WO2019149249A1 - 一种结合bcma的嵌合抗原受体(car)及其应用 - Google Patents
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Definitions
- the present application relates to the field of biomedicine, and in particular to a chimeric antigen receptor capable of specifically binding to a BCMA protein.
- BCMA B cell mature antigen
- BAFF B cell activator receptor
- APRIL B cell proliferation-inducing ligand
- the chimeric antigen receptor is an antigen receptor designed to recognize cell surface antigens in a human leukocyte antigen-independent manner. Attempts to treat these types of patients with genetically modified T cells (CAR-T) expressing CAR have been somewhat successful (Molecular Therapy, 2010, 18: 4, 666-668; Blood, 2008, 112: 2261-2271) .
- BCMA BCMA
- the present application provides a chimeric antigen receptor that can specifically bind BCMA and uses thereof.
- the BCMA chimeric antigen receptors provided herein have one or more of the following properties: 1) have a higher affinity for the BCMA protein; 2) the CAR-T cells prepared using the CAR can stably express the CAR 3) CAR-T cells made using the CAR have a higher CAR positive rate; 4) the CAR can promote the release of cytokines; 5) can be used to treat diseases or conditions associated with the expression of BCMA.
- the application includes a chimeric antigen receptor (CAR), wherein the CAR comprises a BCMA binding domain, a transmembrane domain, a costimulatory domain, and an intracellular signaling domain, the BCMA binding domain
- CAR chimeric antigen receptor
- the CAR comprises a BCMA binding domain, a transmembrane domain, a costimulatory domain, and an intracellular signaling domain, the BCMA binding domain
- An antibody or fragment thereof comprising a specific binding to BCMA, wherein the antibody comprises a heavy chain complementarity determining region 1 (HCDR1), a heavy chain complementarity determining region 2 (HCDR2) and a heavy chain complementarity determining region 3 (HCDR3), said HCDR1
- HCDR1 heavy chain complementarity determining region 1
- HCDR2 heavy chain complementarity determining region 2
- HCDR3 heavy chain complementarity determining region 3
- the antibody comprises a heavy chain variable region, the amino acid sequence of which is set forth in SEQ ID NO: 7. In certain embodiments, the antibody comprises a light chain variable region, the amino acid sequence of which is set forth in SEQ ID NO: 15. In certain embodiments, the antibody is a single chain antibody. In certain embodiments, the antibody comprises the amino acid sequence set forth in SEQ ID NO:43.
- the transmembrane domain comprises the amino acid sequence set forth in SEQ ID NO:27.
- the co-stimulatory domain of the CAR comprises a costimulatory domain derived from a protein selected from the group consisting of CD28, 4-1BB, OX-40, and ICOS.
- the costimulatory domain comprises the amino acid sequence set forth in SEQ ID NO:29 or SEQ ID NO:31.
- the intracellular signaling domain of the CAR comprises a signaling domain derived from CD3 ⁇ . In certain embodiments, the intracellular signaling domain comprises the amino acid sequence set forth in SEQ ID NO:33.
- the CAR further comprises a hinge region joining the BCMA binding domain and the transmembrane domain.
- the hinge region comprises the amino acid sequence set forth in SEQ ID NO:25.
- the CAR further binds to a signal peptide.
- the signal peptide comprises the amino acid sequence set forth in SEQ ID NO:3.
- the CAR is also linked to a cleavage peptide.
- the cleavage peptide comprises an amino acid sequence from a T2A peptide.
- the cleavage peptide comprises the amino acid sequence set forth in SEQ ID NO:35.
- the CAR comprises the amino acid sequence set forth in SEQ ID NO: 49 or SEQ ID NO: 51.
- the application also includes an isolated nucleic acid molecule encoding a CAR as described herein.
- the application further encompasses an isolated nucleic acid molecule encoding a CAR comprising the nucleic acid sequence set forth in SEQ ID NO: 50 or SEQ ID NO: 52.
- the present application also encompasses a vector comprising a nucleic acid molecule as described herein.
- the vector is selected from the group consisting of a plasmid, a retroviral vector, and a lentiviral vector.
- the application further encompasses an immune effector cell comprising a CAR as described herein, a nucleic acid molecule described herein, or a vector described herein.
- the immune effector cells are selected from the group consisting of T lymphocytes and natural killer (NK) cells.
- the present application also encompasses a method of making an immune effector cell comprising introducing a vector described herein into an immune effector cell.
- the present application also encompasses a composition comprising an immune effector cell as described herein.
- the application further comprises the CAR, the nucleic acid molecule, the vector, or the use of the immune effector cell for the preparation of a medicament, wherein the medicament is for use in treating a BCMA expression.
- Disease or condition In certain embodiments, the disease or condition associated with expression of BCMA is cancer or a malignancy.
- Figure 1A shows the structure of the CAR described herein.
- Figure 1B shows the evaluation of the expression of the CAR described herein by GFP signaling.
- FIG. 2A shows a schematic diagram of the process of transient transfection of T cells using the CAR plasmid described herein;
- Figure 2B shows the expression of CAR molecules after transient transfection of T cells by the CAR plasmid described herein.
- Figure 3 shows the expression of CAR molecules after transient transfection of T cells by the CAR plasmid described in the present application.
- Figure 4 shows the results of biotitration detection of the CAR plasmid described in the present application after lentiviral packaging.
- FIG. 5 shows the growth of CAR-T cells as described herein.
- Figure 6 shows the detection of the binding ability of the CAR molecule of the CAR-T cell described in the present application to the BCMA protein.
- Figure 7 shows the results of flow cytometry of the CD107a degranulation experiment of the CAR-T cells described herein.
- Figure 8 shows the results of the CD107a degranulation experiment after incubation of the CAR-T cells described herein with different target cells.
- Figure 9 shows the results of the CD107a degranulation experiment after incubation of the CAR-T cells of the present application with different target cells under BCMA protein competition conditions.
- Figure 10 shows the results of cytokine release assay after co-incubation of CAR-T cells and target cells described herein.
- FIG. 11 shows the results of CAR-T cell function assays for different donors described herein.
- Figure 12 shows the in vitro killing effect test of the CAR-T cells of the present application against target cells.
- Figure 13 shows the results of a tumor-bearing mouse model experiment of the tumor killing effect of the CAR-T cells described in the present application.
- Figure 14 and Figure 15 show changes in cytokines in a tumor-bearing mouse model following administration of CAR-T cells as described herein.
- Figure 16 is a graph showing changes in CAR copy number in peripheral blood of tumor-bearing mice after administration of CAR-T cells as described herein.
- Figure 17 shows the in vivo therapeutic evaluation of the subject's CAR-T cells after administration.
- Figure 18 is a graph showing changes in CAR copy number in peripheral blood of a subject in the present application.
- the embodiments of the present invention are described below by way of specific embodiments, and those skilled in the art can readily appreciate other advantages and effects of the present invention from the disclosure of the present specification.
- the CAR described in the present application can specifically bind to BCMA, and the CAR-T cells prepared by using the CAR can stably express the CAR, and the CAR-T cells prepared by using the CAR have a higher CAR positive rate. Furthermore, the CAR can promote the release of cytokines and can be used to treat diseases or conditions associated with the expression of BCMA.
- CAR Chimeric Antigen Receptor
- CAR-T chimeric antigen receptor T cells
- antigen eg, tumor-associated antigen (TAA)
- TAA tumor-associated antigen
- the CAR can be combined with the T cell receptor-activated intracellular domain based on the antigen (eg, BCMA) specificity of the antibody.
- T cells that are genetically modified to express CAR can specifically recognize and eliminate malignant cells expressing the target antigen.
- BCMA and “B cell maturation antigen” are used interchangeably and generally refer to a protein encoded by the TNFRSF17 gene.
- the BCMA protein is a member of the tumor necrosis factor receptor family.
- the BCMA may be human BCMA, and its GenBank accession number is BAB60895.1.
- BCMA is a type III transmembrane protein with a cysteine-rich domain (CRD) characteristic of members of the TNFR family in the extracellular domain (ECD), which forms a ligand binding motif.
- CCD cysteine-rich domain
- BCMA is a B-cell biomarker expressed in tumor cells (eg, multiple myeloma cells) or on the surface of tumor cells (eg, present in multiple myeloma malignant plasma cells).
- BCMA proteins may also include fragments of BCMA, such as extracellular domains and fragments thereof, such as binding domains, transmembrane domains, costimulatory domains, and intracellular signaling domains, as well as fragments that bind to any of the antibodies of the present application.
- the term "BCMA binding domain” generally refers to a domain that can specifically bind to a BCMA protein.
- the BCMA binding domain can comprise a chimeric antigen receptor or fragment thereof, an anti-BCMA antibody or antigen-binding fragment thereof, which specifically binds to a human BCMA polypeptide expressed on a B cell.
- binding domain "binding domain”, "extracellular domain”, “extracellular binding domain”, “antigen-specific binding domain” and “extracellular antigen-specific binding domain” are used interchangeably herein.
- a domain or fragment of a CAR having the ability to specifically bind to a target antigen of interest eg, BCMA
- the BCMA binding domain can be a natural source, a synthetic source, a semi-synthetic source, or a recombinant source.
- an antibody generally refers to a polypeptide molecule that is capable of specifically recognizing and/or neutralizing a particular antigen.
- an antibody may comprise an immunoglobulin consisting of at least two heavy (H) chains and two light (L) chains inter-connected by disulfide bonds, and includes any molecule comprising an antigen-binding portion thereof.
- the term “antibody” includes monoclonal antibodies, antibody fragments or antibody derivatives including, but not limited to, human antibodies, humanized antibodies, chimeric antibodies, single domain antibodies (eg, dAbs), single chain antibodies (eg, scFv), And antibody fragments that bind to the antigen (eg, Fab, Fab', and (Fab) 2 fragments).
- antibody also includes all recombinant forms of the antibody, such as antibodies expressed in prokaryotic cells, unglycosylated antibodies, and any of the antigen-binding antibody fragments and derivatives thereof.
- Each heavy chain can be composed of a heavy chain variable region (VH) and a heavy chain constant region.
- Each light chain can be composed of a light chain variable region (VL) and a light chain constant region.
- VH and VL regions can be further distinguished as hypervariable regions termed complementarity determining regions (CDRs), which are interspersed in more conserved regions called framework regions (FR).
- CDRs complementarity determining regions
- Each VH and VL can be composed of three CDRs and four FR regions, which can be arranged from the amino terminus to the carboxy terminus in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3 and FR4.
- the variable regions of the heavy and light chains contain a binding domain that interacts with the antigen.
- the constant region of the antibody can mediate binding of the immunoglobulin to host tissues or factors, including various cells of the immune system (eg, effector cells) and the first component (Clq) of the classical complement system.
- the term "antigen-binding molecule” generally refers to a molecule comprising an antigen-binding region or antigen-binding portion capable of binding to a target antigen.
- the antigen binding molecule can be a protein or polypeptide.
- the target antigen is a B cell mature antigen (BCMA)
- BCMA binding molecule when the target antigen is a B cell mature antigen (BCMA), the antigen binding molecule that binds to BCMA is also referred to as a BCMA binding molecule.
- Antigen binding molecules include, for example, antibodies and antigen binding fragments thereof, single chain scFv antibodies, various fusions and conjugates constructed based on scFv, such as scFv-Fc antibodies, immunoconjugates, antibody drug conjugates (ADCs), Multi/bispecific antibody, chimeric antigen receptor (CAR).
- ADCs antibody drug conjugates
- CAR chimeric antigen receptor
- the antigen binding portion of an antibody typically comprises amino acid residues from a "complementarity determining region" or "CDR”.
