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Clinical Trial
. 2016 Sep 29;128(13):1688-700.
doi: 10.1182/blood-2016-04-711903. Epub 2016 Jul 13.

T cells expressing an anti-B-cell maturation antigen chimeric antigen receptor cause remissions of multiple myeloma

Syed Abbas Ali  1 , Victoria Shi  2 , Irina Maric  3 , Michael Wang  4 , David F Stroncek  5 , Jeremy J Rose  2 , Jennifer N Brudno  1 , Maryalice Stetler-Stevenson  6 , Steven A Feldman  7 , Brenna G Hansen  2 , Vicki S Fellowes  2 , Frances T Hakim  2 , Ronald E Gress  2 , James N Kochenderfer  2
Affiliations
Clinical Trial

T cells expressing an anti-B-cell maturation antigen chimeric antigen receptor cause remissions of multiple myeloma

Syed Abbas Ali (V体育官网入口) et al. Blood. .

"VSports最新版本" Abstract

Therapies with novel mechanisms of action are needed for multiple myeloma (MM). B-cell maturation antigen (BCMA) is expressed in most cases of MM. We conducted the first-in-humans clinical trial of chimeric antigen receptor (CAR) T cells targeting BCMA. T cells expressing the CAR used in this work (CAR-BCMA) specifically recognized BCMA-expressing cells. Twelve patients received CAR-BCMA T cells in this dose-escalation trial. Among the 6 patients treated on the lowest 2 dose levels, limited antimyeloma activity and mild toxicity occurred VSports手机版. On the third dose level, 1 patient obtained a very good partial remission. Two patients were treated on the fourth dose level of 9 × 10(6) CAR(+) T cells/kg body weight. Before treatment, the first patient on the fourth dose level had chemotherapy-resistant MM, making up 90% of bone marrow cells. After treatment, bone marrow plasma cells became undetectable by flow cytometry, and the patient's MM entered a stringent complete remission that lasted for 17 weeks before relapse. The second patient on the fourth dose level had chemotherapy-resistant MM making up 80% of bone marrow cells before treatment. Twenty-eight weeks after this patient received CAR-BCMA T cells, bone marrow plasma cells were undetectable by flow cytometry, and the serum monoclonal protein had decreased by >95%. This patient is in an ongoing very good partial remission. Both patients treated on the fourth dose level had toxicity consistent with cytokine-release syndrome including fever, hypotension, and dyspnea. Both patients had prolonged cytopenias. Our findings demonstrate antimyeloma activity of CAR-BCMA T cells. This trial was registered to www. clinicaltrials. gov as #NCT02215967. .

