more steps or actions can be conducted simultaneously.

[0036] The term “consisting of’ excludes any element, step, or ingredient not specifically recited.

[0037] The term “consisting essentially of’ limits the scope of a disclosure to the specified materials or steps and those that do not materially affect the basic and novel characteristic(s) of the disclosure.

[0038] Any example(s) following the term “e.g.” or “for example” is not meant to be exhaustive or limiting.

[0039] The articles “a,” “an” and “the” are used herein to refer to one or to more than one ( / .< ., to at least one) of the grammatical object of the article. By way of example, “an element” means one element or more than one element.

[0040] As used herein, the term “about” modifying the quantity of an ingredient, parameter, calculation, or measurement in the compositions employed in the methods of the disclosure refers to the variation in the numerical quantity that can occur, for example, through typical measuring and liquid handling procedures used for making isolated polypeptides or pharmaceutical compositions in the real world; through inadvertent error in these procedures; through differences in the manufacture, source, or purity of the ingredients employed to make the compositions or carry out the methods; and the like without having a substantial effect on the chemical or physical attributes of the compositions or methods of the disclosure. Such variation can be within an order of magnitude, typically within 10%, more typically still within 5%, of a given value or range. The term “about” also encompasses amounts that differ due to different equilibrium conditions for a composition resulting from a particular initial mixture. Whether or not modified by the term “about,” the paragraphs include equivalents to the quantities. Reference to “about” a value or parameter herein includes (and describes) embodiments that are directed to that value or parameter per se. For example, description referring to “about X” includes description of “X.” Numeric ranges are inclusive of the numbers defining the range.

[0041] The term “or” as used herein should be understood to mean “and / or,” unless the context clearly indicates otherwise.Attorney Docket No.: 000218-0093-W01

[0042] Notwithstanding that the numerical ranges and parameters setting forth the broad scope of the disclosure are approximations, the numerical values set forth in the specific examples are reported as precisely as possible. Any numerical value, however, inherently contains certain errors necessarily resulting from the standard deviation found in their respective testing measurements. Moreover, all ranges disclosed herein are to be understood to encompass any and all subranges subsumed therein. For example, a stated range of “1 to 10” should be considered to include any and all subranges between (and inclusive of) the minimum value of 1 and the maximum value of 10; that is, all subranges beginning with a minimum value of 1 or more, e.g., 1 to 6.1, and ending with a maximum value of 10 or less, e.g., 5.5 to 10. The disclosure of a range should also be considered as disclosure of the endpoints of that range.

[0043] Exemplary methods and materials are described herein, although methods and materials similar or equivalent to those described herein can also be used in the practice or testing of the present application. The materials, methods, and examples are illustrative only and not intended to be limiting.Definitions

[0044] The following terms, unless otherwise indicated, shall be understood to have the following meanings:

[0045] As used herein, the term “ablated” refers to a genetic engineering technique that involves the alteration or deletion of genetic sequence information to silence a gene or make it “null” ( / .< ., a DNA sequence variation that results in no gene product). The RNA or protein encoded by a null gene is either absent or not detectable at the molecular level or does not function at the phenotypic level.

[0046] “Administering” or “administration of’ a substance, a compound or an agent to a subject refers to the contact of that substance, compound or agent to the subject or a cell, tissue, organ or bodily fluid of the subject. For example, a compound or an agent can be administered intravenously. Administering can also be performed, for example, once, a plurality of times, and / or over one or more extended periods. In some embodiments, the administration includes both direct administration, including self-administration, and indirect administration, including the act of prescribing a drug. For example, as used herein, a physician who instructs a subject to self-administer a drug, or to have the drug administered by another and / or who provides a subject with a prescription for a drug is administering the drug to the subject.Attorney Docket No.: 000218-0093-W01

[0047] The term “amino acid,” as used herein, refers to one of the 20 naturally occurring amino acids that are coded for by DNA and RNA.

[0048] As used herein, the term “antibody” or “Ab” refers to an immunoglobulin molecule capable of recognizing and binding to a specific target or antigen, such as a carbohydrate, polynucleotide, lipid, polypeptide, etc., through at least one antigen recognition site, located in the variable region of the immunoglobulin molecule. As used herein, the term “antibody” can encompass any type of antibody, including but not limited to monoclonal antibodies, polyclonal antibodies, human antibodies, engineered antibodies (including humanized antibodies, fully human antibodies, chimeric antibodies, single-chain antibodies, artificially selected antibodies, CDR-granted antibodies, etc.) that specifically bind to a given antigen.

[0049] As used herein, the term “antigen-binding fragment” refers to a portion (or fragment) of an antibody that retains the antibodies binding specificity. Accordingly, as used herein, an antigen-binding fragment retains the six CDRs of the reference antibody. In some embodiments, the antigen-binding fragment includes a VHH antibody. In such cases, the antigen-binding fragment only includes the three heavy chain CDRs of the reference antibody.

[0050] The terms “chimeric antigen receptor” and “CAR” are used interchangeably herein and refer to a receptor protein that has been engineered to target a specific antigen and, upon binding the antigen, activate an immune cell, such as a T cell. The chimeric receptors combine both antigen-binding and T cell activating functions into a single receptor. A CAR typically comprises (a) an ectodomain comprising an antigen recognition region; (b) a transmembrane domain, and (c) an endodomain comprising at least one costimulatory domain.

[0051] As used herein, the term “highly active” SLE refers to a high disease activity status (HDAS), which is a measure of increased SLE disease activity and a higher likelihood of organ damage. HDAS is typically defined using the Systemic Lupus Erythematosus Disease Activity Index-2000 (SLEDAI-2K). The total SLEDAI-2K score falls between 0 and 105. A SLEDAI-2K score of 6 is considered clinically important and affects the decision to treat with a probability of initiating therapy in more than 50% of cases. Meaningful improvement is best defined as a reduction in SLEDAI-2K of 4. See, e.g., Mikdashi J, et al. Arthritis Res Ther. 2015; vol. 17(1): 183, incorporated herein by reference in its entirety.Attorney Docket No.: 000218-0093-W01

[0052] The terms “nucleic acid,” “polynucleotide” and “oligonucleotide” are used interchangeably and refer to a deoxyribonucleotide or ribonucleotide polymer, in linear or circular conformation, and in either single- or double-stranded form. For the purposes of the present disclosure, these terms are not to be construed as limiting with respect to the length of a polymer.

[0053] As used herein, the term “P-CD19CD20-ALLO1” CAR-T cells refers to specific healthy donor-derived allogeneic CAR T cells that targets both CD 19 and CD20 antigens. More details regarding specific genetic modification introduced into P-CD19CD20-ALLO1 CAR-T cells are provided infra.

[0054] The terms “patient,” “subject” and “individual” are used interchangeably herein and refer to a human in need of treatment. In some embodiments, the subject is in need of treatment of a systemic lupus erythematosus (SLE), such as highly active, severe, treatment-refractory SLE.

[0055] As used herein, the terms “polypeptide,” “peptide” and “protein” are used interchangeably to refer to a polymer of amino acid residues. Expression of a protein in a cell can result from delivery of the protein to the cell or by delivery of a polynucleotide encoding the protein to a cell, wherein the polynucleotide is transcribed, and the transcript is translated, to generate the protein. Trans-splicing, polypeptide cleavage and polypeptide ligation can also be involved in expression of a protein in a cell. Methods for polynucleotide and polypeptide delivery to cells are known in the art.

[0056] As used herein, the term “remission” in the context of SLE refers to the definition of remission in SLE (DORIS), which requires an SLEDAL2K score of 0 and a Physician’s Global Assessment (PGA) score less than 0.5 (on the 0-3 scale). See, e.g., van Vollenhoven, et al. Lupus Sci Med. 2021, vol. 8(l):e000538, incorporated herein by reference in its entirety. The term “immunosuppression-free remission” refers to a patient who remains in remission without the need for any immunosuppression therapy.

[0057] As used herein, the term “severe” in the context of systemic lupus erythematosus (SLE) refers to a major organ-threatening form of the disease that can include life-threatening complications. These complications can include: nephritis, cerebritis, myelitis, pneumonitis, and mesenteric vasculitis.

[0058] The terms “therapeutically effective amount” and “effective amount” are used interchangeably herein an refer to that amount of the therapeutic agent being administered, as a single agent or in combination with one or more additional agents,Attorney Docket No.: 000218-0093-W01which will relieve to some extent one or more of the symptoms of the condition being treated. In some embodiments, the therapeutically effective amount is an amount sufficient to effect beneficial or desired clinical results. With respect to the treatment of SLE, a therapeutically effective amount refers to that amount which has at least one of the following effects: reduction a symptom of SLE, elimination of a symptom of SLE, remission, reduction of a biomarker of SLE, elimination of a biomarker of SLE, or improvement in a clinical disease activity assessment. Non-limiting examples of common SLE symptoms include fatigue; fever; joint pain, stiffness and swelling; a butterfly-shaped rash on the face that covers the cheeks and bridge of the nose or rashes elsewhere on the body; skin lesions that appear or worsen with sun exposure; fingers and toes that turn white or blue when exposed to cold or during stressful periods; shortness of breath; chest pain; dry eyes; and headaches, confusion and memory loss. Non-limiting examples of biomarkers include cytokines associated with B cell function and activation (e.g., soluble B cell-activating factor (sBAFF), and B cell maturation antigen (BCMA), autoantibodies associated with SLE, T-cell subsets (e.g., activation and proliferation), and T cell engraftment (IL-7 and IL-15). Non-limiting examples of clinical disease activity assessments include Systemic Lupus Erythematosus Disease Activity Index 2000 (SLEDAI-2K), British Isles Lupus Assessment Group 2004 (BILAG-2004), Physician’s Global Assessment (PGA), Cutaneous Lupus Erythematosus Disease Area and Severity (CLASI), and tender / swollen joint counts. Each of these assessments is described in more detail in Example 7, infra. The effective amount that may be used in the present disclosure varies depending upon the manner of administration, the age, body weight, and general health of the subject. The appropriate amount and dosage regimen can be determined using routine skill in the art.

[0059] As used herein, the terms “treatment” (and grammatical variations thereof such as “treat” or “treating”) and “therapeutic treatment” refers to clinical intervention in an attempt to alter the natural course of a disease in the individual being treated performed during the course of clinical pathology. Desirable effects of treatment include, but are not limited to, preventing occurrence or recurrence of disease, alleviation of symptoms, diminishment of any direct or indirect pathological consequences of the disease, decreasing the rate of disease progression, amelioration or palliation of the disease state, and remission or improved prognosis. In some aspects,Attorney Docket No.: 000218-0093-W01the CAR-T cells of the disclosure are used to delay development of a disease or to slow the progression of a disease.

[0060] As used herein, the term “treatment-refractory” in the context of systemic lupus erythematosus (SLE) refers to a condition in which the disease does not respond to one or more standard therapies. Non-limiting examples of standard therapies for SLE include oral corticosteroids, antimalarial therapies (such as hydroxychloroquine), and conventional immunosuppressants (such as mycophenolate mofetil (MMF), azathioprine, methotrexate, and cyclophosphamide (CYC)). In addition, belimumab, an inhibitor of B cell-activating factor (BAFF) and anifrolumab, an anti-interferon (IFN) a / p receptor mAb, have been approved for patients with SLE who are receiving standard-of-care therapy.

[0061] The terms “variable human heavy-chain domain,” “VHH” and “nanobody” are used interchangeably herein and refer to a single antigen-binding domain derived from heavy-chain antibodies. A heavy-chain antibody is an antibody which consists only of two heavy chains and lacks the two light chains usually found in antibodies. Naturally-occurring heavy-chain antibodies can be produced by camelids, such as alpacas, llamas, camels, and vicunas. Fully human and fully humanized VHHs are known in the art. See, e.g., WO2019126574A1; and Rouet, R. et al., J. Biol. Chem, 2015, vol. 290(19): 11905-17, each of which is incorporated herein by reference in its entirety.

[0062] The terms “VCAR” and “VHH CAR” are used interchangeably herein and refer to a CAR in which the antigen recognition region comprises at least one VHH.

[0063] The sequences referenced herein are provided in Table 1, infra.Table 1: Amino acid sequences used in the methods of the present disclosure.Attorney Docket No.: 000218-0093-W01P-CD19CD20-ALLO1 CAR-T Cells

[0064] P-CD19CD20-ALLO1 CAR-T cells target both CD 19 and CD20 and have been genetically modified using an electroporation-based, non-viral (DNA transposon) gene delivery system called the piggyBac® DNA modification system.Attorney Docket No.: 000218-0093-W01

[0065] P-CD19CD20-ALLO1 CAR-T cells were manufactured from cryopreserved peripheral blood mononuclear cells (PBMCs) obtained by leukapheresis of healthy donors. The primary attributes of P-CD19CD20-ALLO1 are conferred by the genes introduced in the PB transposon, including an anti-CD19 targeting variable human heavy-chain domain (VHH)-based CAR (VCAR) gene, an anti-CD20 targeting VCAR gene, a dihydrofolate reductase (DHFR) selection mutant protein (mutein) gene, and an inducible caspase 9 (iCasp9)-based safety switch gene. See, Figure 1.Additionally, P-CD19CD20-ALLO1 CAR-T cells are deficient for the native T-cell receptor (TCR) and human leukocyte antigen (HLA)-A, B and C major histocompatibility complex I (MHC-I) proteins, as a result of the ablation of the TCRP chain (TCRB) and p2-microglobin (P2M) genes using the Cas-CLOVER™ Site-Specific Gene Editing System. The TCRB and B2M genes are genes known to mediate rejection, e.g., graft-vs-host (GVH) and host-vs-graft (HVG) responses. Accordingly, their removal decreases the likelihood that the subject being treated will reject the P-CD19CD20-ALLO1 allogeneic CAR-T cells being administered.

[0066] P-CD19CD20-ALLO1 CAR-T cells comprise 2 CAR separate expression cassettes. The first CAR expression cassette encodes an extracellular CD 19 binding VHH protein (SEQ ID NO: 1) fused to a CD8a spacer (SEQ ID NO: 4), a CD8a transmembrane domain (SEQ ID NO: 5), an intracellular 4-1BB signaling domain (SEQ ID NO: 6) and a CD3(^ signaling domain (SEQ ID NO: 7). The second CAR expression cassette encodes an extracellular CD20-binding VHH protein (SEQ ID NO: 2) fused to a CD8a spacer (SEQ ID NO: 4), a CD8a transmembrane domain (SEQ ID NO: 5), an intracellular 4-1BB signaling domain (SEQ ID NO: 6) and a CD3(^ signaling domain (SEQ ID NO: 7). See, Figure 2 for a schematic representation of the CD19 VCAR and the CD20 VCAR. Both expression cassettes also include nucleotide sequences encoding a CD8a signal peptide (SEQ ID NO: 3), which gets cleaved from the mature VCAR in the endoplasmic reticulum.