- CDR complementarity determining region
- BCMA binding molecules are used interchangeably with “antibody of the present application” or “anti-BCMA antibodies” depending on the context.
- single-chain antibody may be an antibody obtained by linking the heavy chain variable region and the light chain variable region through a linker peptide.
- transmembrane domain generally refers to a domain in a CAR that crosses a cell membrane, which is linked to an intracellular signal transduction domain and functions to transmit a signal.
- costimulatory domain generally refers to an intracellular domain that can provide an immunostimulatory molecule that is a cell surface molecule required for an effective response of a lymphocyte to an antigen.
- the costimulatory domain may comprise a costimulatory domain of CD28, and may also include a costimulatory domain of the TNF receptor family, such as the costimulatory domains of OX40 and 4-1BB.
- the term "hinge region” generally refers to a junction region between an antigen binding region and an immune cell Fc receptor (FcR) binding region.
- HA-tag generally refers to a protein tag based on the human influenza hemagglutinin antigen, which is chemically derived from a human influenza virus hemagglutinin 98-106 amino acid. Short amino acid sequence. After the HA-tag sequence is ligated to one end of the protein of interest by molecular biological means, the specific antibody against the HA-tag can be used to bind the recombinant protein, which facilitates experiments such as immunohistochemistry (IHC) and Western Blotting (see Schembri, Laura et al. The HA tag is cleaved and loses immunoreactivity during apoptosis. Nature Methods. February 2007, 4(2): 107-108).
- IHC immunohistochemistry
- Western Blotting see Schembri, Laura et al. The HA tag is cleaved and loses immunoreactivity during apoptosis. Nature Methods. February 2007, 4(2): 107-108).
- the term "intracellular signaling domain” generally refers to a domain located within a cell capable of transducing a signal.
- the intracellular signaling domain can signal a signal into a cell.
- the intracellular signaling domain is the intracellular signaling domain of the chimeric antigen receptor.
- the intracellular signaling domain can be selected from the group consisting of a CD3 sputum intracellular domain, a CD28 intracellular domain, a CD28 intracellular domain, a 4-1BB intracellular domain, and an OX40 intracellular domain.
- signal peptide generally refers to a peptide chain that directs protein transfer.
- the signal peptide can be a short peptide chain that can be 5-30 amino acids in length.
- the term "shear peptide” refers to a class of polypeptides capable of effecting the function of a cleavage protein.
- the cleaved peptide can undergo protein cleavage via ribosome skipping rather than protease hydrolysis.
- the cleavage peptide can be a cleavage 2A peptide, which can include T2A, F2A, P2A, and the like.
- the term "marker detection signal” generally refers to a gene, protein or other molecule of a known function or sequence that is capable of specifically labeling a signal that can be detected.
- the label detection signal may be a fluorescent protein such as GFP, RFP, and YFP.
- the marker detection signal can be EGFRt.
- EGFRt generally refers to a gene encoding a truncated human epidermal growth factor receptor polypeptide.
- EGFRt lacks the distal membrane EGF binding domain and the cytoplasmic signaling tail, but retains the extracellular epitope recognized by the anti-EGFR antibody.
- EGFRt can be used as a non-immunogenic selection tool with genetically modified cell function as well as a tracking marker.
- the EGFRt can serve as a marker molecule for CAR-T cells. The EGFRt can abolish the CAR-T cell cetuximab mediated ADCC pathway in vivo if necessary (see US8802374B2).
- Kozak sequence generally refers to a (gcc)gccRccAUGG sequence that is shared in the mRNA of a eukaryote. It plays an important role in the initiation of the translation process and is recognized by the ribosome as a translation initiation site (see De Angioletti M et al. a novel silent beta-thalassaemia mutation, the first in the Kozak sequence. Br J Haematol. 2004, 124 (2): 224–31.).
- the term "isolated” generally refers to an antibody that is an antibody that has been separated from components in its natural environment.
- the antibody is purified to greater than 95% or 99% purity by, for example, electrophoresis (eg, SDS-PAGE, isoelectric focusing (IEF), capillary electrophoresis) or chromatography (eg, ion exchange or Determine by reverse phase HPLC).
- electrophoresis eg, SDS-PAGE, isoelectric focusing (IEF), capillary electrophoresis
- chromatography eg, ion exchange or Determine by reverse phase HPLC.
- nucleic acid molecule generally refers to a nucleotide, deoxyribonucleotide or ribonucleotide or analog thereof in isolated form of any length.
- the nucleic acid molecules described herein can be isolated from the natural environment.
- the nucleic acid molecules described herein can be produced or synthesized by (i) amplification in vitro, such as by polymerase chain reaction (PCR) amplification, (ii) Produced by clonal recombination, (iii) purified, for example by restriction enzyme digestion and gel electrophoresis, or (iv) synthesized, for example by chemical synthesis.
- PCR polymerase chain reaction
- the isolated nucleic acid is a nucleic acid molecule produced by recombinant DNA techniques.
- nucleic acids encoding the antibodies or antigen-binding fragments thereof can be prepared by a variety of methods known in the art including, but not limited to, restriction fragment manipulation or the use of synthetic oligonucleotides. Overlap extension PCR, see Sambrook et al, Molecular Cloning, A Laboratory Manual, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, 1989; and Ausube et al. Current Protocols in Molecular Biology, Greene Publishing and Wiley-Interscience, for specific procedures. New York NY, 1993.
- the "vector” generally refers to a nucleic acid molecule capable of self-replication in a suitable host for transferring the inserted nucleic acid molecule into and/or between host cells.
- the vector may include a vector mainly for inserting DNA or RNA into a cell, a vector mainly for replicating DNA or RNA, and a vector mainly for expression of transcription and/or translation of DNA or RNA.
- the carrier also includes a carrier having a plurality of the above functions.
- the vector may be a polynucleotide that is capable of being transcribed and translated into a polypeptide when introduced into a suitable host cell. Typically, the vector will produce the desired expression product by culturing a suitable host cell comprising the vector.
- one or more of the nucleic acid molecules may be included in the vector.
- other genes may be included in the vector, such as a marker gene that allows selection of the vector in a suitable host cell and under appropriate conditions.
- the vector may further comprise an expression control element that allows the coding region to be correctly expressed in a suitable host.
- control elements are well known to those skilled in the art and may include, for example, promoters, ribosome binding sites, enhancers, and other control elements that regulate gene transcription or mRNA translation.
- the expression control sequence is a tunable element.
- the specific structure of the expression control sequence may vary depending on the function of the species or cell type, but typically comprises 5' non-transcribed sequences and 5' and 3' untranslated sequences, respectively, involved in transcriptional and translational initiation, such as TATA boxes, plus Cap sequence, CAAT sequence, and the like.
- a 5' non-transcribed expression control sequence can comprise a promoter region, and the promoter region can comprise a promoter sequence for transcriptional control of a functionally linked nucleic acid.
- the vector described herein may be selected from the group consisting of a plasmid, a retroviral vector, and a lentiviral vector.
- the plasmid, retroviral vector and lentiviral vector described herein may comprise a CAR.
- plasmid generally refers to a DNA molecule other than a chromosome or a pseudonucleus in an organism such as a bacterium or a yeast.
- the plasmid can exist in the cytoplasm and has the ability to replicate autonomously, enabling it to maintain a constant copy number in the progeny cells and express the genetic information it carries. Plasmids are used as vectors for genes in genetic engineering studies.
- the term "retroviral vector” generally refers to a viral particle that can clone and express a foreign gene but is not self-packaging into a proliferative capacity. Many of these viruses have reverse transcriptase.
- the retrovirus contains at least three genes: gag, a gene containing a protein that constitutes a viral center and structure, pol, a gene containing a reverse transcriptase, and an env, and a gene that constitutes a viral coat.
- retroviral transfection retroviral vectors can randomly and stably integrate the genome and the foreign genes it carries into the host cell genome. For example, CAR molecules can be integrated into host cells.
- the term "lentiviral vector” generally refers to a diploid RNA viral vector belonging to a retrovirus.
- the lentiviral vector is based on the genome of the lentivirus, and removes several sequences related to the activity of the virus, making it biologically safe, and then introducing the sequence of the target gene required for the experiment into the genome of the genome. And a vector prepared by expressing the structure.
- the retroviral vector can randomly and stably integrate the genome and the foreign gene it carries into the host cell genome.
- the CAR molecule can be integrated into the host cell.
- transposon generally refers to a discrete DNA fragment containing a transposase gene.
- the flanking is a terminal inverted repeat (TIR) containing a transposase binding site.
- TIR terminal inverted repeat
- the transposase binds to TIR and allows the transposon to be transferred to a new site.
- the transposon described herein is a two-component system consisting of a plasmid carrying a CAR (transposon) and another plasmid carrying a transposase.
- the transposon can be introduced into a target cell by means of electrical transduction or the like.
- the two components are electroporated into Peripheral Blood Mononuclear Cell (PBMC), and the expressed transposase acts on the terminal inverted repeat (TIR) of both sides of the CAR.
- the CAR (transposon) is cleaved and subsequently integrated into the TA dinucleotide sequence in the genome of the target cell (eg, T cell).
- the CAR protein can be expressed on the surface of target cells (see Cheng Zhang, Jun Liu, Jiang F Zhong, et al. Engineering CAR-T cells. Biomarker Research. 2017, 5:22).
- the term "gene editing” generally refers to a technique for site-directed modification of a genome. It may include zinc finger nucleases (ZFNs), transcription activator like effector nucleases (TALENs), and clustered regular interspaced short palindromic repeats/ Technologies such as CRISPR-associated (Cas9), CRISPR/Cas9). It can efficiently target the genome and modify it by adding, removing or altering genetic material at specific locations in the genome.
- Gene editing as described herein can include introducing a CAR molecule into the genome of a recipient cell by genetic editing techniques (e.g., CRISPR-Cas9).
- the term "immune effector cells” generally refers to immune cells involved in the clearance of foreign antigens and functioning effects in an immune response.
- the immune effector cells can be plasma cells, cytotoxic T cells, NK cells, APSC pluripotent cells, mast cells, and the like.
- pharmaceutically acceptable adjuvant generally refers to a pharmaceutically acceptable formulation carrier, solution or additive that enhances the properties of the formulation.
- additives are well known to those skilled in the art.
- cancer generally refers to a disease caused by a disorder that controls the proliferation of cells.
- hyperproliferative diseases known as cancer include, but are not limited to, solid tumors, such as occurring in the breast, respiratory, brain, reproductive organs, digestive tract, urinary tract, eye, liver, skin, head and neck, thyroid gland, thyroid Glandular cancers, as well as their distant metastases.
- diseases also include lymphomas, sarcomas, and leukemias.
- breast cancer include, but are not limited to, invasive ductal carcinoma, invasive lobular carcinoma, ductal carcinoma in situ, and lobular carcinoma in situ.
- cancers of the respiratory tract include, but are not limited to, small cell lung cancer and non-small cell lung cancer, as well as bronchial adenomas and pleural pulmonary blastomas.
- brain cancer include, but are not limited to, brain stem and hypothalamic keratinoma, cerebellar and cerebral astrocytoma, medulloblastoma, ependymoma, and neuroectoderm and pineal tumors.
- Male genital tumors include, but are not limited to, prostate and testicular cancer.
- Female genital tumors include, but are not limited to, endometrial cancer, cervical cancer, ovarian cancer, vaginal cancer, and vulvar cancer, as well as uterine tumors.
- Gastrointestinal tumors include, but are not limited to, the anus, colon, colorectal, esophagus, gallbladder, stomach, pancreas, rectum, small intestine, and salivary gland cancer.
- Urethral tumors include, but are not limited to, the bladder, penis, kidney, renal pelvis, ureter, and urethra cancer.