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"VSports在线直播" Figures

Figure 1
Figure 1
CAR-BCMA T cells specifically recognized BCMA in vitro and exhibited antimyeloma activity in humans. (A) Diagram of the MSGV-11D5-3-CD828Z γ-retroviral vector encoding the anti-BCMA CAR (CAR-BCMA) is shown. CAR-BCMA contained the 11D5-3 anti-BCMA single chain variable fragment (scFv), CD8α hinge and transmembrane regions, the cytoplasmic portion of the CD28 costimulatory moiety, and the CD3ζ T-cell activation domain. (B) CAR-BCMA expression on the surface of the infusion T cells of patient 10 was detected by staining with a PE-BCMA-Fc protein reagent. The plot is gated on live CD3+ lymphocytes. (C) Demonstrating specificity, the PE-BCMA-Fc reagent did not stain PBMCs that were not transduced with the CAR-BCMA gene. (D) CAR-BCMA T cells of patient 10 were cultured with target cells that either expressed BCMA (BCMA-K562 and RPMI8226) or did not express BCMA (NGFR-K562 and CCRF-CEM). The CAR-BCMA T cells of patient 10 specifically released IFNγ in response to BCMA-expressing target cells in vitro. (E) Patient 8 who had MM that was progressing despite 8 prior lines of therapy obtained a very good partial remission (VGPR) after infusion of CAR-BCMA T cells. Positron emission tomography–computed tomography scans from before and after treatment show elimination of a large number of MM bone lesions. (F) Patient 8’s free κ light chains level decreased after CAR T-cell infusion as the MM entered a VGPR and then increased with progression of the MM. The serum BCMA protein level followed a pattern similar to that of the serum free κ light chains.
Figure 2
Figure 2
Patient 10 entered a stringent complete remission. (A) Before treatment, patient 10’s bone marrow was stained for BCMA by IHC. BCMA expression was uniform but dim on the MM cells (500×). (B) Flow cytometry of patient 10’s bone marrow showed dim but uniform BCMA expression on the CD19-negative malignant plasma cells (red), whereas BCMA expression was absent on CD19+ B lymphocytes (black). The plot shows a combination of the plasma cell gate (CD38bright, CD138+, CD45dim-negative) and the mature B-cell gate (CD45+, CD19+ lymphocytes). (C) Four hours after CAR-BCMA infusion, patient 10 became febrile. He was febrile for 7 days. The plot shows the maximum temperature for each day. The CRP was elevated. (D) Before initiation of protocol treatment, patient 10 had a hypercellular bone marrow (hematoxylin and eosin). Bone marrow cells were ≥90% plasma cells as shown by CD138 staining. BCMA expression was dim but present. (E) Four weeks after CAR T-cell infusion, the bone marrow was hypocellular, and bone marrow plasma cells were completely absent as shown by the negative CD138 and BCMA staining. (F) Eight weeks after CAR T-cell infusion, the bone marrow was recovering with an overall cellularity of 25%, and plasma cells remained absent. Magnification for D-F was 40×. (G) Immediately before CAR-BCMA infusion, the patient had a serum IgA level of 2633 mg/dL. After CAR T-cell infusion on day 0, serum IgA decreased until reaching undetectable levels. (H) IHC staining of patient 10’s bone marrow showed an infiltrate of CD3+ T cells 4 weeks after CAR T-cell infusion. (I) Flow cytometry with a PE-BCMA-Fc reagent showed that 31% of bone marrow T cells were CAR-BCMA T cells 4 weeks after CAR T-cell infusion. The plot is gated on CD45+ and CD3+ lymphocytes.
Figure 3
Figure 3
MM was eliminated from the bone marrow of patient 11 after CAR-BCMA infusion. (A) Before treatment, BCMA expression was uniform and strong on the MM cells of patient 11 (500×). (B) Flow cytometry showed strong uniform BCMA expression on CD19-negative malignant plasma cells (red), whereas BCMA expression was absent on CD19+ B lymphocytes (black). The plot shows a combination of the plasma cell gate (CD38bright, CD138+, CD45dim-negative) and the mature B-cell gate (CD45+, CD19+ lymphocytes). (C) Four hours after CAR-BCMA infusion, patient 11 became febrile. He was febrile until 5 days after the CAR T-cell infusion. The plot shows the maximum temperature for each day. The CRP was also elevated. (D) Before protocol treatment, patient 11 had a hypercellular bone marrow (hematoxylin and eosin ). Bone marrow cells were 80% plasma cells as shown by CD138 staining. (E) Four weeks after CAR T-cell infusion, the bone marrow was hypocellular, and bone marrow plasma cells were completely absent as shown by negative CD138 and BCMA staining. (F) Eight weeks after CAR