[0067] CD 19 and CD20 are lineage-specific antigens expressed on the cell surface of certain B cells. CD 19 is expressed more broadly from early B-cell genesis through maturation and remains present on a subset of plasma cells, while CD20 is acquired during the late stage of B-cell lymphopoiesis and lost upon their differentiation into plasma cells.

[0068] Although monoclonal antibodies targeting CD 19 and CD20 have demonstrated benefit in subsets of patients, relapse is common, and resistance toAttorney Docket No.: 000218-0093-W01monoclonal antibody therapy may be due to incomplete depletion of B cells residing within secondary lymphoid organs. See, e.g., Nunez D, et al. Mol Ther Methods Clin Dev. 2023; vol. 31 : 101104, incorporated herein by reference in its entirety. There are several mechanisms for CD 19 or CD20 antigen loss following treatment with antibodies targeting CD 19 or CD20, including CD 19 or CD20 gene mutations and frameshift mutations leading to protein truncation, non-functional transmembrane domains, and antigen loss; and alternative splicing of the CD 19 or CD20 protein; emergence of point mutations in the CD 19 or CD20 protein leading to altered epitope recognition by CD 19 or CD20 antibodies.

[0069] Moreover, autologous CAR-T cell therapies require well-functioning T cells at apheresis from the patient. For patients who may have received multiple treatment regimens, including anti-CD20 antibodies, which can deplete T cells, it is possible that the T-cell quality is poor at apheresis.

[0070] P-CD19CD20-ALLO1 CAR-T cells address the issues that affect antiCD 19 and / or anti-CD20 therapies. Without being bound by theory, it is unlikely that treatment resistant clones will develop to a CAR-T therapy that is able to recognize two antigens simultaneously. Moreover, because P-CD19CD20-ALLO1 CAR-T cells are allogeneic, the T-cells from which they are derived are obtained from healthy donors that have never received cytotoxic chemotherapy and, therefore should result in a product with highly functional T-cells with good cell killing activity. Accordingly, P-CD19CD20-ALLO1 CAR-T cells may provide deep depletion of tissue resident B cells that are refractory to other B cell-depleting modalities facilitating an “immune reset” where the pathogenic B cell repertoire is eliminated, and a predominantly naive immune system is reconstituted resulting in substantially reduced autoantibody secretion and potentially providing sustained drug-free remission to patients with highly active, severe, and treatment-refractory SLE.

[0071] An iCasp9 “safety switch” is an extra safety feature not found in most CAR-T cells and is designed to allow for rapid killing of P-CD19CD20-ALLO1 cells, if clinically indicated, by intravenous (IV) administration of the activation agent, rimiducid, a synthetic small molecule dimerizing drug. The iCasp9 safety switch is the first gene encoded within the quad-cistronic P-CD19CD20-ALLO1 transgene. Because it is encoded upstream of the CD 19 and CD20 VCARs, there can be no expression of the VCARs or DHFR genes without expression of the iCasp9 switch. The iCasp9 sequence consists of the chemically induced dimerization (CID)-binding domain (i.e.Attorney Docket No.: 000218-0093-W01a mutant human FK506-binding protein 12 (FKBP12) domain) coupled to the signaling domain of caspase-9, an enzyme that is part of the apoptotic pathway. The amino acid sequence of the iCasp9 safety switch is provided in SEQ ID NO: 8 or 9. In some embodiments, the iCasp9 safety switch is encoded by the amino acid sequence of SEQ ID NO:8. In some embodiments, the iCasp9 safety switch is encoded by the amino acid sequence of SEQ ID NO:9. Infusion of rimiducid (AP1903), an investigational product and otherwise bio-inert small-molecule dimerizing drug, triggers dimerization of iCasp9 and subsequent activation of caspase-9, which leads to rapid induction of apoptosis of the iCasp9-containing CAR-T cells, preferentially killing activated cells expressing the transgene. Rimiducid eliminates P-CD19CD20-ALLO1 cells in a dosedependent and time-dependent manner in vitro (0.1 nM [40%], 1 nM [60%], and 10 nM [100%]).

[0072] The selection gene encoding a DHFR mutein enzyme (SEQ ID NO: 10) was used during manufacturing for ex vivo selection of P-CD19CD20-ALLO1 CAR-T cells to produce a more homogeneous product. The wild-type DHFR protein plays a role in DNA synthesis is rapidly dividing cells but is inhibited by methotrexate. Accordingly, when cells are exposed to methotrexate, they eventually die. The modified DHFR selection gene used in P-CD19CD20-ALLO1 CAR-T cells, however, has a lower sensitivity to methotrexate. The modified DHFR gene is the fourth ( / .< ., last) in the quad-ci stronic P-CD19CD20-ALLO1 transgene. Accordingly, it is only expressed in cells that have expressed all of the iCasp9 safety switch, the CD 19 VCAR, and the CD20 VCAR and, therefore, can be used to select for such cells.

[0073] TCR knockout cells do not express surface TCR / CD3 complex and thus expand poorly in response to anti-CD3 agonist antibodies. To overcome this deficiency, the P-CD19CD20-ALLO1 manufacturing process uses an mRNA construct that encodes a booster molecule (CD2z booster) that is co-delivered via electroporation. CD2z mRNA encodes a chimeric fusion protein composed of human full-length CD2 and human CD3(^ intracellular signaling domain. When cells expressing the CD2z booster mRNA are treated with expansion reagents containing anti-CD2 agonist antibodies, binding of the CD2z booster rescues intracellular delivery of CD3(^ signals to the TCR knockout cells and results in optimal levels of expansion.Attorney Docket No.: 000218-0093-W01Method of treatment of Systemic Lupus Erythematosus (SLE), using P-CD19CD20-ALLO1 CAR-T cells

[0074] The present disclosure relates to methods of treating lupus, such as systemic lupus erythematosus (SLE), using P-CD19CD20-ALLO1 CAR-T cells, which are allogeneic dual chimeric antigen receptor T (CAR-T) cells comprising a chimeric antigen receptor (CAR) encoding an anti-CD20 antibody and a CAR encoding an antiCD 19 antibody. Without being bound by theory, P-CD19CD20-ALLO1 CAR-T cells may achieve a greater degree of B cell depletion compared with current monovalent B cell-targeting antibodies, including more extensive loss of tissue-infiltrating B cells and more efficient removal of cells with lower levels of CD 19 or CD20 expression, which may yield improved efficacy. The hypothesized ability of CAR-T cells to provide deep depletion of tissue resident B cells that are refractory to other B cell-depleting modalities may facilitate an “immune reset” where the pathogenic B cell repertoire is eliminated, and a predominantly naive immune system is reconstituted. Therefore, P-CD19CD20-ALLO1 CAR-T cells are hypothesized to deplete autoreactive B cells and plasmablasts, thus limiting the production of autoantibodies in SLE and potentially providing sustained drug-free remission to patients with highly active, severe, and treatment-refractory disease. P-CD19CD20-ALLO1 CAR-T cells are also hypothesized to deplete a subpopulation of CD20-positive T cells that have been described in some autoimmune diseases and that have a high pathogenic potential.

[0075] To confirm antigen-specific cytotoxicity against CD19-positive cells and CD20-positive cells in autoimmune samples, a series of nonclinical in vitro assays were performed to assess P-CD19CD20-ALLO1 CAR-T cell activity against primary peripheral blood mononuclear cells (PBMCs). See, Examples 2 and 3, infra. PBMCs were isolated from either healthy human donor controls or patients with autoimmune disease, including lupus. Three lots of P-CD19CD20-ALLO1 CAR-T cells were manufactured from 3 individual healthy donors and characterized for in vitro cytotoxicity and cytokine release against these PBMCs. All three lots of P-CD19CD20-ALLO1 CAR-T cells demonstrated antigen-specific cytotoxicity and cytokine release against B cells and CD20-expressing T cells. Neither off-target killing nor cytokine release was observed with antigen-negative cells, with the exception of monocytes in some donors, consistent with the well-known nonspecific activation of T cells by monocytes. The data disclosed herein, therefore, demonstrate the antigen-specific potency of P-CD19CD20-ALLO1 CAR-T cells and the potential for their deploymentAttorney Docket No.: 000218-0093-W01for the treatment of SLE, mediated by CD 19-positive and / or CD20-positive B and T cells.

[0076] Additionally, results from a comprehensive battery of in vitro and in vivo nonclinical pharmacology and toxicology studies of P-CD19CD20-ALLO1 CAR-T cells provide evidence of target pharmacology, demonstrate an acceptable safety profile, and support administration in humans. See, Examples 4 and 5, infra. The demonstrated effects reflect the specificity of P-CD19CD20-ALLO1 CAR-T cells for both intended targets present on cell lines expressing CD 19 and / or CD20, but not for cells without these surface markers. Genotoxicity and other toxicity studies demonstrated an acceptable safety profile with no evidence of cellular transformation and with very low frequencies of off-target gene editing and translocation events. The functionality of the iCasp9 safety switch was demonstrated to be intact.

[0077] In a first aspect, the present disclosure provides methods of treating systemic lupus erythematosus (SLE) in a human subject in need thereof comprising administering a P-CD19CD20-ALLO1 chimeric antigen receptor T cell (CAR-T cell) to the subject.

[0078] In a second aspect, the present disclosure provides the use of a P-CD19CD20-ALLO1 chimeric antigen receptor T cell (CAR-T cell) in the manufacture of a medicament for treating systemic lupus erythematosus (SLE) in a human subject in need thereof.

[0079] In a third aspect, the present disclosure provides a P-CD19CD20-ALLO1 chimeric antigen receptor T cell (CAR-T cell) for use in treating systemic lupus erythematosus (SLE) in a human subject in need thereof.

[0080] In some embodiments of any of the above aspects, the SLE is selected from the group consisting of severe SLE; treatment refractory SLE; highly active SLE; severe, treatment-refractory SLE; severe, highly active SLE; highly active, treatmentrefractory SLE; and highly active, severe, treatment-refractory SLE. In some embodiments, the SLE is severe SLE. In some embodiments, the SLE is treatment refractory SLE. In some embodiments, the SLE is highly active SLE. In some embodiments, the SLE is severe, treatment-refractory SLE. In some embodiments, the SLE is severe, highly active SLE. In some embodiments, the SLE is highly active, treatment-refractory SLE. In some embodiments, the SLE is highly active, severe, treatment-refractory SLE. In some embodiments, the highly active SLE is identified by (a) British Isles Lupus Assessment Group (BILAG)-2004 Category A disease in oneAttorney Docket No.: 000218-0093-W01or more organ systems and / or Category B disease in two or more organ systems (excluding constitutional, musculoskeletal, and / or mucocutaneous organ systems for Category B); (b) a Systemic Lupus Erythematosus Disease Activity Index 2000 (SLEDAI-2K) score greater than or equal to 8; and (c) a Physician’s Global Assessment (PGA) score greater than or equal to 1.0 on a 0 to 3 Visual Analog Scale (VAS). The BILAG-2004 index and SLEDAI-2K are both known in the art to be a valid assessment of disease activity in SLE. See, e.g., Yee, CS., et al., Arthritis Rheum. 2007 Dec; 56(12): 4113-4119; and Touma Z, et al. Lupus. 2010 Jan; 19(1):49-51, incorporated herein by reference in its entirety; see also, Example 7, infra. In some embodiments, highly active SLE is defined by a Systemic Lupus Erythematosus Disease Activity Index-2000 (SLEDAI-2K) cut-off of at least 6. In some embodiments, highly active SLE is defined by a SLEDAI-2K cut-off of at least 7. In some embodiments, highly active SLE is defined by a SLEDAI-2K cut-off of at least 8. In some embodiments, highly active SLE is defined by a SLEDAI-2K cut-off of at least 9. In some embodiments, highly active SLE is defined by a SLEDAI-2K cut-off of at least 10. In some embodiments, highly active SLE is defined by a SLEDAI-2K cut-off of at least 11. In some embodiments, highly active SLE is defined by a SLEDAI-2K cut-off of at least 12. As used herein, when calculating a SLEDAI-2K score, points for alopecia, headache, and fever are excluded because of the frequently nonspecific nature of these signs or symptoms and the difficulty in confirming that these features are attributable to active SLE rather than another cause. PGA is also recognized in the art as an efficient assessment tool for SLE. See, e.g., Louthrenoo W, et al. Lupus. 2021, vol. 30(10): 1586-1595, incorporated by reference herein; see also, Example 7, infra. CLASI is a validated instrument for assessing cutaneous lupus erythematosus activity. See, e.g., Albrecht, J, et al., J Invest Dermatol. 2005, vol. 125(5): 889-894; see also, Example 7, infra. Tender / swollen joint counts is a further SLE clinical disease activity assessment. See, e.g., Cipriano E et al., Reumatismo. 2015, vol. 67(2):62-7; see also, Example 7, infra.

[0081] In some embodiments of any of the above aspects, the subject suffers from lupus nephritis. In some embodiments, the lupus nephritis is active Class III or Class IV disease. In some embodiments, the lupus nephritis is active Class III disease. In some embodiments, the lupus nephritis is active Class IV disease. Class III and Class IV lupus nephritis typically require high-dose immunosuppressants for management and carry an increased risk for progression to end-stage renal disease. In someAttorney Docket No.: 000218-0093-W01embodiments of any of the above aspects, the subject does not suffer from lupus nephritis.

[0082] In some embodiments of any of the above aspects, the CAR-T cell is administered to the subject intravenously or is formulated to be administered to the subject intravenously. In some embodiments, the CAR-T cell is administered to the subject intravenously. In some embodiments, the CAR-T cell is formulated to be administered to the subject intravenously. In some embodiments of any of the above aspects, the CAR-T cell is administered as a single dose or is formulated to be administered as a single dose. In some embodiments, the CAR-T cell is administered as a single dose. In some embodiments, the CAR-T cell is formulated to be administered as a single dose. In some embodiments, one or more subsequent doses of the CAR-T cell is administered to the subject. In some embodiments, the one or more subsequent doses of the CAR-T cell are administered to the subject every 3 to 5 years after the first dose. For example, the second dose is administered 3 to 5 years after the first dose, the third dose is administered 3 to 5 years after the second dose, and so on. In some embodiments, the one or more subsequent doses of the CAR-T cell are administered to the subject every 3 years after the first dose. In some embodiments, the one or more subsequent doses of the CAR-T cell are administered to the subject every 3.5 years after the first dose. In some embodiments, the one or more subsequent doses of the CAR-T cell are administered to the subject every 4 years after the first dose. In some embodiments, the one or more subsequent doses of the CAR-T cell are administered to the subject every 4.5 years after the first dose. In some embodiments, the one or more subsequent doses of the CAR-T cell are administered to the subject every 5 years after the first dose.