- Eye cancer includes, but is not limited to, intraocular melanoma and retinoblastoma.
- liver cancer examples include, but are not limited to, hepatocellular carcinoma (hepatoblastoma with or without fibrolamellar variation), cholangiocarcinoma (intrahepatic cholangiocarcinoma), and mixed hepatocellular cholangiocarcinoma.
- Skin cancers include, but are not limited to, squamous cell carcinoma, Kaposi's sarcoma, malignant melanoma, Merkel cell skin cancer, and non-melanoma skin cancer.
- Head and neck cancers include, but are not limited to, laryngeal/hypharyngeal/nasopharynx/ oropharyngeal cancer, and lip and oral cancer.
- Lymphoma includes, but is not limited to, AIDS-related lymphoma, non-Hodgkin's lymphoma, cutaneous T-cell lymphoma, Hodgkin's disease, and central nervous system lymphoma.
- Sarcomas include, but are not limited to, soft tissue sarcoma, osteosarcoma, malignant fibrous histiocytoma, lymphosarcoma, and rhabdomyosarcoma.
- Leukemias include, but are not limited to, acute myeloid leukemia, acute lymphocytic leukemia, chronic lymphocytic leukemia, chronic myeloid leukemia, and hairy cell leukemia.
- the term "about” generally means a range of 0.5% to 10% above or below a specified value, such as 0.5%, 1%, 1.5%, 2%, 2.5% above or below a specified value, Variations within the range of 3%, 3.5%, 4%, 4.5%, 5%, 5.5%, 6%, 6.5%, 7%, 7.5%, 8%, 8.5%, 9%, 9.5%, or 10%.
- the CAR may comprise an extracellular domain, a transmembrane domain, an intracellular costimulatory signaling domain, and an intracellular signaling domain that specifically bind to BCMA.
- the extracellular domain of the CAR may comprise a single chain antibody (scFv) as described herein.
- the single chain antibody can be linked to a transmembrane domain by a hinge region, such as a CD8 hinge.
- the CAR can be used to transduce immune effector cells (eg, T cells) and express them on the cell surface.
- T cells eg, T cells
- the present application can also provide T cells expressing the chimeric antigen receptor, and the use of the T cells and/or the CAR for the preparation of a medicament for treating a B cell related disease.
- the chimeric antigen receptor may comprise a BCMA binding domain, a transmembrane domain, a costimulatory domain, and an intracellular signaling domain.
- the BCMA binding domain may comprise an antibody fragment that specifically binds to BCMA, which antibody may comprise heavy chain complementarity determining region 1 (HCDR1), heavy chain complementarity determining region 2 (HCDR2) and heavy chain complementation determining Region 3 (HCDR3), which may in turn comprise the amino acid sequences set forth in SEQ ID NOs: 9-11, respectively; the antibody may comprise a light chain complementarity determining region 1 (LCDR1), a light chain complementarity determining region 2 ( LCDR2) and light chain complementarity determining region 3 (LCDR3), which may in turn comprise the amino acid sequences set forth in SEQ ID NOS: 17-19, respectively.
- HCDR1 heavy chain complementarity determining region 1
- HCDR2 heavy chain complementarity determining region 2
- HCDR3 heavy chain complementation determining Region 3
- the antibody may comprise a heavy chain variable region, and the heavy chain variable region may comprise the amino acid sequence set forth in SEQ ID NO: 7.
- the antibody may comprise a light chain variable region, and the light chain variable region may comprise the amino acid sequence set forth in SEQ ID NO: 15.
- the antibody may be a single chain antibody.
- the antibody may comprise the amino acid sequence set forth in SEQ ID NO:43.
- the single chain antibody can include scFv0026, the sequence of which is set forth in SEQ ID NO:43.
- a single chain antibody as described herein can be scFv0026, the sequence of which is set forth in SEQ ID NO:43.
- the amino acid sequences of LCDR1-3 of the single-chain antibody scFv0026 are shown in SEQ ID NO: 17, SEQ ID NO: 18, and SEQ ID NO: 19, respectively; the amino acid sequence of VL is shown in SEQ ID NO: 15; The amino acid sequences are shown in SEQ ID NO: 9, SEQ ID NO: 10 and SEQ ID NO: 11, respectively; the amino acid sequence of VH is shown in SEQ ID NO: 7.
- the CAR described herein may comprise a transmembrane domain, which may comprise a transmembrane domain from a protein selected from the group consisting of a, beta or ⁇ chain of a T cell receptor, CD28, CD3e, CD45, CD4, CD5, CD8a, CD9, CD16, CD22, CD33, CD37, CD64, CD80, CD86, CD134, CD137 and CD154.
- the transmembrane domain may comprise the amino acid sequence set forth in SEQ ID NO:27.
- a transmembrane domain of the present application can include a transmembrane domain of CD8a, the sequence of which is set forth in SEQ ID NO:27.
- the costimulatory domain may comprise a costimulatory domain from a protein selected from the group consisting of CD28, 4-1BB, OX40 and ICOS.
- the costimulatory domain may comprise the amino acid sequence set forth in SEQ ID NO: 29 or SEQ ID NO:31.
- the CARs described herein can include an intracellular signaling domain that can comprise a signaling domain from CD3 ⁇ .
- the intracellular signaling domain may comprise the amino acid sequence set forth in SEQ ID NO:33.
- the CAR described herein can include a hinge region that can link the antibody to the transmembrane domain.
- the hinge region may comprise the amino acid sequence set forth in SEQ ID NO:25.
- the CAR described herein may also include an HA-tag, which may be located at the N-terminus of the CAR.
- the HA-tag may comprise the amino acid sequence set forth in SEQ ID NO: 5.
- the expression of the CAR described herein can be detected by using an anti-HA antibody to specifically bind thereto and used to enrich CAR-T cells for functional studies.
- the CAR described herein can be linked to a signal peptide, which can comprise the amino acid sequence set forth in SEQ ID NO:3.
- the signal peptide can be a CD8a signal peptide, the sequence of which is set forth in SEQ ID NO: 3.
- CAR0037, CAR0085, CAR0087 can link the CD8a signal peptide.
- the CAR may also be linked to a cleavage peptide.
- the cleavage peptide may comprise an amino acid sequence from a T2A peptide.
- the cleavage peptide may comprise the amino acid sequence set forth in SEQ ID NO:35.
- the cleavage peptide can be T2A, the sequence of which is set forth in SEQ ID NO:35.
- CAR0037, CAR0087 can link the cleavage peptide T2A.
- the CAR may also be coupled to a marker detection signal, which may be located at the C-terminus of the CAR.
- the marker detection signal can be a fluorescent protein, which can be selected from the group consisting of GFP, RFP and YFP.
- the expression of a CAR molecule can be indirectly evaluated by detecting a signal of GFP.
- the CAR can include CAR0037 with a marker detection signal sequence as set forth in SEQ ID NO:37.
- the marker detection signal can be EGFRt.
- CAR0087 can be ligated to a marker detection signal as shown in SEQ ID NO:39.
- the CAR can be ligated to a Kozak sequence, the sequence of which is set forth in SEQ ID NO: 1.
- the CAR can be linked to a Kozak sequence, which can be located at the N-terminus of the CAR.
- CAR0037, CAR0085 or CAR0087 can be ligated to the Kozak sequence of SEQ ID NO: 1.
- the CAR may comprise the amino acid sequence set forth in SEQ ID NO: 49 or SEQ ID NO: 51.
- the CAR can be selected from CAR0037, the sequence of which is set forth in SEQ ID NO:49.
- the CAR can be selected from CAR0085, the sequence of which is set forth in SEQ ID NO: 51; the CAR can be selected from CAR0087, the sequence of which is set forth in SEQ ID NO:51.
- the CARs described herein can include, in order from the N-terminus, a BCMA binding domain, a transmembrane domain, a costimulatory domain, and an intracellular signaling domain.
- the CAR may comprise a BCMA binding domain, and the BCMA binding domain sequence is set forth in SEQ ID NO:43.
- the BCMA binding domain may comprise HCDR1-3, the sequences thereof are sequentially shown as SEQ ID NOs: 9-11, respectively; and the BCMA binding domain may comprise LCDR1-3, the sequences of which are respectively SEQ ID NO: 17-19.
- the CAR can include CAR0037 or a CAR as described herein having the same LCDR1-3 and HCDR1-3.
- the BCMA binding domain may comprise a heavy chain variable region, the sequence of which is set forth in SEQ ID NO: 7; and the BCMA binding domain may further comprise a light chain variable region, the sequence of which is set forth in SEQ ID NO: 15 Shown.
- the CAR can include CAR0037 or a CAR as described herein having the same light chain variable region and heavy chain variable region.
- a linker peptide may also be included between the light chain variable region and the heavy chain variable region, the sequence of which is set forth in SEQ ID NO:23.
- the CAR can include CAR0037 or a CAR as described herein having the same linker peptide.
- the transmembrane domain may comprise a transmembrane domain from CD8a, the sequence of which may be set forth in SEQ ID NO:27.
- the CAR can include CAR0037 or a CAR as described herein having the same transmembrane domain.
- the costimulatory domain may comprise a costimulatory structure from CD28, the sequence of which may be set forth in SEQ ID NO:29.
- the CAR can include CAR0037 or a CAR as described herein having the same costimulatory domain.
- the intracellular signaling domain may comprise a signaling domain from CD3 ⁇ , the sequence of which is set forth in SEQ ID NO:33.
- the CAR can include CAR0037 or a CAR described herein having the same intracellular signal transduction domain.
- the CAR may further comprise a hinge region, which may be located at the C-terminus of the BCMA binding domain and at the N-terminus of the transmembrane domain, the sequence of which is set forth in SEQ ID NO:25.
- the CAR can include CAR0037 or the CAR described herein having the same hinge region.
- the CAR may also be ligated to an HA-tag, which may be located at the N-terminus of the BCMA binding domain, the sequence of which is set forth in SEQ ID NO:5.
- the CAR can include CAR0037 or the CAR described herein having the same HA-tag.
- the CAR may also be linked to a signal peptide, which may be located at the N-terminus of the CAR, the sequence of which may be as set forth in SEQ ID NO:3.
- the CAR can also be linked to a cleavage peptide, such as: T2A.
- the cleavage peptide can be located at the C-terminus of the intracellular signal transduction domain, and the sequence can be as set forth in SEQ ID NO:35.
- the CAR can also be linked to a label detection signal that can be located at the C-terminus of the CAR (or the shear peptide).
- the marker detection signal can be selected from the group consisting of GFP, RFP and YFP, the sequence of which is set forth in SEQ ID NO:37.
- the CAR described herein may be CAR0037, and the amino acid sequences of LCDR1-3 are shown in SEQ ID NO: 17, SEQ ID NO: 18, and SEQ ID NO: 19, respectively;
- the amino acid sequence of VL is SEQ ID NO: 15 shows the amino acid sequence of HCDR1-3 as shown in SEQ ID NO: 9, SEQ ID NO: 10 and SEQ ID NO: 11, respectively;
- the amino acid sequence of VH is shown in SEQ ID NO: 7; between VH and VL
- the sequence of the linker peptide is set forth in SEQ ID NO: 23; its hinge region is set forth in SEQ ID NO: 25; its transmembrane domain is set forth in SEQ ID NO: 27; and its costimulatory domain is a CD28 costimulatory structure.
- the CD3 intracellular signaling domain is set forth in SEQ ID NO: 33;
- the CAR0043 may further comprise a cleavage peptide as set forth in SEQ ID NO: 35, and ID NO:37 GFP-tag detection signal;
- the CAR0037 may further comprise a KOZAK sequence as shown in SEQ ID NO: 1, such as the CD8a signal peptide set forth in SEQ ID NO: 3, as set forth in SEQ ID NO: The HA-label shown.