T-cell infusion, the marrow was recovering with an overall cellularity of 20%; plasma cells remained absent. Magnification for D-F was 40×. (G) Before protocol treatment, the patient’s serum λ free light chains level was 95.9 mg/dL. The λ free light chains decreased to an undetectable level after CAR-BCMA T-cell infusion. Before treatment, the patient had an IgG λ M-protein level of 3.7 g/dL. The M-protein decreased after treatment. (H) Patient 11’s bone marrow contained an infiltrate of CD3+ T cells 4 weeks after CAR T-cell infusion. (I) Flow cytometry with PE-BCMA-Fc showed that 19% of bone marrow T cells were CAR-BCMA T cells 4 weeks after CAR T-cell infusion. The plot is gated on CD45+, CD3+ lymphocytes.
Figure 4
Figure 4
Patients with the most impressive antimyeloma responses had the highest levels of blood CAR-BCMA+ cells. (A) CAR-BCMA+ PBMCs were quantified in the blood of patients by qPCR. Patients treated on the 2 lower dose levels had low levels of CAR-BCMA+ cells in their blood (left). Patients treated on the higher 2 dose levels had higher levels of CAR-BCMA+ cells in their blood (right). Patients 8, 10, and 11 had the best antimyeloma responses and the highest levels of blood CAR-BCMA+ cells. The results are presented as the absolute number of CAR+ PBMC/μL of blood. Pre stands for pretreatment. (B) IL-6 was measured in the serum of all patients. Patients 10 and 11 (right) had the most severe cytokine-release syndrome, and these patients also had the highest serum IL-6 levels. Serum IL-6 levels of the other patients are shown on the left. Note the different y-axis scales on the left and right graphs. (C) Five days after CAR-BCMA T cell infusion, serum levels of 15 cytokines were measured in the blood of patient 10 and patient 11. Results are expressed as fold increase over pretreatment levels. (D) Serum soluble BCMA of all patients was measured by ELISA pretreatment and posttreatment. The posttreatment serum samples were obtained 22 to 29 days after CAR T-cell infusion for all patients except patient 12. The posttreatment sample of patient 12 was obtained 13 days after CAR T-cell infusion. All patients had detectable serum BCMA. In the patients obtaining the most impressive antimyeloma responses, Patients 8, 10, and 11, serum BCMA decreased posttreatment (right); in contrast, serum BCMA did not decrease substantially posttreatment in the other patients (left).
Figure 5
Figure 5
CAR-BCMA T cells became more differentiated after infusion. For A-L, cells from patient 10 are shown. All plots are gated on live lymphocytes, and the numbers on the plots are the percentages of cells in the indicated regions of the plots. Staining for CAR-BCMA was performed with PE-BCMA-Fc. All postinfusion samples were PBMCs that were collected 16 days after CAR-BCMA T-cell infusion. (A) Cells from the time of infusion (infusion cells) were stained for CD3 and CAR-BCMA. (B) This plot is gated on the CD3+CAR+ infusion cells. (C) Postinfusion cells were stained for CD3 and CAR-BCMA. (D) This plot is gated on postinfusion CD3+CAR+ cells. CAR T cells after infusion were predominantly CD8+. (E) CCR7 and CD45RA expression of CD3+CD4+CAR+ infusion cells is shown. (F) CCR7 and CD45RA expression of CD3+CD8+CAR+ infusion cells is shown. (G) CCR7 and CD45RA expression of postinfusion blood CD3+CD4+CAR+ T cells. (H) CCR7 and CD45RA expression of postinfusion blood CD3+CD8+CAR+ T cells. (I) CD3+CD8+CAR+ infusion cell PD-1 and CD28 expression are shown. (J) PD-1 and CD28 expression is shown on postinfusion blood CD3+CD8+CAR+ T cells. (K) CD57 and CD28 expression on CD3+CD8+CAR+ infusion cells is shown. (L) CD57 and CD28 expression of postinfusion blood CD3+CD8+CAR+ T cells is shown. For M-P, results are shown for all 4 patients with sufficient blood CAR-BCMA+ T cells from 7 to 16 days after infusion for analysis. Gating and analysis were performed as described above in A-L. The results are all for live CD3+CAR+ cells. The symbols shown in M correspond to the same patients for M-P. (M) The CD8+ to CD4+ ratio of CAR-BCMA T cells increased for all 4 assessable patients between infusion and the time the postinfusion samples were collected. The percentage of CD8+ CAR-BCMA T cells with (N) EM or EMRA phenotypes, (O) PD-1 expression, and (P) CD57 expression increased in all 4 assessable patients between infusion and the time a postinfusion sample was collected.
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References

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