[0083] In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 5 x 106and about 1.2 x 109cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 20 x 106and about 1.2 x 109cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 20 x 106and about 800 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 20 x 106and about 480 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a doseAttorney Docket No.: 000218-0093-W01between about 20 x 106and about 320 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 20 x 106and about 160 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 20 x 106and about 60 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 60 x 106and about 1.2 x 109cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 60 x 106and about 800 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 60 x 106and about 480 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 160 x 106and about 1.2 x 109cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 160 x 106and about 800 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 160 x 106and about 480 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 320 x 106and about 1.2 x 109cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 320 x 106and about 800 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 320 x 106and about 480 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 60 x 106and about 320 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 60 x 106and about 160 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 160 x 106and about 320 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 480 x 106and about 1.2 x 109cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between aboutAttorney Docket No.: 000218-0093-W01480 x 106and about 800 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose of about 5 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose of about 20 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose of about 60 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose of about 160 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose of about 320 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose of about 480 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose of about 800 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose of about 1.2 x 109cells.

[0084] In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 5 x 106and 1.2 x 109cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 20 x 106and 1.2 x 109cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 20 x 106and 800 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 20 x 106and 480 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 20 x 106and 320 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 20 x 106and 160 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 20 x 106and 60 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 60 x 106and 1.2 x 109cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 60 x 106and 800 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or isAttorney Docket No.: 000218-0093-W01formulated to be administered at a dose between 60 x 106and 480 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 160 x 106and 1.2 x 109cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 160 x 106and 800 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 160 x 106and 480 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 320 x 106and 1.2 x 109cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 320 x 106and 800 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 320 x 106and 480 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 60 x 106and 320 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 60 x 106and 160 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 160 x 106and 320 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 480 x 106and 1.2 x 109cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 480 x 106and 800 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose of 5 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose of 20 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose of 60 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose of 160 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose of 320 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose of 480 x 106cells. In some embodiments of any of the above aspects, the CAR-T cellAttorney Docket No.: 000218-0093-W01is administered or is formulated to be administered at a dose of 800 x 106cells. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose of 1.2 x 109cells.

[0085] In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 0.0625 x 106and about 15 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 0.0625 x 106and about 10 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 0.0625 x 106and about 6 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 0.0625 x 106and about 2 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 0.0625 x 106and about 0.75 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 0.0625 x 106and about 0.25 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 0.25 x 106and about 15 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 0.75 x 106and about 15 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 2 x 106and about 15 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 6 x 106and about 15 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 10 x 106and about 15 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 0.25 x 106and about 10 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 0.25 x 106and about 6 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 0.25 x 106and about 2 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-Attorney Docket No.: 000218-0093-W01T cell is administered or is formulated to be administered at a dose between about 0.25 x 106and about 0.75 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 0.75 x 106and about 6 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 0.75 x 106and about 2 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between about 2 x 106and about 6 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose of about 0.0625 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose of about 0.25 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose of about 0.75 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose of about 2 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose of about 6 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose of about 10 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose of about 15 x 106cells / kg.

[0086] In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 0.0625 x 106and 15 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 0.0625 x 106and 10 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 0.0625 x 106and 6 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 0.0625 x 106and 2 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 0.0625 x 106and 0.75 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 0.0625 x 106andAttorney Docket No.: 000218-0093-W010.25 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 0.25 x 106and 15 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 0.75 x 106and 15 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 2 x 106and 15 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 6 x 106and 15 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 10 x 106and 15 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 0.25 x 106and 10 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 0.25 x 106and 6 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 0.25 x 106and 2 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 0.25 x 106and 0.75 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 0.75 x 106and 6 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 0.75 x 106and 2 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose between 2 x 106and 6 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose of 0.0625 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose of 0.25 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose of 0.75 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose of 2 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose of 6 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or isAttorney Docket No.: 000218-0093-W01formulated to be administered at a dose of 10 x 106cells / kg. In some embodiments of any of the above aspects, the CAR-T cell is administered or is formulated to be administered at a dose of 15 x 106cells / kg.

[0087] In some embodiments of any of the above aspects, administration of the CAR-T cell results in (a) a detectable number of CAR transgene copies in the blood of the subject, (b) a detectable number CAR-T cells in the blood of the subject, (c) the depletion of B-cell levels in the subject by more than 90 percent, (d) immunosuppression-free remission in the subject for at least 6 months, (e) an time to first SLE flare of at least 6 months; (f) a total number of SLE flares less than one per year; (g) reduced glucocorticoid and / or immunosuppressant use as compared to prior to treatment; (h) reduced titers antinuclear antibody (ANA) and double stranded DNA (dsDNA) as compared to prior to treatment; or (i) reduced titers of lupus-associated autoantibodies (e.g., anti-Smith and anti-RNP) as compared to prior to treatment.

[0088] In some embodiments of any of the above aspects, administration of the CAR-T cell results in a detectable number of CAR transgene copies in the blood of the subject. In some embodiments, the number of CAR transgene copies are measured by droplet digital polymerase chain reaction (ddPCR), which is known in the art to provide accurate and precise detection of vector copy number in CAR T cell products. See, e.g, Murphy LA, et al. Cytotherapy, 2023; vol. 25(l):94-102, incorporated herein by reference in its entirety.

[0089] In some embodiments of any of the above aspects, administration of the CAR-T cell results in a detectable number CAR-T cells in the blood of the subject.

[0090] In some embodiments of any of the above aspects, administration of the CAR-T cell results in the depletion of B-cell levels in the subject by more than 90 percent. In some embodiment, the level of B-cells in the subject is depleted by more than 95 percent. In some embodiment, the level of B-cells in the subject is depleted by more than 97 percent. In some embodiment, the level of B-cells in the subject is depleted by more than 99 percent. In some embodiments, after administration of the CAR-T cells, the level of B-cells in the subject falls below the limit of detection by flow cytometry. In some embodiments, depletion of the number of B-cells occurs within 1 month after administration of the CAR-T cell. In some embodiments, depletion of the number of B-cells occurs within 2 months after administration of the CAR-T cell. In some embodiments, depletion of the number of B-cells occurs within 3 months after administration of the CAR-T cell. In some embodiments, depletion ofAttorney Docket No.: 000218-0093-W01the number of B-cells occurs within 4 months after administration of the CAR-T cell. In some embodiments, depletion of the number of B-cells occurs within 5 months after administration of the CAR-T cell. In some embodiments, depletion of the number of B-cells occurs within 6 months after administration of the CAR-T cell. In some embodiments, depletion of the number of B-cells occurs between 1 and 6 months after administration of the CAR-T cell. In some embodiments, depletion of the number of B-cells occurs between 2 and 6 months after administration of the CAR-T cell. In some embodiments, depletion of the number of B-cells occurs between 3 and 6 months after administration of the CAR-T cell. In some embodiments, depletion of the number of B-cells occurs between 4 and 6 months after administration of the CAR-T cell. In some embodiments, depletion of the number of B-cells occurs between 5 and 6 months after administration of the CAR-T cell. In some embodiments, depletion of the number of B-cells occurs between 1 and 2 months after administration of the CAR-T cell. In some embodiments, depletion of the number of B-cells occurs between 1 and 3 months after administration of the CAR-T cell. In some embodiments, depletion of the number of B-cells occurs between 1 and 4 months after administration of the CAR-T cell. In some embodiments, depletion of the number of B-cells occurs between 1 and 5 months after administration of the CAR-T cell. In some embodiments, depletion of the number of B-cells occurs between 2 and 5 months after administration of the CAR-T cell. In some embodiments, depletion of the number of B-cells occurs between 2 and 4 months after administration of the CAR-T cell. In some embodiments, depletion of the number of B-cells occurs between 2 and 3 months after administration of the CAR-T cell. In some embodiments, depletion of the number of B-cells occurs between 3 and 5 months after administration of the CAR-T cell. In some embodiments, depletion of the number of B-cells occurs between 3 and 4 months after administration of the CAR-T cell. In some embodiments, depletion of the number of B-cells occurs between 4 and 5 months after administration of the CAR-T cell.

[0091] Without being bound by theory, the ability of P-CD19CD20-ALLO1 CAR-T cells to provide deep depletion of tissue resident B cells that are refractory to other B cell-depleting modalities may facilitate an “immune reset” where the pathogenic B cell repertoire is eliminated, and a predominantly naive immune system is reconstituted. In some embodiments, a naive B-cell population is reconstituted in the subject after administration of the CAR-T cell. In some embodiments, a naive B-cell population is reconstituted in the subject about 6 months after administration of theAttorney Docket No.: 000218-0093-W01CAR-T cell. In some embodiments, a naive B-cell population is reconstituted in the subject about 7 months after administration of the CAR-T cell. In some embodiments, a naive B-cell population is reconstituted in the subject about 8 months after administration of the CAR-T cell. In some embodiments, a naive B-cell population is reconstituted in the subject about 9 months after administration of the CAR-T cell. In some embodiments, a naive B-cell population is reconstituted in the subject about 10 months after administration of the CAR-T cell. In some embodiments, a naive B-cell population is reconstituted in the subject about 11 months after administration of the CAR-T cell. In some embodiments, a naive B-cell population is reconstituted in the subject about 12 months after administration of the CAR-T cell. In some embodiments, a naive B-cell population is reconstituted in the subject between 6 and 12 months after administration of the CAR-T cell. In some embodiments, a naive B-cell population is reconstituted in the subject between 6 and 11 months after administration of the CAR-T cell. In some embodiments, a naive B-cell population is reconstituted in the subject between 6 and 10 months after administration of the CAR-T cell. In some embodiments, a naive B-cell population is reconstituted in the subject between 6 and 9 months after administration of the CAR-T cell. In some embodiments, a naive B-cell population is reconstituted in the subject between 6 and 8 months after administration of the CAR-T cell. In some embodiments, a naive B-cell population is reconstituted in the subject between 6 and 7 months after administration of the CAR-T cell. In some embodiments, a naive B-cell population is reconstituted in the subject between 7 and 12 months after administration of the CAR-T cell. In some embodiments, a naive B-cell population is reconstituted in the subject between 7 and 11 months after administration of the CAR-T cell. In some embodiments, a naive B-cell population is reconstituted in the subject between 7 and 10 months after administration of the CAR-T cell. In some embodiments, a naive B-cell population is reconstituted in the subject between 7 and 9 months after administration of the CAR-T cell. In some embodiments, a naive B-cell population is reconstituted in the subject between 7 and 8 months after administration of the CAR-T cell. In some embodiments, a naive B-cell population is reconstituted in the subject between 8 and 12 months after administration of the CAR-T cell. In some embodiments, a naive B-cell population is reconstituted in the subject between 8 and 11 months after administration of the CAR-T cell. In some embodiments, a naive B-cell population is reconstituted in the subject between 8 and 10 months after administration of the CAR-T cell. In some embodiments, a naive B-Attorney Docket No.: 000218-0093-W01cell population is reconstituted in the subject between 8 and 9 months after administration of the CAR-T cell. In some embodiments, a naive B-cell population is reconstituted in the subject between 9 and 12 months after administration of the CAR-T cell. In some embodiments, a naive B-cell population is reconstituted in the subject between 9 and 11 months after administration of the CAR-T cell. In some embodiments, a naive B-cell population is reconstituted in the subject between 9 and 10 months after administration of the CAR-T cell. In some embodiments, a naive B-cell population is reconstituted in the subject between 10 and 12 months after administration of the CAR-T cell. In some embodiments, a naive B-cell population is reconstituted in the subject between 10 and 11 months after administration of the CAR-T cell. In some embodiments, a naive B-cell population is reconstituted in the subject between 11 and 12 months after administration of the CAR-T cell.

[0092] In some embodiments of any of the above aspects, administration of the CAR-T cell results in immunosuppression-free remission in the subject for at least 6 months. In some embodiments, the immunosuppression-free remission in the subject for at least 9 months. In some embodiments, the immunosuppression-free remission in the subject for at least 12 months. In some embodiments, the immunosuppression-free remission in the subject for at least 15 months. In some embodiments, the immunosuppression-free remission in the subject for at least 18 months. In some embodiments, the immunosuppression-free remission in the subject for at least 24 months.

[0093] In some embodiments of any of the above aspects, administration of the CAR-T cell results in a time to first SLE flare of at least 6 months. In some embodiments, the time to first SLE flare is at least 9 months. In some embodiments, the time to first SLE flare is at least 12 months. In some embodiments, the time to first SLE flare is at least 15 months. In some embodiments, the time to first SLE flare is at least 18 months. In some embodiments, the time to first SLE flare is at least 24 months.

[0094] In some embodiments of any of the above aspects, administration of the CAR-T cell results in a total number of SLE flares less than one per year. In some embodiments of any of the above aspects, administration of the CAR-T cell results in reduced glucocorticoid and / or immunosuppressant use by the subject as compared to prior to treatment. In some embodiments of any of the above aspects, administration of the CAR-T cell results in reduced glucocorticoid use by the subject as compared to prior to treatment. In some embodiments of any of the above aspects, administrationAttorney Docket No.: 000218-0093-W01of the CAR-T cell results in reduced immunosuppressant use by the subject as compared to prior to treatment. In some embodiments of any of the above aspects, administration of the CAR-T cell results in reduced glucocorticoid and immunosuppressant use as compared to prior to treatment. In some embodiments of any of the above aspects, administration of the CAR-T cell results in reduced titers antinuclear antibody (ANA) and anti-double stranded DNA (dsDNA) antibodies in the subject as compared to prior to treatment. In some embodiments of any of the above aspects, administration of the CAR-T cell results in reduced titers antinuclear antibody (ANA) in the subject as compared to prior to treatment. In some embodiments of any of the above aspects, administration of the CAR-T cell results in reduced titers anti-dsDNA antibodies in the subject as compared to prior to treatment. In some embodiments of any of the above aspects, administration of the CAR-T cell results in reduced titers of lupus-associated autoantibodies in the subject as compared to prior to treatment. In some embodiments, the lupus-associated autoantibodies comprise one or more of anti-Smith antibodies, anti-Ul ribonucleoprotein antibodies ( / .< ., anti-RNP antibodies), anti-Sjbgren’s-syndrome-related antigen A antibodies (SSA / Ro antibodies), and anti-Sjbgren’s-syndrome-related antigen B (SSB / La antibodies). In some embodiments, the lupus-associated autoantibodies comprise anti-Smith antibodies. In some embodiments, the lupus-associated autoantibodies comprise anti-RNP antibodies. In some embodiments, the lupus-associated autoantibodies comprise SSA / Ro antibodies. In some embodiments, the lupus-associated autoantibodies comprise SSB / La antibodies.