- the CAR described in the present application may be CAR0085, and the amino acid sequences of LCDR1-3 are shown in SEQ ID NO: 17, SEQ ID NO: 18, and SEQ ID NO: 19, respectively;
- the amino acid sequence of VL is SEQ ID NO: 15 shows the amino acid sequence of HCDR1-3 as shown in SEQ ID NO: 9, SEQ ID NO: 10 and SEQ ID NO: 11, respectively;
- the amino acid sequence of VH is shown in SEQ ID NO: 7; between VH and VL
- the sequence of the linker peptide is set forth in SEQ ID NO: 23; its hinge region is set forth in SEQ ID NO: 25; its transmembrane domain is set forth in SEQ ID NO: 27; and its costimulatory domain is 4-1BB.
- a stimulatory domain as set forth in SEQ ID NO: 31; a CD3 sputum signaling domain thereof as set forth in SEQ ID NO: 33; said CAR0085 may further comprise a KOZAK sequence as set forth in SEQ ID NO: 1, such as SEQ ID NO: The CD8a signal peptide shown in 3.
- the CAR described herein may be CAR0087, the amino acid sequences of LCDR1-3 are shown in SEQ ID NO: 17, SEQ ID NO: 18, and SEQ ID NO: 19, respectively; the amino acid sequence of VL is SEQ ID NO: 15 shows the amino acid sequence of HCDR1-3 as shown in SEQ ID NO: 9, SEQ ID NO: 10 and SEQ ID NO: 11, respectively; the amino acid sequence of VH is shown in SEQ ID NO: 7; between VH and VL
- the sequence of the linker peptide is set forth in SEQ ID NO: 23; its hinge region is set forth in SEQ ID NO: 25; its transmembrane domain is set forth in SEQ ID NO: 27; and its costimulatory domain is 4-1BB.
- a stimulatory domain as set forth in SEQ ID NO: 31; a CD3 sputum signaling domain thereof as set forth in SEQ ID NO: 33; said CAR0085 may further comprise a cleavage peptide as set forth in SEQ ID NO: 35, and The EGFRt marker detection signal as set forth in SEQ ID NO: 39; the CAR0087 may further comprise a KOZAK sequence as set forth in SEQ ID NO: 1, such as the CD8a signal peptide set forth in SEQ ID NO:3.
- a protein, polypeptide and/or amino acid sequence referred to in this application is also understood to include at least the following range: a functional variant or homologue having the same or similar function as the protein or polypeptide.
- the functional variant may be one or more substitutions, deletions or additions in the amino acid sequence of the protein and/or the polypeptide (eg, an antibody or fragment thereof that specifically binds BCMA) A protein or polypeptide of an amino acid.
- the functional variant may comprise having been substituted by at least 1, for example 1-30, 1-20 or 1-10, for example 1 , 2, 3, 4 or 5 amino acids.
- the functional variant may substantially retain the biological properties of the protein or the polypeptide prior to alteration (eg, substitution, deletion or addition).
- the functional variant can retain at least 60%, 70%, 80%, 90%, or 100% of the biological activity (eg, antigen binding capacity) of the protein or the polypeptide prior to the alteration.
- the substitution can be a conservative substitution.
- the homolog may be at least about 85% (eg, having at least about) the amino acid sequence of the protein and/or the polypeptide (eg, an antibody or fragment thereof that specifically binds BCMA) 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99% or more) sequence homology Sexual protein or polypeptide.
- the polypeptide eg, an antibody or fragment thereof that specifically binds BCMA
- the homology generally refers to the similarity, similarity or association between two or more sequences.
- the "percentage of sequence homology” can be calculated by comparing two sequences to be aligned in a comparison window to determine the presence of the same nucleic acid base in both sequences (eg, A, T, C, G, I) Or the same amino acid residues (eg, Ala, Pro, Ser, Thr, Gly, Val, Leu, Ile, Phe, Tyr, Trp, Lys, Arg, His, Asp, Glu, Asn, Gln, Cys, and Met)
- the number of positions is obtained to obtain the number of matching positions, the number of matching positions is divided by the total number of positions in the comparison window (ie, the window size), and the result is multiplied by 100 to generate a sequence homology percentage.
- Alignment to determine percent sequence homology can be accomplished in a variety of ways known in the art, for example, using publicly available computer software such as BLAST, BLAST-2, ALIGN or Megalign (DNASTAR) software.
- BLAST BLAST-2
- ALIGN ALIGN
- Megalign DNASTAR
- One of skill in the art can determine suitable parameters for aligning the sequences, including any algorithms needed to achieve maximum alignment within the full length sequence being compared or within the target sequence region.
- the homology can also be determined by the following methods: FASTA and BLAST. A description of the FASTA algorithm can be found in WRPearson and DJ Lipman, "Improved Tools for Biological Sequence Comparison," Proc. Natl. Acad.
- the application provides an isolated nucleic acid molecule that encodes a CAR as described herein.
- An isolated nucleic acid molecule encoding a CAR which may comprise a nucleic acid sequence set forth in SEQ ID NO: 50 or SEQ ID NO: 52, or a functional variant thereof.
- the nucleic acid molecules described herein can be isolated. For example, it may be produced or synthesized by (i) amplification in vitro, such as by polymerase chain reaction (PCR) amplification, (ii) production by clonal recombination, (iii) purification. , for example, by fractionation by gel digestion and gel electrophoresis, or (iv) synthetic, for example by chemical synthesis.
- the isolated nucleic acid is a nucleic acid molecule produced by recombinant DNA techniques.
- the application provides a vector that can comprise the nucleic acid molecule.
- the vector may be selected from one or more of a plasmid, a retroviral vector, and a lentiviral vector.
- the lentiviral vector described herein may comprise a CAR.
- a lentiviral vector as described herein can comprise a nucleic acid sequence set forth in SEQ ID NO: 50 and/or SEQ ID NO: 52, or a functional variant thereof.
- other genes may be included in the vector, such as a marker gene that allows selection of the vector in a suitable host cell and under appropriate conditions.
- the vector may further comprise an expression control element that allows the coding region to be correctly expressed in a suitable host.
- control elements are well known to those skilled in the art and may include, for example, promoters, ribosome binding sites, enhancers, and other control elements that regulate gene transcription or mRNA translation.
- the expression control sequence is a tunable element.
- the specific structure of the expression control sequence may vary depending on the function of the species or cell type, but typically comprises 5' non-transcribed sequences and 5' and 3' untranslated sequences, respectively, involved in transcriptional and translational initiation, such as TATA boxes, plus Cap sequence, CAAT sequence, and the like.
- a 5&apos non-transcribed expression control sequence can comprise a promoter region, and the promoter region can comprise a promoter sequence for transcriptional control of a functionally linked nucleic acid.
- nucleic acid molecules described herein can be operably linked to the expression control element.
- vectors may include, for example, plasmids, cosmids, viruses, bacteriophages or other vectors commonly used, for example, in genetic engineering.
- the vector is an expression vector, including a vector scFv plasmid and/or a CAR plasmid.
- the vector comprising the virus can be a lentiviral vector, which can comprise a vector scFv plasmid and/or a CAR plasmid.
- the virus can be a lentiviral LV0002, which can comprise a vector scFv plasmid PXL0008, which can comprise a nucleic acid scFv0008 molecule; and/or a CAR plasmid PXL0009, which can comprise a nucleic acid CAR0009 molecule.
- the virus can be lentiviral LV0011, which can comprise the vector scFv plasmid PXL0008, which can comprise a nucleic acid scFv0008 molecule, and/or a CAR plasmid PXL0041, which can comprise a nucleic acid CAR0041 molecule.
- the virus can be lentiviral LV0007, which can comprise the vector scFv plasmid PXL0026, which can include nucleic acid scFv0026, and/or CAR plasmid PXL0037, which can comprise nucleic acid CAR0037.
- the virus can be lentivirus LV0020, which can comprise the vector scFv plasmid PXL0026, which can comprise the nucleic acid scFv0026, and/or the CAR plasmid PXL0085, which can include the nucleic acid CAR0085.
- the virus can be lentivirus LV0021, which can comprise the vector scFv plasmid PXL0026, which can comprise the nucleic acid scFv0026, and/or the CAR plasmid PXL0087, which can comprise the nucleic acid CAR0087.
- the vector comprising the virus may further comprise a retroviral vector, which may include the scFv plasmid and/or CAR plasmid.
- the application provides an immune effector cell, which can comprise a CAR, a nucleic acid molecule, or a vector described herein.
- the immune effector cell can be a mammalian cell.
- the immune effector cells may be selected from the group consisting of T lymphocytes and natural killer (NK) cells.
- the T lymphocytes may include thymocytes, natural T lymphocytes, immature T lymphocytes, mature T lymphocytes, resting T lymphocytes, or activated T lymphocytes.
- the T cell may be a helper T cell (Th), such as a helper T cell 1 (Th1) or helper T cell 2 (Th2) cell.
- the T lymphocytes may be CD4 + helper T cells (HTL; CD4 + T cells), cytotoxic T cells (CTL; CD8 + T cells), tumor infiltrating cytotoxic T cells (TIL; CD8 + T cells), CD4 + /CD8 + T cells, CD4 - /CD8 - T cells or any other T lymphocyte subtype.
- the T lymphocytes can be naive T cells (T N cells). In certain embodiments, the T lymphocytes can be central memory T cells (T CM ). In certain embodiments, the T lymphocytes can be effector T cells (T EM cells). In certain embodiments, the T lymphocytes can be NKT cells. In the present application, the T lymphocytes may be derived from peripheral blood cells, cord blood cells, and/or white blood cells.
- the T lymphocytes may be T CM cells, which may be characterized by CD45RO + /CD62L + .
- the T lymphocytes may be T EM cells, which may be characterized by CD45RO + /CD62L ⁇ .
- the T lymphocytes may be T N cells, which may be characterized by CD45RO - /CD62L + .
- the T lymphocytes may be NKT cells, which are classified into NK1.1 + and NK1.1 - , CD4 + , CD4 - , CD8 + , and CD8 - .
- NKT cells can produce large amounts of interferon-gamma, IL-4 (interleukin 4), and granulocyte-macrophage colony-stimulating factor.
- IL-4 interleukin 4
- granulocyte-macrophage colony-stimulating factor IL-4 (interleukin 4)
- NKT cells can also produce some cytokines and chemokines (such as IL-2, IL-13, IL-17, IL-21, tumor necrosis factor- ⁇ ).
- the application provides a method of making an immune effector cell, which can comprise introducing a vector described herein into an immune effector cell.
- a vector described herein can be introduced into the immune effector cells, such as T lymphocytes or natural killer (NK) cells.
- each or each cell can comprise one or one of the vectors described herein.
- each or each cell can comprise multiple (eg, 2 or more) or multiple (eg, 2 or more) vectors as described herein.
- the vector can be introduced into an immune effector cell and the vector described herein can be introduced into the cell by methods known in the art.
- the immune effector cells can be transfected by a retroviral vector, and the viral genome carrying the CAR molecule can be integrated into the host genome to ensure long-term and stable expression of the target gene.
- a transposon is used, and a plasmid carrying a CAR (transposon) and a plasmid carrying a transposase are introduced into a target cell.
- CAR molecules can be added to the genome by genetic editing (eg, CRISPR/Cas9).
- a vector carrying a CAR molecule described herein can be introduced into the cell by methods known in the art, such as electroporation, lipofectamine 2000 (Invitrogen), and the like.
- the application provides a composition comprising the immune effector cells and a pharmaceutically acceptable adjuvant.