[0095] In some embodiments of any of the above aspects, administration of the CAR-T cell results in (a) an incidence of Grade 2 cytokine release syndrome (CRS) less than 30 percent, (b) an incidence of immune effector cell-associated neurotoxicity syndrome (ICANS) less than 20 percent, (c) an incidence of hemophagocytic lymphohistiocytosis (HLH) less than 10 percent, (d) a risk of serious infection no more than 10% more compared to the incidence of serious infection in patients receiving standard of care at 6 months after treatment, (e) an incidence of immune rejection and graft-versus-host disease (GVHD) less than 10 percent, or (f) an incidence of new T-cell malignancies less than 0.1%.

[0096] In some embodiments of any of the above aspects, administration of the CAR-T cell results in an incidence of Grade 2 cytokine release syndrome (CRS) less than 30 percent. In some embodiments, the incidence if Grade 2 CRS is less than 25Attorney Docket No.: 000218-0093-W01percent. In some embodiments, the incidence if Grade 2 CRS is less than 20 percent. In some embodiments, the incidence if Grade 2 CRS is less than 15 percent. In some embodiments, the incidence if Grade 2 CRS is less than 10 percent. In some embodiments, the incidence if Grade 2 CRS is less than 5 percent. As used herein, CRS is assessed according to the American Society for Transplantation and Cellular Therapy (ASTCT) Consensus CRS Grading. See, e.g., Lee, DW et al., Biol Blood Marrow Transplant. 2019 Apr;25(4):625-638, incorporated herein by reference in its entirety.

[0097] In some embodiments of any of the above aspects, administration of the CAR-T cell results in an incidence of immune effector cell-associated neurotoxicity syndrome (ICANS) less than 20 percent. In some embodiments, the incidence of ICANS is less than 15 percent. In some embodiments, the incidence of ICANS is less than 10 percent. In some embodiments, the incidence of ICANS is less than 5 percent. As used herein, ICANS is assessed according to ASTCT Consensus ICANS Grading. See, e.g., Lee, DW et al., Biol Blood Marrow Transplant. 2019 Apr;25(4):625-638, incorporated herein by reference in its entirety.

[0098] In some embodiments of any of the above aspects, administration of the CAR-T cell results in an incidence of hemophagocytic lymphohistiocytosis (HLH) less than 10 percent. In some embodiments, the incidence of HLH is between 5 and 10 percent. In some embodiments, the incidence of HLH is less than 5 percent. As used herein, HLH is assessed according to ASTCT Consensus HLH Grading. See, e.g., Hines, MR. et al., Transplantation and Cellular Therapy, 2023, vol. 29(7): 438.el-438. el 6, incorporated herein by reference in its entirety.

[0099] In some embodiments of any of the above aspects, administration of the CAR-T cell results in a risk of serious infection no more than 10% more compared to the incidence of serious infection in patients receiving standard of care at 6 months after treatment. In some embodiments, the risk of serious infection no more than 5% more compared to the incidence of serious infection in patients receiving standard of care at 6 months after treatment. In some embodiments, the risk of serious infection is similar to the incidence of serious infection in patients receiving standard of care at 6 months after treatment. The incidence of serious infection in patients receiving standard of care is known in the art. See, e.g., Rodziewicz, M et al. The Lancet Rheumatology, 2023, Volume 5, Issue 5, e284 - e292, incorporated herein by reference in its entirety.

[0100] In some embodiments of any of the above aspects, administration of the CAR-T cell results in an incidence of immune rejection and graft-versus-host diseaseAttorney Docket No.: 000218-0093-W01(GVHD) less than 10 percent. As used herein, GVHD will be assessed according to the International Bone Marrow Transplant Registry (IBMTR) criteria. See, e.g., Rowlings, PA. et al. British J. of Haematology, 1997, vol. 97(4): 855-864, incorporated herein by reference in its entirety.

[0101] In some embodiments of any of the above aspects, administration of the CAR-T cell results in an incidence of new T-cell malignancies less than 0.1%.

[0102] In some embodiments of any of the above aspects, the subject has not been previously administered a therapeutic agent to treat SLE. In some embodiments of any of the above aspects, the subject has been previously administered a therapeutic agent to treat SLE. In some embodiments, the subject is refractory to the previously administered therapeutic agent. In some embodiments, the previously administered therapeutic agent is selected from a corticosteroid, an antimalarial therapy, an immunosuppressant, a B-cell depleting therapy, and a cytokine blocker. In some embodiments, the previously administered therapeutic agent is a corticosteroid. In some embodiments, the previously administered therapeutic agent is an antimalarial therapy. In some embodiments, the antimalarial therapy is hydroxychloroquine. In some embodiments, the previously administered therapeutic agent is an immunosuppressant. In some embodiments, the immunosuppressant is selected from the group consisting of cyclophosphamide, azathioprine, mycophenolate mofetil (MMF), mycophenolic acid, and methotrexate. In some embodiments, the previously administered therapeutic agent is a B-cell depleting therapy. In some embodiments, the B-cell depleting therapy is selected from the group consisting of rituximab, ocrelizumab, ofatumumab, belimumab and obinutuzumab. In some embodiments, the cytokine blocker is anifrolumab. In some embodiments, the previously administered therapeutic agent is a cytokine blocker.

[0103] In some embodiments of any of the above aspects, the CAR-T cell is administered or formulated to be administered in combination with one or more additional therapeutic agents. In some embodiments, the CAR-T cell is administered or formulated to be administered sequentially with the one or more additional therapeutic agents. In some embodiments, the CAR-T cell is administered or formulated to be administered simultaneously with the one or more additional therapeutic agents. In some embodiments, the one or more additional therapeutic agents is selected from an immunosuppressant, a B-cell depleting therapy, a calcineurin inhibitor, another biologic or targeted therapy, an IL-1 inhibitor, a corticosteroid, aAttorney Docket No.: 000218-0093-W01kinase inhibitor, a cytokine blocker or a combination thereof. In some embodiments, the one or more additional therapeutic agents is an immunosuppressant. In some embodiments, the immunosuppressant is selected from the group consisting of cyclophosphamide, azathioprine, mycophenolate mofetil (MMF), mycophenolic acid, and methotrexate. In some embodiments, the one or more additional therapeutic agents is a B-cell depleting therapy. In some embodiments, the B-cell depleting therapy is selected from the group consisting of rituximab, ocrelizumab, ofatumumab, belimumab and obinutuzumab. In some embodiments, the one or more additional therapeutic agents is a calcineurin inhibitor. In some embodiments, the calcineurin inhibitor is selected from the group consisting of ciclosporin, tacrolimus, and voclosporin. In some embodiments, the one or more additional therapeutic agents is another biologic or targeted therapy. In some embodiments, the one or more additional therapeutic agents is an IL-1 inhibitor. In some embodiments, the one or more additional therapeutic agents is a corticosteroid. In some embodiments, the one or more additional therapeutic agents is a kinase inhibitor. In some embodiments, the kinase inhibitor is ruxolitinib. In some embodiments, the one or more additional therapeutic agents is a cytokine blocker. In some embodiments, the cytokine blocker is selected from the group consisting of ruxolitinib, etoposide, or emapalumab.

[0104] In some embodiments of any of the above aspects, the subject has been treated with a lymphodepleting chemotherapy (LDC) regimen prior to the administration of CAR-T cell. In some embodiments, the LDC regimen comprises cyclophosphamide (300-750 mg / m2IV) and fludarabine (0-30 mg / m2IV) for 3 consecutive days. In some embodiments, the LDC regimen comprises cyclophosphamide (300-750 mg / m2IV) and fludarabine (30 mg / m2IV) for 3 consecutive days. In some embodiments, the LDC regimen comprises cyclophosphamide (500 mg / m2IV) and fludarabine (30 mg / m2IV) for 3 consecutive days. In some embodiments, the LDC regimen comprises cyclophosphamide (300-750 mg / m2IV) for 3 consecutive days. In some embodiments, the LDC regimen is administered on Days -5, -4, and -3, and the CAR-T cells is administered or to be administered on Day 1.Attorney Docket No.: 000218-0093-W01Articles of Manufacture

[0105] A fourth aspect of the present disclosure provides a dose infusion bag comprising a composition comprising P-CD19CD20-ALLO1 CAR-T cells. In some embodiments, the dose infusion bag is configured from intravenous administration.

[0106] In some embodiments, the composition comprises from about 5 x 106and about 1.2 x 109cells. In some embodiments, the composition comprises from about 20 x 106and about 1.2 x 109cells. In some embodiments, the composition comprises from about 20 x 106and about 800 x 106cells. In some embodiments, the composition comprises from about 20 x 106and about 480 x 106cells. In some embodiments, the composition comprises from about 20 x l06and about 320 x lO6cells. In some embodiments, the composition comprises from about 20 x 106and about 160 x 106cells. In some embodiments, the composition comprises from about 20 x 106and about 60 x 106cells. In some embodiments, the composition comprises from about 60 x 106and about 1.2 x 109cells. In some embodiments, the composition comprises from about 60 x 106and about 800 x 106cells. In some embodiments, the composition comprises from about 60 x 106and about 480 x 106cells. In some embodiments, the composition comprises from about 160 x 106and about 1.2 x 109cells. In some embodiments, the composition comprises from about 160 x 106and about 800 x 106cells. In some embodiments, the composition comprises from about 160 x 106and about 480 x 106cells. In some embodiments, the composition comprises from about 320 x 106and about 1.2 x 109cells. In some embodiments, the composition comprises from about 320 x 106and about 800 x 106cells. In some embodiments, the composition comprises from about 320 x 106and about 480 x 106cells. In some embodiments, the composition comprises from about 60 x 106and about 320 x 106cells. In some embodiments, the composition comprises from about 60 x l06and about 160 x lO6cells. In some embodiments, the composition comprises from about 160 x 106and about 320 x 106cells. In some embodiments, the composition comprises from about 480 x 106and about 1.2 x 109cells. In some embodiments, the composition comprises from about 480 x 106and about 800 x 106cells. In some embodiments, the composition comprises about 5 x 106cells. In some embodiments, the composition comprises about 20 x 106cells. In some embodiments, the composition comprises about 60 x 106cells. In some embodiments, the composition comprises about 160 x 106cells. In some embodiments, the composition comprises about 320 x lO6cells. In some embodiments, the composition comprises about 480 x 106cells. In some embodiments, the compositionAttorney Docket No.: 000218-0093-W01comprises about 800 x 106cells. In some embodiments, the composition comprises about 1.2 x 109cells.

[0107] In some embodiments, the composition comprises from 5 x 106and 1.2 x 109cells. In some embodiments, the composition comprises from 20 x 106and 1.2 x 109cells. In some embodiments, the composition comprises from 20 x 106and 800 x 106cells. In some embodiments, the composition comprises from 20 x 106and 480 x 106cells. In some embodiments, the composition comprises from 20 x 106and 320 x 106cells. In some embodiments, the composition comprises from 20 x 106and 160 x 106cells. In some embodiments, the composition comprises from 20 x 106and 60 x 106cells. In some embodiments, the composition comprises from 60 x 106and 1.2 x 109cells. In some embodiments, the composition comprises from 60 x 106and 800 x 106cells. In some embodiments, the composition comprises from 60 x 106and 480 x 106cells. In some embodiments, the composition comprises from 160 x 106and 1.2 x 109cells. In some embodiments, the composition comprises from 160 x 106and 800 x 106cells. In some embodiments, the composition comprises from 160 x 106and 480 x 106cells. In some embodiments, the composition comprises from 320 x 106and 1.2 x 109cells. In some embodiments, the composition comprises from 320 x 106and 800 x 106cells. In some embodiments, the composition comprises from 320 x 106and 480 x 106cells. In some embodiments, the composition comprises from 60 x 106and 320 x 106cells. In some embodiments, the composition comprises from 60 x 106and 160 x 106cells. In some embodiments, the composition comprises from 160 x 106and 320 x 106cells. In some embodiments, the composition comprises from 480 x 106and 1.2 x 109cells. In some embodiments, the composition comprises from 480 x 106and 800 x 106cells. In some embodiments, the composition comprises 5 x 106cells. In some embodiments, the composition comprises 20 x 106cells. In some embodiments, the composition comprises 60 x 106cells. In some embodiments, the composition comprises 160 x 106cells. In some embodiments, the composition comprises 320 x 106cells. In some embodiments, the composition comprises 480 x 106cells. In some embodiments, the composition comprises 800 x 106cells. In some embodiments, the composition comprises 1.2 x 109cells.EXEMPLARY EMBODIMENTS

[0108] Particular embodiments of the disclosure are set forth in the following numbered paragraphs:Attorney Docket No.: 000218-0093-W011. A method of treating systemic lupus erythematosus (SLE) in a human subject in need thereof comprising administering a P-CD19CD20-ALLO1 chimeric antigen receptor T cell (CAR-T cell) to the subject.2. Use of a P-CD19CD20-ALLO1 chimeric antigen receptor T cell (CAR-T cell) in the manufacture of a medicament for treating systemic lupus erythematosus (SLE) in a human subj ect in need thereof.3. A P-CD19CD20-ALLO1 chimeric antigen receptor T cell (CAR-T cell) for use in treating systemic lupus erythematosus (SLE) in a human subject in need thereof.4. The method according to paragraph 1, the use according to paragraph 2, or the CAR-T cell for use according to paragraph 3, wherein the systemic lupus erythematosus (SLE) is severe SLE; treatment refractory SLE; highly active SLE; severe, treatment-refractory SLE; severe, highly active SLE; highly active, treatmentrefractory SLE; or highly active, severe, treatment-refractory SLE.5. The method, use or CAR-T cell for use of paragraph 4, wherein the highly active SLE is identified by (a) British Isles Lupus Assessment Group (BILAG)-2004 Category A disease in one or more organ systems and / or Category B disease in two or more organ systems (excluding constitutional, musculoskeletal, and / or mucocutaneous organ systems for Category B); (b) a Systemic Lupus Erythematosus Disease Activity Index 2000 (SLEDAI-2K) score greater than or equal to 8; and (c) a Physician’s Global Assessment (PGA) score greater than or equal to 1.0 on a 0 to 3 Visual Analog Scale (VAS).6. The method, use or CAR-T cell for use of paragraph 4 or 5, wherein the subject suffers from lupus nephritis.7. The method, use or CAR-T cell for use of paragraph 4 or 5, wherein the subject does not suffer from lupus nephritis.8. The method according to any one of paragraphs 1 and 4-7, the use according to any one of paragraphs 2 and 4-7, or the CAR-T cell for use according to any one of paragraphs 3-7, wherein the CAR-T cell is administered to the subject intravenously or is formulated to be administered to the subject intravenously.9. The method according to any one of paragraphs 1 and 4-8, the use according to any one of paragraphs 2 and 4-8, or the CAR-T cell for use according to any one of paragraphs 3-8, wherein the CAR-T cell is administered as a single dose or is formulated to be administered as a single dose.Attorney Docket No.: 000218-0093-W0110. The method according to any one of paragraphs 1 and 4-8, the use according to any one of paragraphs 2 and 4-8, or the CAR-T cell for use according to any one of paragraphs 3-8, wherein the CAR-T cell is administered in multiple doses or is formulated to be administered in multiple doses, optionally wherein each of the multiple doses is administered between 3 and 5 years after the previous dose.11. The method according to any one of paragraphs 1 and 4-10 the use according to any one of paragraphs 2 and 4-10, or the CAR-T cell for use according to any one of paragraphs 3-10, wherein the CAR-T cell is administered at a dose of about 5 x 106cells to about 1.2 x 109cells or is formulated to be administered at a dose of about 5 x 106to about 1.2 x 109cells.12. The method according to any one of paragraphs 1 and 4-11, the use according to any one of paragraphs 2 and 4-11, or the CAR-T cell for use according to any one of paragraphs 3-11, wherein administration of the CAR-T cell results in (a) a detectable number of CAR transgene copies in the blood of the subject, (b) a detectable number of CAR-T cells in the blood of the subject, (c) the depletion of B-cell levels in the subject by at least 90% percent, (d) immunosuppression-free remission in the subject for at least 6 months, (e) a time to first SLE flare of at least 6 months; (f) a total number of SLE flares less than one per year (g) reduced glucocorticoid and / or immunosuppressant use as compared to prior to treatment; (h) reduced titers antinuclear antibody (ANA) and double stranded DNA (dsDNA) as compared to prior to treatment; or (i) reduced titers of lupus-associated autoantibodies (e.g., anti-Smith and anti-RNP) as compared to prior to treatment.13. The method according to any one of paragraphs 1 and 4-12, the use according to any one of paragraphs 2 and 4-12, or the CAR-T cell for use according to any one of paragraphs 3-12, wherein administration of the CAR-T cell results in (a) an incidence of Grade 2 cytokine release syndrome (CRS) less than 30%, (b) an incidence of immune effector cell-associated neurotoxicity syndrome (ICANS) less than 20%, (c) an incidence of hemophagocytic lymphohistiocytosis (HLH) less than 10%, (d) a risk of serious infection no more than 10% more compared to the incidence of infection resulting from standard of care at 6 months after treatment, (e) an incidence of immune rejection and graft-versus-host disease (GVHD) less than 10%, or (f) an incidence of new T-cell malignancies less than 0.1%.14. The method according to any one of paragraphs 1 and 4-13, the use according to any one of paragraphs 2 and 4-13, or the CAR-T cell for use according toAttorney Docket No.: 000218-0093-W01any one of paragraphs 3-13, wherein the subject has not been previously administered a therapeutic agent to treat SLE.15. The method according to any one of paragraphs 1 and 4-13, the use according to any one of paragraphs 2 and 4-13, or the CAR-T cell for use according to any one of paragraphs 3-13, wherein the subject has been previously administered a therapeutic agent to treat SLE.16. The method, use, or CAR-T cell for use according to paragraph 15, wherein the subject is refractory to the previously administered therapeutic agent.17. The method, use, or CAR-T cell for use according to paragraph 15 or 16, wherein the previously administered therapeutic agent is selected from a corticosteroid, an antimalarial therapy, an immunosuppressant, a B-cell depleting therapy, and a cytokine blocker.18. The method, use, or CAR-T cell for use according to paragraph 17, wherein the antimalarial therapy is hydroxychloroquine.19. The method, use, or CAR-T cell for use according to paragraph 17, wherein the immunosuppressant is selected from the group consisting of cyclophosphamide, azathioprine, mycophenolate mofetil (MMF), mycophenolic acid, and methotrexate.20. The method, use, or CAR-T cell for use according to paragraph 17, wherein the B-cell depleting therapy is selected from the group consisting of rituximab, ocrelizumab, ofatumumab, belimumab and obinutuzumab.21. The method, use, or CAR-T cell for use according to paragraph 17, wherein the cytokine blocker is anifrolumab.22. The method according to any one of paragraphs 1 and 4-21, the use according to any one of paragraphs 2 and 4-21, or the CAR-T cell for use according to any one of paragraphs 3-21, wherein the CAR-T cell is administered or formulated to be administered in combination with one or more additional therapeutic agents.23. The method, use, or CAR-T cell for use according to paragraph 22, wherein the CAR-T cell is administered or formulated to be administered sequentially with the one or more additional therapeutic agents.24. The method, use, or CAR-T cell for use according to paragraph 22, wherein the CAR-T cell is administered or formulated to be administered simultaneously with the one or more additional therapeutic agents.Attorney Docket No.: 000218-0093-W0125. The method, use, or CAR-T cell for use according to any one of paragraphs 22-24, wherein the one or more additional therapeutic agents is selected from an immunosuppressant, a B-cell depleting therapy, a calcineurin inhibitor, another biologic or targeted therapy, an IL-1 inhibitor, a corticosteroid, a kinase inhibitor, or a cytokine blocker.26. The method, use, or CAR-T cell for use according to paragraph 25, wherein the immunosuppressant is selected from the group consisting of cyclophosphamide, azathioprine, mycophenolate mofetil (MMF), mycophenolic acid, and methotrexate.27. The method, use, or CAR-T cell for use according to paragraph 25, wherein the B-cell depleting therapy is selected from the group consisting of rituximab, ocrelizumab, ofatumumab, belimumab and obinutuzumab.28. The method, use, or CAR-T cell for use according to paragraph 25, wherein the calcineurin inhibitor is selected from the group consisting of ciclosporin, tacrolimus, and voclosporin.29. The method, use, or CAR-T cell for use according to paragraph 25, wherein the kinase inhibitor is ruxolitinib.30. The method, use, or CAR-T cell for use according to paragraph 25, wherein the cytokine blocker is selected from the group consisting of ruxolitinib, etoposide, or emapalumab.31. The method according to any one of paragraphs 1 and 4-30, the use according to any one of paragraphs 2 and 4-30, or the CAR-T cell for use according to any one of paragraphs 3-30, wherein the subject receives a lymphodepleting chemotherapy (LDC) regimen prior to the administration of CAR-T cell.32. The method, use, or CAR-T cell for use of paragraph 31, wherein the LDC regimen comprises cyclophosphamide (300-750 mg / m2IV) and fludarabine (0-30 mg / m2IV) for 3 consecutive days.33. The method, use, or CAR-T cell for use of paragraph 31 or 32, wherein the LDC regimen is administered on Days -5, -4, and -3, and the CAR-T cells is administered or to be administered on Day 1.Attorney Docket No.: 000218-0093-W01EXAMPLESExample 1: Production of P-CD19CD20-ALLO1 cells

[0109] Healthy donor PBMCs were harvested by a standard leukapheresis and cryostored at < 130°C until use. After thaw, CD4+ and CD8+ T cells were isolated from the leukapheresis material and electroporated with the Cas-CLOVER™ mRNA, the Super piggyBac® (SPB) transposase mRNA, CD2z Booster mRNA, guide RNAs for P2M and TCR P chain, and PB transposon carrying the CD 19 and CD20 transgene. After ex vivo cell expansion during manufacture, followed by purification to remove TCR-positive T cells, modified T cells were formulated and cryopreserved to yield the P-CD19CD20-ALLO1 CAR-T cells.Example 2: In Vitro Primary pharmacology of P-CD19CD20-ALLO1

[0110] Primary pharmacology in vitro studies characterized P-CD19CD20-ALLO1 CAR-T cells for phenotype and target-specific activity. P-CD19CD20-ALLO1 CAR-T cells all had high viability > 91% and demonstrated consistently high knockout frequencies. All lots of P-CD19CD20-ALLO1 CAR-T cells were comprised of 43.5% to 63.9% stem cell-like memory T cells (TSCM) in CD8+ T cells. Table 2, infra, summarizes the cell surface marker expression profile, CD3 and B2M KO frequencies, CAR expression (CD19-CAR and CD20-CAR double-positive of CD4- or CD8-positive cells) and viability of P-CD19CD20-ALLO1 CAR-T products. P-CD19CD20-ALLO1 produced potent, antigen-specific cytotoxicity (Figures 3 A and 3B) and secretion of the proinflammatory cytokine interferon gamma (fFN-y) (Figures 4A and 4B) and granzyme B (Figures 4C and 4D).Attorney Docket No.: 000218-0093-W01Table 2: Summary of Surface Marker Expression and Viability for RUO ProductionsAbbreviations: B2M = beta-2 -microglobulin; CAR = chimeric antigen receptor; CD = cluster of differentiation; KO = knockout; NC-200 = NucleoCounter®-200; TSCM = stem cell memory T cells.Example 3: In Vitro P -CD19CD20 -ALLO1 CAR-T Cell Anti-B Cell Activity Assessment

[0111] The in vitro potency activity of 3 lots of P-CD19CD20-ALLO1 CAR-T cells was evaluated on PBMC’s isolated from patients with SLE. After 48 hours of coculture, all three lots of P-CD19CD20-ALLO1 CAR-T cells consistently demonstrated antigen-specific and robust depletion of B cells (Figure 5). Overall, NK cells and T cells were minimally targeted, although we noted cytotoxic activity against a population of CD20-positive T cells present in some autoimmune disease PBMCs. While monocyte depletion was noted with some PBMC samples, co-culture assays with isolated monocytes did not demonstrate significant cytotoxicity, suggesting the cytotoxicity observed in PBMCs was likely due to bystander killing.

[0112] Similar levels of antigen-specific cytotoxic activity of P-CD19CD20-ALLO1 CAR-T cells were observed in co-culture assays with HD PBMCs or isolated T or B cells (Figures 6A-6C). Cytotoxicity against B cells obtained from healthy donors and patients suffering from autoimmune disease was similar to that observed against the Raji tumor cell line, which expresses both CD 19 and CD20 antigens at a higher level than normal B cells. See, Example 5, infra. The higher background targeting of NK cells, T cells and monocytes that was observed with Donor 3 in PBMCs was not observed against isolated T cells from the same donor indicating that this background is most likely due to bystander killing by activated P-CD19CD20-ALLO1 CAR-T cells rather than specific targeting. In addition to cytotoxic activity, P-CD19CD20-ALLO1 CAR-T cells also demonstrated significant cytokine release against antigen-positive B from both autoimmune patients and healthy donors, but not antigen-negative cells.Attorney Docket No.: 000218-0093-W01

[0113] T cells in autoimmune diseases can express low levels of CD20 and have a high pathogenic potential. P-CD19CD20-ALLO1 T cells induced a strong and E:T-dependent reduction of the number of CD20-positive T cells in an SLE Donor (Figure 7), demonstrating a potential to target both pathogenic B cells and CD20-expressing T cells.Example 4: In Vitro Rimiducid (AP1903) Killing of P-CD19CD20-ALLO1 Cells Expressing Inducible Caspase-9 Protein

[0114] The ability of the P-CD19CD20-ALLO1 iCasp9 safety switch to induce cellular apoptosis in response to administration of rimiducid (AP1903) was evaluated. The iCasp9 gene, an inducible apoptosis system is incorporated into the P-CD19CD20-ALLO1 transgene to eliminate CAR-T cells after administration of another investigational drug, rimiducid (AP1903) in patients, if indicated. Upon binding to API 903, iCasp9 protein dimerizes and triggers the apoptotic caspase cascade, leading to cell death.

[0115] API 903 eliminated P-CD19CD20-ALLO1 cells in a dose-dependent and time-dependent manner in vitro; 0.1, 1 and 10 nM API 903 eliminated approximately 40%, 60%, and 100% of P-CD19CD20-ALLO1 cells by 24 hours, respectively (Figure 8 A), and approximately 40%, 65%, and 100% of P-CD19CD20-ALLO1 cells by 48 hours (Figure 8B). Activated P-CD19CD20-ALLO1 cells were more sensitive to AP1903.Example 5: In Vitro P-CD19CD20-ALLO1 Reactivity Assessment Against Normal Human Cells

[0116] The reactivity of P-CD19CD20-ALLO1 CAR-T cells was evaluated against a panel of normal human cells representative of a broad range of organs and tissue types including 12 normal primary human cells and 8 tumor cell lines along with concurrent assessment of the surface expression of CD 19 and CD20 on target cells by flow cytometry (Figure 9A).

[0117] P-CD19CD20-ALLO1 CAR-T cells specifically lysed the CD19 / CD20 double-positive tumor cell line Raji compared to tumor alone and had no nonspecific killing against the CD19 / CD20 negative cell line Jurkat. Similarly, P-CD19CD20-ALLO1 demonstrated antigen-specific cytotoxicity against all tumor cell lines that expressed CD19 and / or CD20 including against single-positive Raji CD20KO, RajiAttorney Docket No.: 000218-0093-W01CD19KO, Jeko-1, Nalm-6, and Mec-1 cells, and the B Lymphoblast cell line COLO829BL. Of the normal human primary cells, specific cytotoxicity was only observed against bone marrow and PBMCs, both of which had a small fraction of cells that stained positive for CD 19 and CD20 by flow cytometry, as expected (Figure 9B).Example 6: In Vitro Potency Assessment of P-CD19CD20-ALLO1 compared to CD19- and CD20-Single Targeting Counterparts

[0118] The potency of P-CD19CD20-ALLO1 was compared to the potency of its CD 19- and CD20-single targeting counterparts, and potency advantages were observed. In vitro potency was evaluated using serial restimulation against the CD 19-and CD20-positive RAJI cell line, as well as RAJI cells engineered to express only CD 19 or only CD20. In these assays, P-CD19CD20-ALLO1 CAR-T cells showed superior potency against WT (CD19+CD20+) RAJI cells compared to the CD19- or CD20-single targeting products (Figure 11 A). Interestingly, this superiority was also observed against Raji cells expressing only one of the two target antigens, CD 19 or CD20 (Figures 1 IB and 11C), suggesting that the increased potency of P-CD19CD20-ALLO1 is not solely due to the dual binding of CAR molecules to more antigens on the same target cell. Mechanistically, it was observed that over three restimulations with WT (CD19+CD20+) RAJI cells, P-CD19CD20-ALLO1 CAR-T cells expressed and / or sustained higher expression of effector cytokines such as IL-2, TNFa, IFNy, Granzyme A, Granzyme B, perforin and Granulysin than either CD 19 or CD20 single-antigen targeting CAR-T cells (Figures 11D-11J).