- the pharmaceutically acceptable adjuvant may include buffers, antioxidants, preservatives, low molecular weight polypeptides, proteins, hydrophilic polymers, amino acids, sugars, chelating agents, counterions, metal complexes, and/or nonionic surfaces. Active agent, etc.
- the composition can be formulated for oral administration, intravenous administration (for example, intravenous, IV), intramuscular administration (for example, intramuscular injection, IM), in situ at the tumor site.
- intravenous administration for example, intravenous, IV
- intramuscular administration for example, intramuscular injection, IM
- in situ at the tumor site for example, intramuscular injection, IM
- compositions described herein can comprise a therapeutically effective amount of the antibody or antigen-binding fragment thereof.
- the therapeutically effective amount is a dose that is capable of preventing and/or treating (at least partially treating) a condition or disorder (eg, cancer) and/or any of its complications in a subject having or at risk of developing.
- the application provides the use of the CAR, the nucleic acid molecule, the vector or the immune effector cell for the preparation of a medicament, wherein the medicament is for treating a gene associated with expression of BCMA A disease or condition.
- the disease or condition associated with expression of BCMA can be a cancer or a malignant tumor.
- the cancer or malignancy may be selected from a plasma cell malignancy disease, such as multiple myeloma, and may also be selected from a B cell malignant disease, such as Hodgkin's lymphoma and non-Hodgkin's lymphoma. .
- the application provides the CAR, the nucleic acid molecule, the vector or the immune effector cell, which treats a disease or condition associated with expression of BCMA.
- the application provides a method of treating a disease or condition associated with expression of BCMA comprising administering to the patient the CAR, the nucleic acid molecule, the vector, or the immune effector cell .
- KOZAK (its nucleotide sequence is SEQ ID NO: 2)
- CD8a signal peptide (its nucleotide sequence is SEQ ID NO: 4)
- HA-tag (its nucleotide The sequence is SEQ ID NO: 6)
- scFv0026 (the nucleotide sequence of which is SEQ ID NO: 44)
- the hinge region (the nucleotide sequence of which is SEQ ID NO: 26), and the transmembrane region (the nucleotide sequence thereof is SEQ ID NO: 28)
- CD28 costimulatory factor (its nucleotide sequence is SEQ ID NO: 30), 4-1BB costimulatory domain (its nucleotide sequence is SEQ ID NO: 32), CD3 intracellular signal a transduction domain (the nucleotide sequence of which is SEQ ID NO: 34), a T2A cleavage peptide (the nucleotide sequence of which is SEQ ID NO:
- the scFv0008 molecule was constructed as a control, and the amino acid sequence of the scFv0008 molecule is shown as SEQ ID NO: 41 (see US9034324, SEQ ID NO: 3 and SEQ ID NO: 4).
- the HA-tag is located at the N-terminus of the CAR molecule, which is directly linked to the CD8a signal peptide.
- the HA-tag can be used as a tag for detecting CAR molecules or enriching CAR-T cells.
- GFP is located at the most C-terminus of the CAR molecule, which is directly linked to the T2A cleavage peptide. Equal amounts of CAR molecules and GFP proteins are formed upon T2A cleavage (see Szymczak et al. correction of multi-gene deficiency in vivo using a single self-cleaving 2A peptide-based retroviral vector. nature biotechnology, 2004.
- FIG. 1B specifically shows the detection process as follows: lentiviral particles (1) enter the cell through cell membrane fusion (2), then de-package (3), followed by reverse transcription (4), followed by integration (5) and transcription (6) ), translation (7), and shearing by T2A cleavage peptide (8). Its transduction efficiency can be assessed by the expression of GFP (9), the binding efficiency of scFv and BCMA protein can be studied by BCMA-Fc (10), anti-HA antibody can be used to detect the expression of CAR and used to enrich CAR-T cells. Perform a functional analysis (11).
- the expression of CAR molecules can also be carried out by other methods.
- a suitable amount of biotinylated BCMA and PE streptavidin can be used to label the CAR molecule and then reflect the expression of the CAR molecule by the PE signal.
- a CAR molecule can be labeled for detection by an appropriate amount of biotinylated anti-HA mAb and streptavidin.
- the plasmid containing the scFv molecule, the plasmid containing the CAR molecule and the corresponding lentivirus thereof are shown in Table 1:
- the lentiviral vector PLVX-EF1alpha-IRES-Puro was digested with NotI and MluI, and the vector fragment was recovered.
- the candidate scFv plasmid PXL0026 (the nucleotide sequence of which is SEQ ID NO: 44) was PCR-amplified, and the NotI restriction site (containing the protective base), CD8a signal peptide, HA was sequentially carried in the 5' end by extension PCR.
- the T4 linkage was constructed to obtain the CAR plasmid numbered PXL0037 (the nucleotide sequence of CAR0037 is shown as SEQ ID NO: 50).
- the CAR plasmid numbered PXL0085 was obtained in a similar manner, and the lentiviral vector PLVX-EF1alpha-IRES-Puro was double-digested with NotI and MluI, and the vector fragment was recovered.
- the candidate scFv plasmid PXL0026 was PCR-amplified, and the NotI restriction site (containing the protective base), CD8a signal peptide, hinge region, transmembrane region, and 4-1BB costimulatory factor were sequentially carried in the 5' end by extension PCR.
- a CAR plasmid numbered PXL0087 was obtained in a similar manner, and the lentiviral vector PLVX-EF1alpha-IRES-Puro was double-digested with NotI and MluI, and the vector fragment was recovered.
- the candidate scFv plasmid PXL0026 was PCR-amplified, and the NotI restriction site (containing the protective base), CD8a signal peptide, hinge region, transmembrane region, and 4-1BB costimulatory factor were sequentially carried in the 5' end by extension PCR.
- nucleotide sequence is SEQ ID NO: 32
- CD3 sputum intracellular signaling domain T2A cleavage peptide
- EGFRt its nucleotide sequence is SEQ ID NO: 40
- gene synthesis PCR amplification; then with overlap
- a PCR fragment having a MluI site at the 3' end of the NotI restriction site at the 5' end was obtained by PCR, and the fragment was digested with NotI and MluI, and recovered.
- the T4 linkage was constructed to obtain the CAR plasmid numbered PXL0087 (the nucleotide sequence of the CAR molecule portion of CAR0087 is shown as SEQ ID NO: 52).
- a CAR plasmid containing the scFv plasmid PXL0008 was also constructed as a control:
- the CAR plasmid numbered PXL0041 was then obtained in the same manner (the nucleotide sequence of the CAR molecule portion of CAR0041 is shown as SEQ ID NO: 48).
- the lentiviral vector PLVX-EF1alpha-IRES-Puro was digested with NotI and MluI, and the vector fragment was recovered.
- the candidate scFv plasmid PXL0008 (the nucleotide sequence of which is SEQ ID NO: 42) was PCR-amplified, and the NotI restriction site (containing the protective base), CD8a signal peptide, HA was sequentially carried in the 5' end by extension PCR.
- the T4 linkage was constructed to obtain the CAR plasmid numbered PXL0041 (the nucleotide sequence of the CAR molecule portion of CAR0041 is shown as SEQ ID NO: 48).
- a CAR plasmid numbered PXL0009 was obtained in a similar manner, and the lentiviral vector PLVX-EF1alpha-IRES-Puro was double-digested with NotI and MluI, and the vector fragment was recovered.
- the candidate scFv plasmid PXL0008 was amplified by PCR, and the NotI restriction site (containing the protective base) and the CD8a signal peptide were sequentially carried out at the 5' end by extension PCR; hinge region, transmembrane region, CD28 costimulatory factor, CD3 cell Internal signal transduction domain gene synthesis, PCR amplification; then PCR PCR was used to obtain the PCR fragment of the 5' end with the MluI site at the 3' end of the NotI restriction site, and the fragment was digested with NotI and MluI, and Recycling.
- the T4 linkage was constructed to obtain the CAR plasmid numbered PXL0009 (the nucleotide sequence of the CAR molecule portion of CAR0009 is shown as SEQ ID NO: 46).
- PEI of PXL0009, PXL0041 and PXL0037 prepared in Example 1 was transiently transfected into 293T cells using PEI as a transfection reagent to obtain PXL0009-293T cells, PXL0041-293T cells and PXL0037-293T cells, respectively.
- PXL0009-293T cells, PXL0041-293T cells, and PXL0037-293T cells after 72 hours of transient transfection were used to evaluate the expression ability of candidate CAR molecules.
- 1 indicates plasmid and transfection reagent
- 2 indicates transient transfection
- 3 indicates expression and T2A cleavage peptide cleavage.
- the expression of the CAR molecule was detected by the method described in Example 1. Specifically, in the presence of excess and fixed concentrations of PE steptavidin, the amount of biotinylated BCMA can be changed by gradient dilution to obtain a change in PE signal (as shown in Figure 2B). Show).
- the X-axis is the GFP protein signal expressed in the cell
- the Y-axis is the PE signal obtained by using a gradient-diluted biotinylated BCMA (biotinylated BCMA) and a fixed amount of PE-streptavidin-labeled CAR molecule.
- PE-signaling curves obtained by transiently transfected 293T cells from candidate CAR plasmids were detected using a gradient diluted biotinylated BCMA protein (295.86 nM to 3.79 pM) (shown in Figure 3). Calculated from the curve fitting BCMA protein to the cell surface binding molecules CAR EC 50 values shown in Table 2.
- PXL0009 is a reference plasmid known to normally express CAR molecules, which does not contain a gene encoding a GFP protein and an HA tag, and its sequence is shown in SEQ ID NO:45.
- the GFP signal should not be detected in the cell sample, nor the PE signal of the CAR binding to the biotinylated anti-HA mAb. Therefore, the PXL0009-293T cell sample can be used as a control sample for flow cytometry.
- PXL0041 and PXL0037 which are reference plasmids, encode GFP and HA tags, so GFP signaling can be detected in PXL0041-293T cell samples, and CAR and biotinylated BCMA can be detected separately.
- Protein or binding to a biotinylated anti-HA mAb (PE signal) can be used as a positive control.
- the CAR plasmids numbered PXL0009, PXL0041 and PXL0037 prepared in Example 1 required simultaneous co-transfection of 293T cells with a shuttle plasmid and other packaging plasmids for lentiviral packaging in cells. Specific steps are as follows:
- lentivirus package in a 10 cm culture dish 293T cells were seeded at a density of 6 ⁇ 10 4 /cm 2 in DMEM medium containing 10% FBS and placed at 37 ° C, 5% CO. 2 The culture was carried out under an environment of saturated humidity, and transfection was carried out 3 days after the inoculation. Two EP tubes were taken before transfection, each with 500 ⁇ l of opti-MEM, and one of the tubes was added with lentiviral helper vector 3 ⁇ g PSPAX2, 2pMD2.G and the vector prepared in Example 1 (CAR plasmids numbered PXL0009, PXL0041 or PXL0037).
- the supernatant was collected into a centrifuge tube, centrifuged at 3000 g for 4 min at 4 ° C, and the supernatant was filtered through a 0.45 ⁇ m filter to prepare for purification.
- LV0002 (corresponding to the CAR plasmid of PXL0009), LV0011 (corresponding to the CAR plasmid of PXL0041), and LV0007 (the CAR plasmid corresponding to PXL0037) lentivirus were obtained, respectively.
- the packaging efficiency of lentivirus can be evaluated by detecting the virus titer (biological titer) with transduction activity in the supernatant harvested during lentiviral packaging.