[0119] In addition to improved in vitro potency, the in vivo efficacy of the dualtargeting product in comparison with the single-targeting products was assessed in a stress xenograft model of WT (CD19+CD20+) RAJI. P-CD19CD20-ALLO1 CAR-T cells were superior to the CD19-targeting CAR-T product (data not shown). P-CD19CD20-ALLO1, a dual-targeting, fully allogeneic TSCM-rich CAR-T product for CD 19 and CD20-positive B-cell malignancies, demonstrates robust antigen-specific activity against diffuse large B-cell lymphoma (DLBCL) and chronic lymphocytic leukemia (CLL) models and outperforms its single-targeting counterparts even in the presence of only a single antigen on target cells.Attorney Docket No.: 000218-0093-W01Example 7: Open-label, multicenter study of P-CD19CD20-ALLO1 CAR-T Cells

[0120] A Phase I, open-label, non-randomized, multicenter study to evaluate the safety, tolerability, pharmacokinetics (PK), cellular kinetics (CK), pharmacodynamics (PD), and efficacy of P-CD19CD20-ALLO1 CAR-T cells will be conducted in subjects with severe, treatment-refractory systemic lupus erythematosus (SLE) with or without lupus nephritis (LN). The study will include a dose-escalation stage followed by an expansion stage. An overview of the study design, including the dosing plan for each stage, is provided in Figures 12 and 13.

[0121] Up to approximately 132 participants with severe, treatment-refractory SLE will be enrolled in this study, in accordance with the inclusion and exclusion criteria. The study population is carefully selected to include patients with highly active disease, autoantibody positivity, and hypocomplementemia who have failed to achieve adequate disease control with at least two prior therapies. This patient population is at increased risk for major organ involvement, damage accrual, death, and / or complications of treatment (Bruce et al. 2015) and therefore is appropriate for study in the proposed clinical trial.

[0122] For establishing that a sufficient level of disease activity is present for eligibility, high scores in disease activity indices are required. Systemic Lupus Erythematosus Disease Activity Index 2000 (SLEDAI-2K) points for alopecia, headache, and fever are excluded because of the frequently nonspecific nature of these signs or symptoms and the difficulty in confirming that these features are attributable to active SLE rather than another cause (Cruciani et al. 2023). A higher SLEDAI-2K score is associated with increased mortality and damage accrual, while British Isles Lupus Assessment Group (BILAG) disease activity and immunosuppressant use are also linked to increased mortality. Patients with LN must have active Class III or Class IV disease, which requires high-dose immunosuppressants for management and carries an increased risk for progression to end-stage renal disease (Arnaud and Tektonidou 2020).Inclusion Criteria:Signed Informed Consent Form.Age 18-75 years at time of signing Informed Consent Form.Attorney Docket No.: 000218-0093-W01■ Ability to comply with the study protocol, including remaining within an approximately 1-hour travel radius of the treatment site for 28 days after P- CD19CD20-ALLO1 infusion.■ Sufficient proficiency in the language of the investigator and study staff, or ability to communicate with the study staff using a qualified interpreter.■ Agreement to adhere to the contraception requirements .■ Diagnosis of SLE according to the 2019 European League Against Rheumatism (EULAR) / American College of Rheumatology (ACR) Classification Criteria > 24 weeks prior to screening:o Patients with LN are eligible if they meet the following criteria:■ Revised International Society of Nephrology / Renal Pathology Society (ISN / RPS) 2018 Class III or IV active LN (± Class V) demonstrated on renal biopsy performed in the 12 months prior to or during screening.• Active LN is indicated by a modified National Institutes of Health activity index > 1 / 24.• Participants with isolated Class V disease in the absence of Class III or IV LN are not eligible. o Urine protein-to-creatinine ratio (UPCR) > 1 g / g based on a 24-hour collection at screening.Note: A minimum of 15 participants with a diagnosis of LN are planned to be enrolled in each cohort of the expansion stage.■ Antinuclear antibody (ANA) titer > 1 : 80 at screening.■ Anti-dsDNA and / or anti-Sm above the upper limit of normal (ULN) at screening.■ Low C3 or low C4 or low CH50 complement level at screening:o Low C3 is required in the presence of known genetic deficiency of C4.■ Insufficient response, in the investigator’s judgment, to at least two of the following treatments for at least 3 months: Glucocorticoids, cyclophosphamide, mycophenolic acid or its derivatives, belimumab, methotrexate, azathioprine, anifrolumab, rituximab, obinutuzumab, cyclosporin, tacrolimus, orvoclosporin:Attorney Docket No.: 000218-0093-W01o An insufficient response is defined as a lack of sustained response to appropriate doses. Intolerance is not considered insufficient response.■ High disease activity at screening, as defined by all the following:o BILAG-2004 Category A disease in > 1 organ system and / or Category B disease in > 2 organ systems (excluding constitutional, musculoskeletal, and / or mucocutaneous organ systems for Category B) at Screening Visit 1■ Only clinical manifestations and laboratory abnormalities that are present at the time of Screening Visit 1 will be considered.■ Any participant with renal involvement as the organ system resulting in a qualifying BILAG-2004 assessment of disease activity must have Revised ISN / RPS 2018 Class III or IV active LN (± Class V) to be eligible (i.e., Class I or II LN not eligible unless participant meets BILAG-2004 criterion based on other qualifying organ involvement).o SLEDAI-2K score > 8 at Screening Visit 1 and at Screening Visit 3, with additional requirements listed below for determination of eligibility:■ SLEDAI-2K points for alopecia, headache, and fever are excluded.■ Points for arthritis requires > 3 swollen and tender joints, of which > 1 must be in the hands or wrists.■ Points for rash requires a morphology and distribution characteristic of active cutaneous lupus with a CLASI Total Activity Score > 2 excluding points for alopecia and mucosal ulcers.■ Points for mucosal ulcers requires BILAG Category A severe mucosal ulceration, defined as disabling (significantly interfering with oral intake), extensive and deep ulceration, and must have been observed by a physician.Attorney Docket No.: 000218-0093-W01■ Points for hematuria and / or pyuria require the presence of SLEDAI-2K proteinuria. These items should only be scored if the participant has Class III or IV active LN (± Class V).■ Only laboratory abnormalities that are present at the time of Screening Visit 1 will be considered for eligibility assessment at both Screening Visits 1 and 3.■ Physician’ s Global Assessment (PGA) score > 1.0 on a 0 to 3 Visual Analog Scale (VAS) at Screening Visit 1 and at Screening Visit 3.Exclusion Criteria:Individuals who meet any of the following criteria will be excluded from study entry:■ Pregnant or breastfeeding, or intention of becoming pregnant within the timeframe in which contraception is required:o Participants of childbearing potential must have a negative serum pregnancy test result at Screening Visit 1 and a negative urine pregnancy test result at Screening Visit 3 (within 3 days prior to the initiation of LDC [Day -5]) and prior to P-CD19CD20-ALLO1 administration (on Day 1). If Day -5 is > 14 days from the initial screening serum pregnancy test, the serum pregnancy test must be repeated at Screening Visit 3. If a urine pregnancy test is positive, it must be confirmed by a serum pregnancy test.■ Active severe or unstable lupus-associated neuropsychiatric disease or where, in the opinion of the investigator, it is likely to require treatment with protocol- disallowed therapies:o Examples of neuropsychiatric lupus manifestations include, but are not limited to, meningitis, retinitis, cerebral vasculitis, myelopathy, demyelination syndromes, acute confusional state, psychosis, acute stroke or stroke syndrome, cranial neuropathy, status epilepticus or seizures, cerebellar ataxia, and mononeuritis multiplex.■ Active overlap syndrome with mixed connective tissue disease or systemic sclerosis or catastrophic or severe antiphospholipid syndrome within 12 months prior to screening or during screening.Attorney Docket No.: 000218-0093-W01■ History of non-SLE inflammatory skin or joint disease within 12 months of Day 1 that, in the opinion of the investigator, could interfere with assessments of skin or joint manifestations of SLE.■ History of any non-SLE disease treated with oral, IV, or IM corticosteroids > 10 mg / day for more than 14 days in total during the 12 months prior to Day 1.■ High risk for clinically significant bleeding or any condition requiring plasmapheresis, IV immunoglobulin, or acute blood product transfusions. ■ Prior treatment with CAR T-cell therapy, B cell-targeting T cell-dependent bispecific antibody, gene therapy product, total body irradiation, allograft organ transplant, or hematopoietic stem cell transplant.■ Prior history of HLH or macrophage activation syndrome.■ Severe renal impairment, as defined by eGFR < 30 mL / min / 1.73 m2(as estimated using the CKD-EPI 2021 equation) or the need for dialysis or renal transplantation.■ Sclerosis in > 50% of glomeruli on renal biopsy.■ Presence of rapidly progressive glomerulonephritis, defined by any of the following:o Crescent formation in > 50% of glomeruli assessed on renal biopsy. o Sustained doubling of serum creatinine during the 2 months prior to screening.o The investigator’s opinion that the patient has rapidly progressive glomerulonephritis.■ Any of the following laboratory parameters:o AST or ALT > 2.5 x ULNo Amylase or lipase > 2 x ULNo Neutrophils < 1.5 x 103 / pL with one exception:■ Individuals with benign ethnic neutropenia (BEN): ANC < 1.3 x 109 / L (1300 / pL)■ BEN (also known as constitutional neutropenia) is an inherited cause of mild or moderate neutropenia that is not associated with any increased risk for infections or other clinical manifestations (Atallah-Yunes et al. 2019). BEN is referred to as ethnicAttorney Docket No.: 000218-0093-W01neutropenia because of its increased prevalence in people of African descent and other specific ethnic groups.o CD4+ T cells < 250 / mLo Hemoglobin < 9 g / dLo Platelet count < 80,000 / pL■ Baseline oxygen saturation < 92% on room air■ Left ventricular ejection fraction (LVEF) < 45%.■ Has New York Heart Association (NYHA) Class III or IV heart failure, unstable angina, or a history of myocardial infarction or significant arrhythmia (e.g., atrial fibrillation, sustained [> 30 seconds] ventricular tachyarrhythmias, etc.) ■ Significant or uncontrolled medical disease which, in the investigator’ s opinion, would preclude patient participation or that may require treatment with systemic corticosteroids or immunosuppressants during the study.■ Contraindications to mandatory premedications (i.e., acetaminophen and antihistamines) for IRRs or medications used for management of potential risks (corticosteroids), including previous uncontrolled psychosis with corticosteroids or history of closed-angle glaucoma with antihistamines.■ Receipt of any of the following excluded therapies:o Any B cell -depleting (e.g., anti-CD20 or anti-CD19 monoclonal antibodies) or anti-plasma cell therapy such as, but not limited to, obinutuzumab, rituximab, ocrelizumab, ofatumumab, or bortezomib less than 6 months prior to enrollment, except for B cell-targeting T celldependent bispecific antibodies, which are prohibited regardless of timing.o Conventional immunosuppressants (azathioprine, MMF, mycophenolic acid, methotrexate) during the 2 weeks prior to enrollment (treatment of SLE with systemic corticosteroids < 10 mg / day prednisone or equivalent is permitted).o Cyclophosphamide, tacrolimus, ciclosporin, or voclosporin during the 2 weeks prior to enrollmento Any biologic therapy (other than anti-CD20, anti-CD19, or anti-plasma cell) such as, but not limited to, belimumab, ustekinumab, anifrolumab, secukinumab, or atacicept during the 2 months prior to enrollment.Attorney Docket No.: 000218-0093-W01o Inhibitors of Janus-associated kinase (JAK), Bruton’s tyrosine kinase (BTK), or tyrosine kinase 2 (TYK2), including baricitinib, tofacitinib, upadacitinib, filgotinib, ibrutinib, deucravacitinib, or fenebrutinib during the 2 weeks prior to enrollmento Any investigational therapy within 30 days or 5 drug-elimination halflives (whichever is longer) prior to enrollment.o Any live vaccine during the 28 days prior to screeningo IV immunoglobulin (IVIG) or plasmapheresis within 12 weeks of enrollment.■ Known active or latent tuberculosis infection.o If the investigator considers a potential patient to be at an increased risk for infection with Mycobacterium tuberculosis, latent tuberculosis diagnostic procedures must be followed according to local practice standards during the screening period.■ Positive HIV 1 or 2 infection■ Positive HTLV type 1 or type 2 infection.■ Acute or chronic HBV (defined as positive hepatitis B surface antigen [HBsAg] serology [Conners et al. 2023])o Participants who test negative for HBsAg and hepatitis B surface antibody (HBsAb) tests and positive for total hepatitis B core antibody (HBcAb) test are ineligible.■ Active hepatitis C virus (HCV) antibody test at screening, except in individuals who meet either of the following sets of criteria:o Participants with positive HCV antibody test at screening are eligible only if PCR is negative for HCV RNA. The HCV RNA test will be performed only for patients who have a positive HCV antibody test. ■ Positive EBV viral capsid antigen (VCA) IgM testo If EBV VCA IgM test is negative, an EBV PCR test should be performed if acute or chronic active EBV infection is clinically suspected. Participants with a PCR-confirmed EBV infection are excluded from the study.■ Positive human herpes virus (HHV)-6 or HHV-7 infection by PCRo Participants may be included in the study if they are HHV-6 or HHV-7 IgG antibody positive but PCR negative.Attorney Docket No.: 000218-0093-W01■ Any major episode of infection that fulfills any of the following criteria:o Requires hospitalization during 8 weeks prior to screening or during screening.o Requires treatment with IV antibiotics (or anti-infective medications) during 8 weeks prior to screening or during screening.o Requires treatment with oral antibiotics (or anti -infective medications) during 2 weeks prior to screening or during screening.■ Antibiotics or anti-infective medications given in the absence of a major episode of infection are not exclusionary.■ History of serious recurrent or chronic infection■ History of progressive multifocal leukoencephalopathy (PML)■ History of cancer, including solid tumors, hematological malignancies, and carcinoma in situ, within the past 5 yearso Participants with non-melanomatous carcinomas of the skin that have been treated or excised and have resolved are eligible.■ Major surgery requiring hospitalization during 4 weeks prior to screening or during screening or any planned surgery or procedure requiring hospitalization during the 12 weeks following P-CD19CD20-ALLO1 infusion.■ Current alcohol or drug abuse or history of alcohol or drug abuse within 12 months prior to screening or during screeningo Investigators may elect to obtain a urine drug screen if clinically indicated.■ History of severe allergic reactions attributed to compounds of similar chemical or biologic composition to agents used in study (including, but not limited to, cyclophosphamide and fludarabine used in the LDC, DMSO used as a cryoprotectant in the cell media, etc.).

[0123] Potential participants will be screened for study eligibility prior to enrollment. Eligible participants will be enrolled before starting a lymphodepleting chemotherapy (LDC) regimen of cyclophosphamide (300-750 mg / m2 / day) and fludarabine (0-30 mg / m2 / day) administered by IV for 3 days starting on Day -5 (i.e., Days -5, -4, and -3). On Day 1 (3 days after the final day of LDC), participants will receive a single IV infusion of P-CD19CD20-ALLO1 at a starting dose of 60 x 106cells. See Figure 13. Up to four dose levels of P-CD19CD20-ALLO1 will be testedAttorney Docket No.: 000218-0093-W01with the maximum dose not exceeding 480 x 106cells. If de-escalation is required, a dose of 20 x 106cells will be administered.

[0124] Participants will be closely monitored for adverse events throughout the study. Adverse events will be graded according to the CTCAE v5.0 (National Cancer Institute's Common Terminology Criteria for Adverse Events, Version 5.0) grading scale. Cytokine release syndrome (CRS), immune effector cell -associated neurotoxicity syndrome (ICANS), and hemophagocytic lymphohistiocytosis (HLH) will be assessed according to the American Society for Transplantation and Cellular Therapy (ASTCT) criteria, with specific signs and symptoms associated with the adverse event graded according to CTCAE v5.0. GVHD will be assessed according to the International Bone Marrow Transplant (IBMTR) criteria. Rimiducid may be administered as a rescue therapy at a dose of 0.4 mg / kg and infused over approximately 2 hours.

[0125] The primary objective for this study is to evaluate the safety and tolerability of P-CD19CD20-ALLO1 on the basis of the following endpoints:■ Nature and incidence of dose-limiting toxicity (DLT) at each dose level of P- CD19CD20-ALLO1;■ Incidence and severity of adverse events, with severity determined according to the CTCAE v5.0 grading scale, except in cases of cytokine release syndrome (CRS), immune effector cell-associated neurotoxicity syndrome (ICANS), and hemophagocytic lymphohistiocytosis (HLH), which will be assessed according to American Society for Transplantation and Cellular Therapy (ASTCT) criteria, and graft-versus-host disease (GVHD), which will be assessed according to International Bone Marrow Transplant Registry (IBMTR) criteria;■ Incidence of clinically significant abnormal vital signs;■ Incidence of abnormal clinical laboratory test results; and■ Incidence of abnormal ECG parameters.

[0126] The secondary objectives for this study are:■ To characterize the cellular kinetic (CK) profile of P-CD19CD20-ALLO1 on the basis of the number of CAR transgene copies in the blood at specified timepoints by ddPCR;Attorney Docket No.: 000218-0093-W01■ To evaluate the pharmacodynamic effects of P-CD19CD20-ALLO1 on circulating B cells on the basis of B-cell levels in the blood (CD19+, absolute counts in blood) at specified timepoints;■ To evaluate the clinical efficacy of P-CD19CD20-ALLO1 on the basis of the proportion of participants who achieve sustained (from Week 24 through Week 52) drug-free definition of remission in SLE (DORIS) defined as a clinical Systemic Lupus Erythematosus Disease Activity Index 2000 (SLEDAI-2K) = 0 and a Physician’s Global Assessment (PGA) = 0.5 (on a 3-point PGA-VAS), without corticosteroids or other immunosuppressants;■ To evaluate the immunogenicity of P-CD19CD20-ALLO1 on the basis of achieving humoral immunity: Prevalence of anti-CAR-T antibodies at baseline and their incidence during the study; and■ To identify a recommended dose level of P-CD19CD20-ALLO1 for subsequent studies on the basis of the relationship between P-CD19CD20-ALLO1 dose level, safety, and efficacy endpoints and CK parameters for P-CD19CD20- ALLO1.

[0127] The exploratory endpoints for this study are:■ To characterize the CK profile of P-CD19CD20-ALLO1 on the basis of the number of CAR T cells in blood at specified timepoints by flow cytometry; ■ To evaluate the impact of cell dose on P-CD19CD20-ALLO1 expansion, engraftment, and persistence on the basis of the relationship between P- CD19CD20-ALLO1 dose and P-CD19CD20-ALLO1 exposure (blood concentration or CK parameters of P-CD19CD20-ALLO1) and cytokine markers related to engraftment (e.g., IL- 15 and IL-7);■ To evaluate the preliminary relationship of P-CD19CD20-ALLO1 cell dose / exposure with clinical efficacy and safety on the basis of the following endpoints:o Relationship between P-CD19CD20-ALLO1 dose / exposure and change from baseline of the following at various timepoints, including but not limited to:■ SLE Disease Activity Assessments (SLEDAI-2K, BILAG-2004, PGA, CLASI, tender / swollen joint counts);Attorney Docket No.: 000218-0093-W01■ Patient-reported outcomes (PtGA and FACIT-F);■ Anti-dsDNA titers, C3, and C4;o Proportion of participants achieving the following at various timepoints, including but not limited to:■ SLE treatment response Indices (SRI-4 and BICLA); ■ Disease activity states (DORIS and LLDAS);■ Complete renal response (CRR), as applicable for participants with LN;o Additional endpoints, including but not limited to:■ Time to first SLE flare;■ Total number of SLE flares;■ Glucocorticoid and / or immunosuppressant use; and ■ Safety endpoints.■ To identify and / or evaluate biomarkers that can provide evidence of P- CD19CD20-ALLO1 activity (i.e., pharmacodynamic biomarkers), are associated with susceptibility to developing adverse events or can lead to improved adverse event monitoring or investigation (i.e., safety biomarkers), or can increase the knowledge and understanding of disease biology and drug safety or CK on the basis of the following endpoints:o Relationship between biomarkers and safety, CK, immunogenicity, activity, or other biomarker endpoints;o Longitudinal changes in phenotypic and functional markers of P-CD19CD20-ALLO1 cells;o Longitudinal changes in phenotypic and functional markers in PBMCs including but not limited to B-cell or other immune cell subset profiling (e.g., naive, active, and memory cells) and / or immune cell production of cytokines or other products;o Longitudinal changes in CRS-associated biomarkers (e.g., IL-6 and IL- 10);o Proportion of patients achieving B-cell counts below the limit of detection by flow cytometry; ando Duration of B-cell depletion.■ To evaluate the effect of P-CD19CD20-ALLO1 on lupus disease activity biomarkers on the basis of the following endpoints:Attorney Docket No.: 000218-0093-W01o Longitudinal changes in titers of antinuclear antibody (ANA) and double stranded DNA (dsDNA);o Longitudinal changes in titers of other lupus-associated autoantibodies (e.g., anti-Smith and anti-RNP);o Longitudinal changes in complement C3 and C4;■ To evaluate potential effects of immunogenicity to P-CD19CD20-ALLO1 on the basis of the following endpoints:o Relationship between P-CD 19CD20- ALLO 1 anti-drug antibody (ADA) status and safety, P-CD19CD20-ALLO1 CK, or efficacy endpoints; ando Relationship between the presence of CAR-specific cytotoxic T cells and safety, P-CD19CD20-ALLO1 CK, or efficacy endpoints.■ To evaluate the effect of rimiducid on P-CD19CD20-ALLO1 related AEs, as applicable on the basis of the incidence and severity of adverse events, with severity determined according to the CTCAE v5.0 grading scale before and after rimiducid use.■ To evaluate the pharmacokinetics of fludarabine and cyclophosphamide on the basis of the longitudinal changes in circulating immune cell numbers, phenotype, and function and / or production of cytokines or other products.

[0128] To evaluate cellular immunogenicity of P-CD19CD20-ALLO1 on the basis of cellular immunity (i.e., the cellular immune response to the CAR construct (extracellular domain of CD 19 and CD20 variable heavy chain) at baseline and during the study.

[0129] Clinical disease activity assessments that will be used in this study include Systemic Lupus Erythematosus Disease Activity Index 2000 (SLEDAI-2K), British Isles Lupus Assessment Group 2004 (BILAG-2004), Physician’s Global Assessment (PGA), Cutaneous Lupus Erythematosus Disease Area and Severity Index (CLASI), and tender / swollen joint counts.Systemic Lupus Erythematosus Disease Activity Index 2000 (SLEDAI-2K)

[0130] SLEDAI-2K is a valid, reliable, and sensitive instrument for assessing global disease activity. SLEDAI-2K contains 24 lupus-related items / descriptors thatAttorney Docket No.: 000218-0093-W01are marked as “present,” and most likely attributable to SLE or as “absent” in the participant within the past month. Each item has a fixed number of points based on general clinical meaningfulness (range: 1-8), with a maximum possible total score of 105. The SLEDAI-2K is valid for assessment of lupus-related items within the past 10 days and the past 30 days. For this study, a timeframe of 28 days (4 weeks) will be used.

[0131] A key principle for SLEDAI-2K scoring is attribution: SLEDAI-2K items will only be scored if the feature is most likely attributable to active SLE rather than alternative causes or chronic damage alone. By indicating that an item is present, the assessor believes the feature represents an active, inflammatory manifestation of SLE (e.g., hematuria due to active lupus nephritis). If any SLEDAL2K item is marked as “absent” due to an alternative non-SLE explanation (e.g, fever due to infection, thrombocytopenia due to drug cause), this alternative explanation must be documented.

[0132] SLEDAI-2K scoring will follow the instrument scoring instructions with protocol-specific clarifications (see Table 3, infra). Laboratory-based items will reflect the laboratory results obtained from the same visit for which SLEDAI-2K is completed (e.g., results from samples collected at the Week 4 visit should be used to complete the Week 4 SLEDAI-2K rather than using Week 2 laboratory results).

[0133] At Screening Visits 1 and 3, the SLEDAI-2K will also be used as part of the Eligibility Disease Activity Assessment (EDAA) to assist in eligibility determination. The EDAA is a modified version of SLEDAI-2K incorporating additional scoring requirements described in the Inclusion Criteria, supra. Participants must have an EDAA total activity score of greater than or equal to 8 at Screening Visits 1 and 3 to be considered eligible for enrollment.Table 3: Protocol-Specific Clarifications for SLEDAI-2K ItemsAttorney Docket No.: 000218-0093-W01Note for all laboratory items: If central labs for the current visit cannot be reported due to processing or laboratory error, these labs may be repeated within 14 days of the visit date and may be included in SLEDAI-2K scoring. Labs repeated between visits will also count towards the next visit’sSLEDAI-2K assessment if completed within 28 days of the assessment.aMitsikostas et al. 2004.bRosenbaum et al. 2019.British Isles Lupus Assessment Group 2004 (BILAG-2004)

[0134] BILAG 2004 is a global disease activity assessment that considers activity within nine organ systems. After referring to the item-specific descriptions in the BILAG-2004 Index Glossary to complete the 97-item BILAG-2004 Index, which records the severity of lupus-specific manifestation(s) and change over time by comparing the most recent 4 weeks of disease activity to that of the previous 4 weeks, each domain is assigned a category grade from A through E to describe the severity of organ involvement (A being the most severe and E being never involved) using the domain-specific algorithms detailed in the BILAG-2004 Index Scoring guidance.Attorney Docket No.: 000218-0093-W01

[0135] At Screening Visit 1 only, upon completion of the BILAG-2004 Index, the investigator should assign Category A and / or B grades to applicable domains for eligibility determination. All other visits involving BILAG-2004 assessment only require completion of the BILAG 2004 Index (with the Sponsor responsible for assigning domain-specific category grades upon receipt of data recorded on the BILAG-2004 Index).

[0136] Laboratory -based items will reflect the laboratory results obtained from the same visit at which the BILAG-2004 Index is completed (e.g., results from samples collected at the Week 4 visit should be used to complete the Week 4 BILAG-2004 Index rather than using Week 2 laboratory results).

[0137] In most cases, BILAG Category A and B scores correspond to worsening and / or significant SLE disease activity. However, the BILAG 2004 Index Scoring algorithms give unexpected results when a Category A or B manifestation improves and stabilizes but remains present to a degree that fulfills the higher category definition in the BILAG-2004 Index Scoring because it still meets severity criteria described in the BILAG-2004 Index Glossary. These “False A” or “False B” scores occur because only clinical improvement, not stability, results in an improved letter score from prior assessments.

[0138] To prevent participants from being assigned worsening scores despite stable disease, a protocol specific BILAG-2004 scoring rule has been added: Manifestations fulfilling “same” criteria after fulfilling “improving” criteria will not result in an increase in the BILAG category grade in the absence of worsening.

[0139] The Sponsor or its representatives will review BILAG 2004 scoring to ensure the data are consistent and accurate.Physician’s Global Assessment (PGA)

[0140] PGA is the physician’s assessment of overall SLE disease severity over the past 4 weeks. This instrument factors in all aspects of the participant’s SLE disease activity, including both clinical and laboratory assessments. It does not reflect non-SLE medical conditions.

[0141] The PGA must be completed by a trained and certified investigator. Given the inherent subjectivity in this assessment, the same assessor must evaluate a given participant over time whenever possible.Attorney Docket No.: 000218-0093-W01

[0142] The PGA is scored from 0 (no SLE activity) to 3 (the most severe SLE activity) on a Visual Analog Scale (VAS).