- the specific detection steps are as follows:
- 293T cells were seeded in a 6-well plate at a dose of 1 ⁇ 10 5 /well, and cultured in 500 ⁇ l of DMEM medium containing 10% FBS under an environment of 37 ° C, 5% CO 2 and saturated humidity. After 24 hours of cell culture, lentiviral transduction was carried out, and 100 ⁇ l, 50 ⁇ l, 25 ⁇ l, and 12.5 ⁇ l of the above supernatant were added to a six-well plate (two wells per sample amount). The cells were then cultured at 37 ° C, 5% CO 2 and saturated humidity. After 72 hours of lentiviral transduction, 293T cells were digested and resuspended for flow cytometry.
- LV0007-transduced 293T cells express both CAR and GFP.
- Flow cytometric detection of GFP fluorescence in 293T cells can be used to calculate lentiviral bio titers (called GFP titers) in the supernatant:
- Biological titer (GFP positive rate ⁇ 293T cell number) / virus sampling volume
- LV0007-transduced 293T cells were labeled with biotinylated BCMA (biotinylated BCMA) and PE-streptavidin, and the CAR positive rate (called CAR titer) was detected by flow:
- Biological titer (CAR positive rate ⁇ 293T cell number) / virus sampling volume
- Table 4 shows the biological titer data for the lentiviral packaging supernatant.
- GFP titer generally refers to the detection of the GFP positive rate in the 293T cells transduced by the virus to be tested by the GFP signal, and the calculated titer
- CAR titer generally refers to the BCMA and PE chains through biotinylation. PE-streptavidin was used to detect the CAR positive rate in the 293T cells transduced by the virus to be tested, and the obtained titer was calculated.
- T cells were further obtained by CD3 MicroBeads sorting.
- the sorted T cells were activated using CD3/CD28 Dynabeads.
- the transduced T cells were exchanged once. Thereafter, the count was performed every day, and the cell density was maintained between (0.6 and 2.0) ⁇ 10 6 cells/ml, and the growth curve of the cells was plotted.
- CAR CAR
- GFP GFP
- the growth curves of LV0007-CAR-T cells, LV0011-CAR-T cells and T cell control groups are shown in Fig. 5.
- the data such as the positive CAR positive rate detected by flow cytometry are shown in Table 6.
- candidate CAR molecules expressed on T cells can normally bind to BCMA protein.
- EC 50 values of cell samples from different batches of preparation of the assay This difference may be due to differences in cell types, sample preparation methods, and batches, resulting in different cell surface CAR molecule expression densities.
- CD107a is a marker of intracellular microvesicles. When microvesicles loaded with granzymes are fused with cell membranes, CD107a on the cell membrane increases, when monasein (purchased from BioLegend) blocks its recovery. It can quantitatively reflect the intensity of microvesicle release. When CAR-T is stimulated by the target antigen on the target cell, granzyme release is caused, and the activation of T cells can be judged by flow detection of an increase in CD107a.
- the CAR-T cells (LV0007-CAR-T cells or LV0011-CAR-T cells) obtained in Example 4 were first incubated with target cells U266, monensin and CD107a antibodies for 3 to 6 hours, CAR-T cells.
- the cell density with the target cells was 5 ⁇ 10 5 cells/ml.
- the sample was then labeled with CD8, PD1 antibody and flow tested.
- CAR-positive cells in candidate CAR-T cells were detected by co-expressed GFP, while LV0011-CAR-T cells as a control labeled CAR-positive cells by biotinylated BCMA-Fc and PE streptomycin.
- the negative control in the experiment was co-incubated with K562 cells and CAR-T cells, and the positive control used a cocktail instead of the target cells to activate CAR-T cells.
- the P1 gate was selected on the FSC:SSC scatter plot to remove the cell debris in the lower left corner; for the cells in the P1 gate, CD8:PD1 could be further analyzed to obtain CD8+ /PD1-cell population (Q3); in the CD8+/PD1-cell population, GFP:CD107a can be analyzed again, and CD8+/PD1-/CAR+ cell population (CAR-positive cells labeled with co-expressed GFP signal) and CD8+ can be obtained, respectively.
- the ratio of CD107a expressed in the /PD1-/CAR-cell population was selected on the FSC:SSC scatter plot to remove the cell debris in the lower left corner; for the cells in the P1 gate, CD8:PD1 could be further analyzed to obtain CD8+ /PD1-cell population (Q3); in the CD8+/PD1-cell population, GFP:CD107a can be analyzed again, and CD8+/PD1-/CAR+ cell population (CAR-positive cells labeled with
- the cells were divided into two groups, P1 and P2, in which the cells in the P2 group showed almost no CD8, so it may be K562 cells;
- the cells in the gate can be further analyzed for CD8:PD1; in the CD8+/PD1-cell population (Q3), GFP:CD107a can be analyzed again, thereby also obtaining CD8+/PD1-/CAR+ and CD8+/PD1-/CAR-cell populations.
- the ratio of CD107a is expressed.
- CAR-T cell samples (LV0007-CAR-T and LV0011-CAR-T cells) were incubated with target cells U266 (BCMA positive) or K562 (BCMA negative) for 3 hours, respectively. Flow detection.
- the results of CD107a degranulation test data of cell samples are shown in Table 8 and Figure 8. Both Table 8 and Figure 8 show CD107a positive cells expressed in CD8+/PD1-/CAR+ and CD8+/PD1-/CAR- two subpopulation cells. ratio.
- the CD107a values on CD8+/PD1-/CAR+ subpopulations in CAR-T samples co-incubated with U266 cells can reflect the specific activation of CAR-T cells;
- the CD107a values on CD8+/PD1-/CAR+ subpopulations in co-incubated CAR-T samples can reflect the non-specific activation of CAR-T cells. Comparing the CD107a values on CD8+/PD1-/CAR+ subpopulation cells during U266 co-incubation, it can be seen that the CD8+/PD1-/CAR+ subpopulation of LV0007-CAR-T cells can be specifically activated by BCMA positive cells (U266). And the activation of LV0007-CAR-T cells was stronger than that of LV0011-CAR-T cells as a control.
- soluble BCMA Soluble BCMA is elevated in the serum of MM patients, and the concentration is positively correlated with the degree of tumor malignancy. Therefore, when CAR-T cells were co-incubated with target cells, 1 ⁇ g/ml BCMA protein was added to the medium to evaluate the effect of soluble BCMA on CAR-T cell activation.
- Cytokine release assay was performed by incubating CAR-T cells (5 ⁇ 10 5 cells, 100 ⁇ l) and target cells (5 ⁇ 10 5 cells, 100 ⁇ l) in RPMI-1640 medium for 24 hours. The supernatant was cultured, and then the secretion of factors such as IL-2, IL-4, IL-6, IL-10, IL-17A, TNF- ⁇ , and IFN- ⁇ was measured by CBA method.
- cytokine release assay results of cytokine release assay after co-incubation of CAR-T cell samples with target cells are shown in Table 10 and Figure 10.
- the amount of cytokine release shown in Figure 10 is a percentage relative to the maximum value detected in the sample.
- Table 10 and Figure 10 after stimulation with BCMA-positive target cells U266 (+U226), TNF- ⁇ , IFN- ⁇ , and IL-2 secreted by LV0007-CAR-T cells were greatly increased.
- BCMA-negative K562 stimulation (+K562) did not show increased secretion of TNF- ⁇ , IFN- ⁇ and IL-2.
- LV0002-CAR-T cells and LV0021-CAR-T cells were prepared by transduction of the lentiviruses LV0002 and LV0021 prepared in Example 3, using LV0021 virus (corresponding to the PXL0087 plasmid-encoded CAR molecule).
- the co-stimulatory factor is 4-1BB) and the LV0002 virus (corresponding to the CAR molecule encoded by the PXL0009 plasmid, the co-stimulatory factor is CD28) transfects T cells of different donors to prepare CAR-T cells.
- the T cell of LV0002 virus transduction donor 1 was named LV0002-D01-CAR-T cell, and the T cell of LV0002 virus transduction donor 1 was named LV0002-D02-CAR-T cell, and LV0002 virus was transferred.
- the T cell of the donor 3 was named LV0002-D03-CAR-T cells;
- the T cell of the LV0021 virus transduction donor 1 was named LV0021-D01-CAR-T cells, and the LV0021 virus was transduced into the T of the donor 2
- the cells were named LV0021-D01-CAR-T cells, and the T cells transducing donor LV of LV0021 virus were named LV0021-D03-CAR-T cells.
- donor T cells were used as controls.
- the T cell from the donor 1 was named T cell-D01
- the T cell from the donor 2 was named T cell-D02
- the T cell derived from the donor 3 was named T cell-D03.
- the function of these CAR-T cells was also examined using the CD107a degranulation experiment of Example 6, and the results are shown in Table 11 and Figure 11.
- results of LV0021-CAR-T cells from two different donors were similar, and all of them could be stimulated by BCMA-positive U266 cells to produce CD107a, while those stimulated by BCMA-negative K562 cells produced CD107a values and not.
- the stimulated values are similar (blank).
- LV0002-CAR-T cells as a control can produce higher CD107a under U266 cell stimulation, probably due to the difference in scFv and costimulatory factors.
- LV0002-CAR-T cells had higher CD107a values in both unstimulated (blank) and stimulated by K562 cells.
- the ability of LV0021-CAR-T to produce CD107a was less affected by free BCMA protein, while the ability of LV0002-CAR-T to produce CD107a was significantly affected by free BCMA protein.
- the in vitro killing function was detected using the CAR-T cell LV0021-D02-CAR-T prepared in Example 8, which can be specifically detected by calcein fluorescence method.
- the detection procedure was as follows: 5 ⁇ 10 5 BCMA-positive U266 cells and BCMA-negative K562 cells were respectively taken and resuspended in a PBS+4% FBS solution to a cell suspension of 1 ⁇ 10 6 cells/ml. U266 and K562 cells were then labeled with 25 ⁇ M calcein-AM, respectively.
- the labeled U266 and K562 cells were inoculated into the U-bottom 96-well plate according to the amount of 5000 cells per well; then, in the corresponding wells, according to the ratio of effector cells to target cells (E:T value) 50: 1.
- the ratio of 25:1, 5:1 was added to the CAR-T cells to be tested or T cells as a control, and the volume of the solution per well was 200 ⁇ l.
- PBS solution was used instead of effector cells to be added to U266 or K562 cells as a negative control for detection
- cell lysate solution was used instead of effector cells to be added to U266 or K562 cells as a positive control for detection.
- the relative proportion of target cells (U266 or K562) killed by effector cells and cleaved to release calcein can be calculated by the following formula:
- F test is the average fluorescence value between the target cells and the duplicate wells of the T/CAR-T cells to be tested
- F spont is the average fluorescence value between the duplicate wells containing the target cells and PBS
- F max is the mean fluorescence value between the cells and the duplicate wells of the lysate.
- CAR-T cells have a strong killing effect on BCMA-positive U266 cells, and the killing effect is The increase in the E:T value is enhanced by the fact that a greater proportion of U266 cells are cleaved to release calcein.
- CAR-T cells have a poor killing effect on BCMA-negative K562 cells.
- T cells have a certain non-specific killing of U266 cells, and non-specific killing does not change with the increase of E:T value. Therefore, CAR-T cells have a significant killing effect against BCMA.
- Example 10 In vivo tumor suppression test in a tumor-bearing animal model
- CAR-T cells (LV0021-D02-CAR-T) from donor 2 prepared in Example 8.
- the LV0021-CAR-T cells were named XL103-07.
- T cells (T cell-D02) from donor 2 were used as control T cells.
- the U266 cells in the expansion were subcutaneously injected into immunodeficient NSG mice in an amount of 2 ⁇ 10 6 cells per mouse to establish a U266 subcutaneous tumor-bearing model.
- the tumor-bearing mice were divided into 5 groups according to Table 12, and injected with CAR-T cells (XL103-07), control T cells (T cells-D02), PBS or cells, respectively.
- a cryopreservation solution (which included 7.5% DMSO, 23% human albumin, 32.5% compound electrolyte, 35% glucose injection, and 2% saline). The grouping and dosing schedules are shown in Table 12.
- mice were continuously fed and tumor volume and mouse body weight, as well as mouse survival status, were recorded. The result is shown in FIG.
- the results of the mouse model experiments showed that after a single injection of CAR-T cells, both the high-dose group and the low-dose group had good tumor killing effect, and the tumor completely disappeared 19 days after the injection. Tumors in mice injected with control T cells, PBS, or cell cryopreservation continued to grow.
- Example 11 cytokine detection in a tumor-bearing animal model
- cytometric bead array method cytometric beads array, CBA
- CBA cytometric beads array
- BD TM CBA Flex Set Reagents and BD FACS Array TM Bioanalyzer cytometric beads array, CBA
- XL103-07-H stands for high dose group, the dose is 10 ⁇ 10 6 cells (XL103-07 cells)/animal;
- XL103-07-L stands for low dose group, the dose is 2 ⁇ 10 6 cells (XL103- 07 cells) / animals.
- the results of IFN ⁇ and IL2 are shown in Figure 14.
- the results showed that the peaks of IFN ⁇ and IL2 in the low-dose group were on days 10 and 7 respectively; the peaks of IFN ⁇ and IL2 in the high-dose group were on days 3-7 and 3, respectively; The dose group is advanced.
- the level of the control T cell group was very low and there was no significant change trend.
- IL10, IL7, IL6 and TNF- ⁇ are shown in Fig. 15.
- the results in Figure 15 show that the cytokine levels are lower and there is no significant change trend compared to the control T cell group.
- XL103-07 in the genomic DNA of the peripheral blood cells of the animal model of the animal model in Example 10 was detected by quantitative PCR method, and the amplification of CAR-T cells in mice was analyzed. The results are shown in Fig. 16. Shown. Similarly, tumor-bearing mice were divided into three groups: control T cell group, XL103-07 high dose group and XL103-07 low dose group, 5 rats in each group. Among them, XL103-07-H stands for high dose group, the dose is 10 ⁇ 10 6 cells (XL103-07 cells)/animal; XL103-07-L stands for low dose group, the dose is 2 ⁇ 10 6 cells (XL103- 07 cells) / animals.
- the DNA copy number of CAR in peripheral blood of the 5 subjects described in Example 13 was examined using a droplet digital PCR method (see Bio-Rad QX200 specification) to evaluate the pharmacokinetic profile of CAR-T cells. The result is shown in Fig. 18. After administration, the product is rapidly amplified in vivo and can be detected at 60 days. In most subjects, the peak time of CAR-T was about 10 days, and the time of 001 was 17 days.
- the DNA copy number changes of CAR in Example 14 were further analyzed using the R 3.5.0 software NonCompart package, and the pharmacokinetic related parameters were calculated. The results are shown in Table 13.
- T0 indicates the initial non-zero concentration time
- the peak concentration (Cmax) indicates the maximum peak reached by the drug concentration
- the peak time (Tmax) indicates the time required to reach the peak concentration.
- AUC (0-28) refers to Indicates the integrated area under the curve from 0 to 28 days.
- AUC (0-CLST) refers to the integrated area under the curve from 0 to the last observation time.
- the first dose group had an average Cmax (98644 ⁇ 5000) copy number/ ⁇ g DNA, and the average AUC (0-28) was (1306653.3 ⁇ 100000); the second dose group had an average Cmax of (32487.5 ⁇ 2000) copies/ ⁇ g. DNA, the average AUC (0-28) was (317816.95 ⁇ 150,000).
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Abstract
Description
序号 | scFv质粒 | scFv分子 | CAR质粒 | CAR分子 | 慢病毒 |
1 | PXL0008 | scFv0008 | PXL0009 | CAR0009 | LV0002 |
2 | PXL0008 | scFv0008 | PXL0041 | CAR0041 | LV0011 |
3 | PXL0026 | scFv0026 | PXL0037 | CAR0037 | LV0007 |
4 | PXL0026 | scFv0026 | PXL0085 | CAR0085 | LV0020 |
5 | PXL0026 | scFv0026 | PXL0087 | CAR0087 | LV0021 |
样品 | GFP% | 用500ng BCMA标记的CAR% | CAR%:GFP% | EC 50(nM) |
PXL0037-293T | 38.5 | 9.2 | 0.24 | 0.64 |
PXL0041-293T | 41.8 | 16.1 | 0.39 | 0.30 |
PXL0009-293T | N/A | 18.0 | N/A | 1.59 |
样品 | GFP% | 用500ng anti-HA mAb标记的CAR% | CAR%:GFP% |
PXL0037-293T | 35.9 | 29.4 | 0.82 |
PXL0041-293T | 43.1 | 18.8 | 0.44 |
PXL0009-293T | N/A | N/A | N/A |
样品 | CAR滴度(TU/ml) | GFP滴度(TU/ml) |
LV0002 | 1.52E+05 | N/A |
LV0007 | 8.73E+05 | 1.05E+06 |
LV0011 | 4.68E+05 | 5.38E+05 |
样品 | CAR% | CAR MFI | CD8% | CD4中的CAR% | CD8中的CAR% |
LV0007-CAR-T | 56.3 | 25.1 | 35.6 | 54.0 | 56.5 |
LV0011-CAR-T | 62.8 | 30.7 | 37.6 | 58.3 | 68.5 |
T细胞 | N/A | N/A | 29.6 | N/A | N/A |
样品 | EC 50 |
LV0007-CAR-T | 6.00 |
LV0011-CAR-T | 0.31 |
样品 | IL-2 | IL-17A | IL-4 | IL-6 | IL-10 | IFN-γ | TNF-α |
LV0011-CAR-T | 44.72 | 4.61 | 14.12 | 10.83 | 5.85 | 277.35 | 569.91 |
LV0011-CAR-T+U226 | 8181.1 | 343.74 | 160.77 | 31.82 | 47.66 | 4847.04 | 3265.38 |
LV0011-CAR-T+K562 | 117.31 | 4.1 | 3.04 | 126.72 | 0.45 | 4.25 | 942.7 |
LV007-CAR-T | 1487.07 | 36.95 | 20.63 | 17.46 | 15.15 | 1248.25 | 986.38 |
LV0007-CAR-T+U226 | 8181.1 | 443.26 | 96.62 | 43.42 | 48.17 | 3555.68 | 7658.65 |
LV0007-CAR-T+K562 | 1652.22 | 18.65 | 5 | 279.96 | 10.6 | 1438.33 | 19.54 |
T | 7.03 | N/A | 8.17 | 8.35 | 3.82 | 171.96 | 315.4 |
T+U226 | N/A | N/A | N/A | 5.29 | 2.47 | 21.85 | 83.8 |
T+K562 | N/A | N/A | N/A | 55.54 | N/A | N/A | 123.2 |
Claims (30)
- 嵌合抗原受体(CAR),其中所述CAR包含BCMA结合结构域、跨膜结构域、共刺激结构域和胞内信号传导结构域,所述BCMA结合结构域包含特异性结合BCMA的抗体或其片段,其中所述抗体包含重链互补决定区1(HCDR1),重链互补决定区2(HCDR2)和重链互补决定区3(HCDR3),所述HCDR1的氨基酸序列如SEQ ID NO:9所示,所述HCDR2的氨基酸序列如SEQ ID NO:10所示且所述HCDR3的氨基酸序列如SEQ ID NO:11所示。
- 根据权利要求1所述的CAR,其中所述抗体包含轻链互补决定区1(LCDR1),轻链互补决定区2(LCDR2)和轻链互补决定区3(LCDR3),所述LCDR1的氨基酸序列如SEQ ID NO:17所示,所述LCDR2的氨基酸序列如SEQ ID NO:18所示且所述LCDR3的氨基酸序列如SEQ ID NO:19所示。
- 根据权利要求1-2中任一项所述的CAR,其中所述抗体包含重链可变区,所述重链可变区的氨基酸序列如SEQ ID NO:7所示。
- 根据权利要求1-3中任一项所述的CAR,其中所述抗体包含轻链可变区,所述轻链可变区的氨基酸序列如SEQ ID NO:15所示。
- 根据权利要求1-4中任一项所述的CAR,其中所述抗体为单链抗体。
- 根据权利要求1-5中任一项所述的CAR,其中所述抗体包含SEQ ID NO:43所示的氨基酸序列。
- 根据权利要求1-6中任一项所述的CAR,其中所述跨膜结构域包含源自选自下述蛋白的跨膜结构域:T细胞受体的α,β或ζ链、CD28、CD3e、CD45、CD4、CD5、CD8a、CD9、CD16、CD22、CD33、CD37、CD64、CD80、CD86、CD134、CD137和CD154。
- 根据权利要求1-7中任一项所述的CAR,其中所述跨膜结构域包含SEQ ID NO:27所示的氨基酸序列。
- 根据权利要求1-8中任一项所述的CAR,其中所述共刺激结构域包含源自选自下述蛋白的共刺激结构域:CD28、4-1BB、OX-40和ICOS。
- 根据权利要求1-9中任一项所述的CAR,其中所述共刺激结构域包含SEQ ID NO:29或SEQ ID NO:31所示的氨基酸序列。
- 根据权利要求1-10中任一项所述的CAR,其中所述胞内信号传导结构域包含源自CD3ζ的信号传导结构域。
- 根据权利要求1-11中任一项所述的CAR,其中所述胞内信号传导结构域包含SEQ ID NO:33所示的氨基酸序列。
- 根据权利要求1-12中任一项所述的CAR,其中所述CAR还包含铰链区,所述铰链 区连接所述BCMA结合结构域和所述跨膜结构域。
- 根据权利要求13所述的CAR,其中所述铰链区包含SEQ ID NO:25所示的氨基酸序列。
- 根据权利要求1-14中任一项所述的CAR,其中所述CAR还连接信号肽。
- 根据权利要求15所述的CAR,其中所述信号肽包含SEQ ID NO:3所示的氨基酸序列。
- 根据权利要求1-16中任一项所述的CAR,其中所述CAR还连接剪切肽。
- 根据权利要求17所述的CAR,其中所述剪切肽包含来自T2A肽的氨基酸序列。
- 根据权利要求17-18中任一项所述的CAR,其中所述剪切肽包含SEQ ID NO:35所示的氨基酸序列。
- 根据权利要求1-19中任一项所述的CAR,其包含SEQ ID NO:49或SEQ ID NO:51所示的氨基酸序列。
- 分离的核酸分子,其编码权利要求1-20中任一项所述的CAR。
- 编码CAR的分离的核酸分子,其包含SEQ ID NO:50或SEQ ID NO:52所示的核酸序列。
- 一种载体,其包含权利要求21-22中任一项所述的核酸分子。
- 根据权利要求23所述的载体,其中所述载体选自质粒、逆转录病毒载体和慢病毒载体。
- 一种免疫效应细胞,其包含权利要求1-20中任一项所述的CAR,权利要求21-22中任一项所述的核酸分子,或权利要求23-24中任一项所述的载体。
- 根据权利要求25所述的细胞,其中所述免疫效应细胞选自T淋巴细胞和自然杀伤(NK)细胞。
- 一种制备免疫效应细胞的方法,其包括向免疫效应细胞中引入权利要求26所述的载体。
- 组合物,其包含权利要求25-26中任一项所述的免疫效应细胞。
- 权利要求1-20中任一项所述的CAR,权利要求21-22中任一项所述的核酸分子,权利要求23-24所述的载体,或权利要求26-27中任一项所述的免疫效应细胞用于制备药物的用途,其中所述药物用于治疗与BCMA的表达相关的疾病或病症。
- 根据权利要求29所述的用途,其中所述与BCMA的表达相关的疾病或病症为癌症或恶性肿瘤。
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US16/650,327 US11026975B2 (en) | 2018-02-01 | 2019-01-31 | Chimeric antigen receptor (CAR) binding to BCMA, and uses thereof |
CA3074526A CA3074526C (en) | 2018-02-01 | 2019-01-31 | Chimeric antigen receptor (car) binding to bcma and application thereof |
JP2020518469A JP6968389B2 (ja) | 2018-02-01 | 2019-01-31 | Bcmaに結合するキメラ抗原受容体(car)及びその応用 |
KR1020207008556A KR102160061B1 (ko) | 2018-02-01 | 2019-01-31 | Bcma에 결합하는 키메릭 항원 수용체(car) 및 그의 용도 |