[0143] When scoring PGA, the score from the previous visit must be reviewed first, and the score for the present visit must be assigned relative to the previous visit’s disease activity.Cutaneous Lupus Erythematosus Disease Area and Severity Index (CLASI)

[0144] CLASI is a validated instrument for assessing cutaneous lupus erythematosus activity. It reports two global scores: one for current inflammatory activity (Total Activity Score), and one for damage (Total Damage Score).

[0145] To calculate the total scores described above, trained assessors evaluate erythema, scale / hypertrophy, dyspigmentation, and scarring / atrophy / panniculitis at 13 anatomic locations plus additional assessments for mucosal lesions and alopecia.

[0146] As described in the SLE Disease Activity Assessments Guide, CLASI scoring must be consistent with corresponding items in SLEDAI 2K (rash, alopecia, mucosal ulcers, vasculitis) and BILAG 2004 scoring.Tender / Swollen Joint Counts

[0147] A 28 joint count of swollen and tender joints is performed at each visit. The following joints on left and right sides are examined:• Metacarpophalangeal 1-5• Proximal interphalangeal 1-5 and• Shoulder, elbow, wrist, knee.

[0148] Each joint is evaluated separately for tenderness (via palpation) and swelling. Joints with intra articular injection within the preceding 4 weeks must be excluded.

[0149] As described in the SLE Disease Activity Assessments Guide, joint counts must be consistent with corresponding items in SLEDAI 2K and BILAG 2004 scoring.American Society for Transplantation and Cellular Therapy (ASTCT) Consensus Grading.

[0150] The severity of CRS will be assessed through use of the ASTCT grading scale for CRS (see Table 4, infra).Attorney Docket No.: 000218-0093-W01Table 4: ASTCT Grading Scale for Cytokine Release Syndrome (CRS)> > > >ASTCT = American Society for Transplantation and Cellular Therapy; BiPAP= bi-level positive airway pressure; CPAP= continuous positive airway pressure; CRS = cytokine release syndrome; CTCAE v5.0D National Cancer Institute's Common Terminology Criteria for Adverse Events, Version 5.0.Note: Organ toxicities associated with CRS may be graded according to CTCAE v5.0, but they do not influence CRS grading.a Fever may not be prominent in participants who have received antipyretics, corticosteroids, or anti-cytokine therapy. In such cases, the grade is driven by hypotension and / or hypoxia.bLow flow is defined as oxygen delivered at < 6 L / min via nasal cannula or oxygen delivered via blow-by. High flow is defined as oxygen delivered at > 6 L / min.0Participant may or may not be receiving vasopressin but must be receiving one vasopressor other than vasopressin.dParticipant may or may not be receiving vasopressin but must be receiving two or more vasopressors other than vasopressin.

[0151] The severity of ICANS will be assessed through use of the ASTCT grading scale for ICANS (see Table 5, infra), which takes into account the ICE score (see Table 6, infra).Attorney Docket No.: 000218-0093-W01Table 5: ASTCT Grading Scale for Immune Effector Cell-AssociatedNeurotoxicity Syndrome>ASTCT = American Society for Transplantation and Cellular Therapy; CTCAE v5.0 = National Cancer Institute's Common Terminology Criteria for Adverse Events, Version 5.0; ICANS = immune effector cell- associated neurotoxicity syndrome; ICE = immune effector cell-associated encephalopathy; NA=not applicable.aICANS grade is determined by the most severe event (ICE score, or neurologic symptoms of depressed level of consciousness, seizure activity, motor weakness, or elevated intracranial pressure or cerebral edema) not attributable to any other cause. For example, a participant with an ICE score of 3 who has a generalized seizure is classified as Grade 3 ICANS.bA participant with an ICE score of 0 who is awake with global aphasia may be classified as Grade 3 ICANS, but a participant with an ICE score of 0 who is unarousable may be classified as Grade 4 ICANS.cDepressed level of consciousness should not be attributable to any other cause (e.g., sedating medication).dTremors and myoclonus associated with immune effector cell therapies do not influence ICANS grading. They may be graded according to CTCAE v5.0.eIntracranial hemorrhage with or without associated edema is not considered a neurotoxicity feature and does not influence ICANS grading. It may be graded according to CTCAE v5.0.Attorney Docket No.: 000218-0093-W01Table 6: ICE Score" " """"ICE = immune effector cell-associated encephalopathy.a1 point for each correct answer.

[0152] The severity of HLH will be assessed through use of the ASTCT grading scale for HLH (see Table 7, infra).Table 7: ASTCT Grading Scale for Hemophagocytic LymphohistiocytosisASTCT = American Society for Transplantation and Cellular Therapy; CRS = cytokine release syndrome; CTCAE v5.0= National Cancer Institute's Common Terminology Criteria for Adverse Events, Version 5.0; HLH= hemophagocytic lymphohistiocytosis; ULN= upper limit of normal.Note: The ASTCT grading scale (Hines et al. 2023) is based on the CTCAE v5.0 "immune system disorders, other" category.aHLH is a pathological and biochemical hyperinflammatory syndrome that may present with fever, hepatosplenomegaly, organ failure, and neurologic toxicities and is associated with progression or new onset of hyperferritinemia, cytopenias, coagulopathy with hypofibrinogenemia, and / or elevation of liver enzymes. HLH and CRS that occur subsequent to T cell-redirecting therapy have overlapping features, and HLH-like symptoms may be seen in individuals with severe CRS. Although HLH is distinct from CRS in this setting, overlapping clinical features may make it difficult to distinguish HLH from CRS. Temporally, HLH may have a delayed onset and can occur as CRS is resolving or after CRS has resolved. In addition, HLH can occur in the absence of CRS.Attorney Docket No.: 000218-0093-W01International Bone Marrow Transplant Registry (IBMTR) Grading Scale For Graft Versus Host Disease.

[0153] The severity of GVHD will be assessed through use of the IBMTR grading scale for GVHD (see Table 8, infra).Table 8: Criteria for IBMTR Severity Index for Acute GVHD< < < > > >IBMTR= International Bone Marrow Transplant Registry.aAssign index based on maximum involvement in an individual organ system.Source: Rowlings et al. 1997; Schoemans et al. 2018.

Claims

Attorney Docket No.: 000218-0093-W01CLAIMS1. A method of treating systemic lupus erythematosus (SLE) in a human subject in need thereof comprising administering a P-CD19CD20-ALLO1 chimeric antigen receptor T cell (CAR-T cell) to the subject.

2. Use of a P-CD19CD20-ALLO1 chimeric antigen receptor T cell (CAR-T cell) in the manufacture of a medicament for treating systemic lupus erythematosus (SLE) in a human subj ect in need thereof.

3. A P-CD19CD20-ALLO1 chimeric antigen receptor T cell (CAR-T cell) for use in treating systemic lupus erythematosus (SLE) in a human subject in need thereof.

4. The method according to claim 1, the use according to claim 2, or the CAR-T cell for use according to claim 3, wherein the systemic lupus erythematosus (SLE) is severe SLE; treatment refractory SLE; highly active SLE; severe, treatmentrefractory SLE; severe, highly active SLE; highly active, treatment-refractory SLE; or highly active, severe, treatment-refractory SLE.

5. The method, use or CAR-T cell for use of claim 4, wherein the highly active SLE is identified by (a) British Isles Lupus Assessment Group (BILAG)-2004 Category A disease in one or more organ systems and / or Category B disease in two or more organ systems (excluding constitutional, musculoskeletal, and / or mucocutaneous organ systems for Category B); (b) a Systemic Lupus Erythematosus Disease Activity Index 2000 (SLEDAI-2K) score greater than or equal to 8; and (c) a Physician’s Global Assessment (PGA) score greater than or equal to 1.0 on a 0 to 3 Visual Analog Scale (VAS).

6. The method, use or CAR-T cell for use of claim 4 or 5, wherein the subject suffers from lupus nephritis.

7. The method, use or CAR-T cell for use of claim 4 or 5, wherein the subject does not suffer from lupus nephritis.

8. The method according to any one of claims 1 and 4-7, the use according to any one of claims 2 and 4-7, or the CAR-T cell for use according to any one of claims 3-7, wherein the CAR-T cell is administered to the subject intravenously or is formulated to be administered to the subject intravenously.

9. The method according to any one of claims 1 and 4-8, the use according to any one of claims 2 and 4-8, or the CAR-T cell for use according to any one of claimsAttorney Docket No.: 000218-0093-W013-8, wherein the CAR-T cell is administered as a single dose or is formulated to be administered as a single dose.

10. The method according to any one of claims 1 and 4-8, the use according to any one of claims 2 and 4-8, or the CAR-T cell for use according to any one of claims 3-8, wherein the CAR-T cell is administered in multiple doses or is formulated to be administered in multiple doses, optionally wherein each of the multiple doses is administered between 3 and 5 years after the previous dose.

11. The method according to any one of claims 1 and 4-10 the use according to any one of claims 2 and 4-10, or the CAR-T cell for use according to any one of claims 3-10, wherein the CAR-T cell is administered at a dose of about 5 x 106cells to about 1.2 x 109cells or is formulated to be administered at a dose of about 5 x 106to about 1.2 x 109cells.

12. The method according to any one of claims 1 and 4-11, the use according to any one of claims 2 and 4-11, or the CAR-T cell for use according to any one of claims 3-11, wherein administration of the CAR-T cell results in (a) a detectable number of CAR transgene copies in the blood of the subject, (b) a detectable number of CAR-T cells in the blood of the subject, (c) the depletion of B-cell levels in the subject by at least 90% percent, (d) immunosuppression-free remission in the subject for at least 6 months, (e) a time to first SLE flare of at least 6 months; (f) a total number of SLE flares less than one per year (g) reduced glucocorticoid and / or immunosuppressant use as compared to prior to treatment; (h) reduced titers antinuclear antibody (ANA) and double stranded DNA (dsDNA) as compared to prior to treatment; or (i) reduced titers of lupus-associated autoantibodies (e.g., anti-Smith and anti-RNP) as compared to prior to treatment.

13. The method according to any one of claims 1 and 4-12, the use according to any one of claims 2 and 4-12, or the CAR-T cell for use according to any one of claims 3-12, wherein administration of the CAR-T cell results in (a) an incidence of Grade 2 cytokine release syndrome (CRS) less than 30%, (b) an incidence of immune effector cell-associated neurotoxicity syndrome (ICANS) less than 20%, (c) an incidence of hemophagocytic lymphohistiocytosis (HLH) less than 10%, (d) a risk of serious infection no more than 10% more compared to the incidence of infection resulting from standard of care at 6 months after treatment, (e) an incidence of immune rejection and graft-versus-host disease (GVHD) less than 10%, or (f) an incidence of new T-cell malignancies less than 0.1%.Attorney Docket No.: 000218-0093-W0114. The method according to any one of claims 1 and 4-13, the use according to any one of claims 2 and 4-13, or the CAR-T cell for use according to any one of claims 3-13, wherein the subject has not been previously administered a therapeutic agent to treat SLE.

15. The method according to any one of claims 1 and 4-13, the use according to any one of claims 2 and 4-13, or the CAR-T cell for use according to any one of claims 3-13, wherein the subject has been previously administered a therapeutic agent to treat SLE.

16. The method, use, or CAR-T cell for use according to claim 15, wherein the subject is refractory to the previously administered therapeutic agent.

17. The method, use, or CAR-T cell for use according to claim 15 or 16, wherein the previously administered therapeutic agent is selected from a corticosteroid, an antimalarial therapy, an immunosuppressant, a B-cell depleting therapy, and a cytokine blocker.

18. The method, use, or CAR-T cell for use according to claim 17, wherein the antimalarial therapy is hydroxychloroquine.

19. The method, use, or CAR-T cell for use according to claim 17, wherein the immunosuppressant is selected from the group consisting of cyclophosphamide, azathioprine, mycophenolate mofetil (MMF), mycophenolic acid, and methotrexate.

20. The method, use, or CAR-T cell for use according to claim 17, wherein the B-cell depleting therapy is selected from the group consisting of rituximab, ocrelizumab, ofatumumab, belimumab and obinutuzumab.

21. The method, use, or CAR-T cell for use according to claim 17, wherein the cytokine blocker is anifrolumab.

22. The method according to any one of claims 1 and 4-21 , the use according to any one of claims 2 and 4-21, or the CAR-T cell for use according to any one of claims 3-21, wherein the CAR-T cell is administered or formulated to be administered in combination with one or more additional therapeutic agents.

23. The method, use, or CAR-T cell for use according to claim 22, wherein the CAR-T cell is administered or formulated to be administered sequentially with the one or more additional therapeutic agents.

24. The method, use, or CAR-T cell for use according to claim 22, wherein the CAR-T cell is administered or formulated to be administered simultaneously with the one or more additional therapeutic agents.Attorney Docket No.: 000218-0093-W0125. The method, use, or CAR-T cell for use according to any one of claims 22-24, wherein the one or more additional therapeutic agents is selected from an immunosuppressant, a B-cell depleting therapy, a calcineurin inhibitor, another biologic or targeted therapy, an IL-1 inhibitor, a corticosteroid, a kinase inhibitor, or a cytokine blocker.

26. The method, use, or CAR-T cell for use according to claim 25, wherein the immunosuppressant is selected from the group consisting of cyclophosphamide, azathioprine, mycophenolate mofetil (MMF), mycophenolic acid, and methotrexate.

27. The method, use, or CAR-T cell for use according to claim 25, wherein the B-cell depleting therapy is selected from the group consisting of rituximab, ocrelizumab, ofatumumab, belimumab and obinutuzumab.

28. The method, use, or CAR-T cell for use according to claim 25, wherein the calcineurin inhibitor is selected from the group consisting of ciclosporin, tacrolimus, and voclosporin.

29. The method, use, or CAR-T cell for use according to claim 25, wherein the kinase inhibitor is ruxolitinib.

30. The method, use, or CAR-T cell for use according to claim 25, wherein the cytokine blocker is selected from the group consisting of ruxolitinib, etoposide, or emapalumab.

31. The method according to any one of claims 1 and 4-30, the use according to any one of claims 2 and 4-30, or the CAR-T cell for use according to any one of claims 3-30, wherein the subject receives a lymphodepleting chemotherapy (LDC) regimen prior to the administration of CAR-T cell.

32. The method, use, or CAR-T cell for use of claim 31, wherein the LDC regimen comprises cyclophosphamide (300-750 mg / m2IV) and fludarabine (0-30 mg / m2IV) for 3 consecutive days.

33. The method, use, or CAR-T cell for use of claim 31 or 32, wherein the LDC regimen is administered on Days -5, -4, and -3, and the CAR-T cells is administered or to be administered on Day 1.

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