AU2019214163A AU2019214163B2 (en) | 2018-02-01 | 2019-01-31 | Chimeric antigen receptor (CAR) binding to BCMA, and uses thereof |
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Cited By (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2020018825A1 (en) * | 2018-07-19 | 2020-01-23 | Regeneron Pharmaceuticals, Inc. | Chimeric antigen receptors with bcma specificity and uses thereof |
US11384153B2 (en) | 2018-07-19 | 2022-07-12 | Regeneran Pharmaceuticals, Inc. | Bispecific anti-BCMA x anti-CD3 antibodies and uses thereof |
WO2023044633A1 (zh) * | 2021-09-22 | 2023-03-30 | 南京驯鹿医疗技术有限公司 | Bcma car-t在制备用于治疗自身免疫病的药物中的应用 |
Families Citing this family (23)
Publication number | Priority date | Publication date | Assignee | Title |
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CA3200509A1 (en) | 2020-12-31 | 2022-07-07 | Sonja SCHREPFER | Methods and compositions for modulating car-t activity |
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WO2024119157A1 (en) | 2022-12-02 | 2024-06-06 | Sana Biotechnology, Inc. | Lipid particles with cofusogens and methods of producing and using the same |
Citations (8)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2013154760A1 (en) | 2012-04-11 | 2013-10-17 | The United States Of America, As Represented By The Secretary, Department Of Health And Human Services | Chimeric antigen receptors targeting b-cell maturation antigen |
US8802374B2 (en) | 2009-11-03 | 2014-08-12 | City Of Hope | Truncated epiderimal growth factor receptor (EGFRt) for transduced T cell selection |
US9034324B2 (en) | 2009-03-10 | 2015-05-19 | Biogen Idec Ma Inc. | Anti-BCMA antibodies |
WO2016014789A2 (en) | 2014-07-24 | 2016-01-28 | Bluebird Bio, Inc. | Bcma chimeric antigen receptors |
CN105777911A (zh) * | 2016-04-12 | 2016-07-20 | 上海优卡迪生物医药科技有限公司 | 抗bcma嵌合抗原受体、编码基因、重组表达载体及其构建方法和应用 |
CN105837693A (zh) * | 2016-05-30 | 2016-08-10 | 李斯文 | 一种基于bcma的抗原嵌合受体及其制备方法和应用 |
CN106687483A (zh) * | 2014-07-21 | 2017-05-17 | 诺华股份有限公司 | 使用人源化抗‑bcma嵌合抗原受体治疗癌症 |
CN107207598A (zh) * | 2014-12-12 | 2017-09-26 | 蓝鸟生物公司 | Bcma嵌合抗原受体 |
Family Cites Families (12)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CA2706300C (en) * | 2007-11-29 | 2017-05-09 | Cytos Biotechnology Ag | Human monoclonal nicotine specific antibodies |
PL3415531T3 (pl) * | 2011-05-27 | 2024-02-26 | Glaxo Group Limited | Białka wiążące antygen |
TW201425336A (zh) * | 2012-12-07 | 2014-07-01 | Amgen Inc | Bcma抗原結合蛋白質 |
GB201317929D0 (en) * | 2013-10-10 | 2013-11-27 | Ucl Business Plc | Chimeric antigen receptor |
EP4310097A3 (en) * | 2014-12-05 | 2024-04-03 | Memorial Sloan Kettering Cancer Center | Chimeric antigen receptors targeting b-cell maturation antigen and uses thereof |
US10294304B2 (en) * | 2015-04-13 | 2019-05-21 | Pfizer Inc. | Chimeric antigen receptors targeting B-cell maturation antigen |
PL3298033T5 (pl) * | 2015-05-18 | 2023-10-30 | TCR2 Therapeutics Inc. | Kompozycje i zastosowania medyczne do reprogramowania TCR z zastosowaniem białek fuzyjnych |
MX2018010733A (es) * | 2016-03-04 | 2019-07-04 | Novartis Ag | Celulas que expresan multiples moleculas del receptor de antigeno quimerico (car) y usos de las mismas. |
IL312206A (en) | 2016-04-15 | 2024-06-01 | Novartis Ag | Preparations and methods for selective protein expression |
AU2017276706B2 (en) * | 2016-06-07 | 2024-08-15 | Max-Delbrück-Centrum Für Molekulare Medizin In Der Helmholtz-Gemeinschaft | Chimeric antigen receptor and CAR-T cells that bind BCMA |
US11365252B2 (en) * | 2016-07-20 | 2022-06-21 | University Of Utah Research Foundation | CD229 CAR T cells and methods of use thereof |
CN111601825B (zh) * | 2018-02-01 | 2022-11-29 | 信达生物制药(苏州)有限公司 | 全人源的抗b细胞成熟抗原(bcma)单链抗体及其应用 |
-
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Patent Citations (9)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US9034324B2 (en) | 2009-03-10 | 2015-05-19 | Biogen Idec Ma Inc. | Anti-BCMA antibodies |
US8802374B2 (en) | 2009-11-03 | 2014-08-12 | City Of Hope | Truncated epiderimal growth factor receptor (EGFRt) for transduced T cell selection |
WO2013154760A1 (en) | 2012-04-11 | 2013-10-17 | The United States Of America, As Represented By The Secretary, Department Of Health And Human Services | Chimeric antigen receptors targeting b-cell maturation antigen |
CN104379179A (zh) * | 2012-04-11 | 2015-02-25 | 美国卫生和人力服务部 | 靶向b-细胞成熟抗原的嵌合抗原受体 |
CN106687483A (zh) * | 2014-07-21 | 2017-05-17 | 诺华股份有限公司 | 使用人源化抗‑bcma嵌合抗原受体治疗癌症 |
WO2016014789A2 (en) | 2014-07-24 | 2016-01-28 | Bluebird Bio, Inc. | Bcma chimeric antigen receptors |
CN107207598A (zh) * | 2014-12-12 | 2017-09-26 | 蓝鸟生物公司 | Bcma嵌合抗原受体 |
CN105777911A (zh) * | 2016-04-12 | 2016-07-20 | 上海优卡迪生物医药科技有限公司 | 抗bcma嵌合抗原受体、编码基因、重组表达载体及其构建方法和应用 |
CN105837693A (zh) * | 2016-05-30 | 2016-08-10 | 李斯文 | 一种基于bcma的抗原嵌合受体及其制备方法和应用 |
Non-Patent Citations (22)
Title |
---|
"Current Protocols in Immunology", 1991 |
"GenBank", Database accession no. BAB60895.1 |
ANAND: "Techniques for the Analysis of Complex Genomes", 1992, ACADEMIC PRESS |
AUSUBEL ET AL.: "Current Protocols in Molecular Biology", 1993, GREENE PUBLISHING AND WILEY-INTERSCIENCE |
BLOOD, vol. 112, 2008, pages 2261 - 2271 |
CHENG ZHANGJUN LIUJIANG F ZHONG ET AL.: "Engineering CAR-T cells", BIOMARKER RESEARCH, vol. 5, 2017, pages 22, XP055551810, DOI: 10.1186/s40364-017-0102-y |
D. J. LIPMANW. R. PEARSON: "Rapid and Sensitive Protein Similarity Searches", SCIENCE, vol. 227, 1989, pages 1435 - 1441 |
DE ANGIOLETTI M ET AL.: "a novel silent beta-thalassaemia mutation, the first in the Kozak sequence", BR J HAEMATOL., vol. 124, no. 2, 2004, pages 224 - 31 |
DOTTI GGOTTSCHALK SSAVOLDO BBRENNER MK: "Design and development of therapies using chimeric antigen receptor-expressing T cells", IMMUNOL REV., vol. 257, no. 1, 2014, pages 107 - 126, XP055552726, DOI: 10.1111/imr.12131 |
FLATMAN, S. ET AL., J.CHROM.B, vol. 848, 2007, pages 79 - 87 |
GLOVER: "Short Protocols in Molecular Biology: A Compendium of Methods from Current Protocols in Molecular Biology", vol. I & II, 1985, GREENE PUB.ASSOCIATES AND WILEY-INTERSCIENCE |
HARLOWLANE: "Antibodies", 1998, COLD SPRING HARBOR LABORATORY PRESS |
MANIATIS ET AL., MOLECULAR CLONING: A LABORATORY MANUAL, 1982 |
MOLECULAR THERAPY, vol. 18, no. 4, 2010, pages 666 - 668 |
PERBAL: "A Practical Guide to Molecular Cloning", 1984 |
RICKERT ET AL., IMMUNOLOGICAL REVIEWS, vol. 244, 2011, pages 115 - 133 |
S. ALTSCHULW. GISHW. MILLERE. W. MYERSD. LIPMAN: "A Basic Local Alignment Search Tool", JOURNAL OF MOLECULAR BIOLOGY, vol. 215, 1990, pages 403 - 410 |
SADELAIN MBRENTJENS RRIVI 'ERE I: "The basic principles of chimeric antigen receptor design", CANCER DISCOV., vol. 3, no. 4, 2013, pages 388 - 398, XP055287277, DOI: 10.1158/2159-8290.CD-12-0548 |
SCHEMBRI, LAURA ET AL.: "The HA tag is cleaved and loses immunoreactivity during apoptosis", NATURE METHODS, vol. 4, no. 2, February 2007 (2007-02-01), pages 107 - 108 |
SZYMCZAK ET AL.: "correction of multi-gene deficiency in vivo using a single self-cleaving 2A peptide-based retroviral vector", NATURE BIOTECHNOLOGY, vol. 22, 2004, pages 589, XP055564203, DOI: 10.1038/nbt957 |
TURTLE CJHUDECEK MJENSEN MCRIDDELL SR: "Engineered T cells for anticancer therapy", CURR OPIN IMMUNOL., vol. 24, no. 5, 2012, pages 633 - 639 |
W. R. PEARSOND. J. LIPMAN: "Improved Tools for Biological Sequence Comparison", PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA (PROC. NATL. ACAD. SCI., vol. 85, 1988, pages 2444 - 2448 |
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Publication number | Priority date | Publication date | Assignee | Title |
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WO2020018825A1 (en) * | 2018-07-19 | 2020-01-23 | Regeneron Pharmaceuticals, Inc. | Chimeric antigen receptors with bcma specificity and uses thereof |
US11384153B2 (en) | 2018-07-19 | 2022-07-12 | Regeneran Pharmaceuticals, Inc. | Bispecific anti-BCMA x anti-CD3 antibodies and uses thereof |
EP4324851A3 (en) * | 2018-07-19 | 2024-05-15 | Regeneron Pharmaceuticals, Inc. | Chimeric antigen receptors with bcma specificity and uses thereof |
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