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Long‐term safety and activity of NY-ESO‐1 SPEAR T cells after autologous stem cell transplant for myeloma

Edward A. Stadtmauer, Thomas H. Faitg, Daniel E. Lowther, Ashraf Z. Badros, Karen Chagin, Karen Dengel, Malini Iyengar, Luca Melchiori, Jean-Marc Navenot, Elliot Norry, Trupti Trivedi, Ruoxi Wang, Gwendolyn K. Binder, Rafael Amado and Aaron P. Rapoport

Data supplements

Article Figures & Data

Figures

  • Figure 1.

    Clinical responses (ITT population). (A) Patient response profiles. Swimmer plot showing duration of clinical response and survival after disease progression. Patients 207, 256, and 264 were disease progression-free at more than 3 years post-NY-ESO-1c259 T-cell therapy. Day 0, ASCT; day 2, NY-ESO-1c259 T-cell infusion. (B) PFS, Kaplan-Meier plot. Surviving (censored) patients are represented by tick marks. (C) OS, Kaplan-Meier plot. Surviving (censored) patients are represented by tick marks.

  • Figure 2.

    Persistence of NY-ESO-1 SPEAR T cells in the peripheral blood and bone marrow. (A) Persistence of NY-ESO-1 SPEAR T cells in PBMC through 1 year after T-cell infusion measured by quantitative polymerase chain reaction methodology is shown for responders (left) and nonresponders (right). Note: Responder/nonresponder status based on BOR up to year 1. (Assumptions: average of 1 vector copy per cell based on transduction efficiency; 1 μg DNA = ∼158 000 cells. Lower limit of quantification of the assay was established at 50 copies/μg DNA at the University of Pennsylvania, and 100 copies/μg DNA at Cambridge Biomedical.) (B) Persistence of NY-ESO-1 SPEAR T-cells in PBMC through 100 days after T-cell infusion was confirmed via flow cytometry measurements. Box plot shows the mean, upper, and lower quartiles. (C) Persistence of NY-ESO-1 SPEAR T cells was compared in PBMC and bone marrow (BM) from patients with paired samples, and is shown as a percentage of the total CD4+ (or CD8+) T cells. Box plot shows the mean, upper, and lower quartiles. (D) NY-ESO-1 SPEAR T-cell proliferation in the BM and PBMC was demonstrated by Ki-67 staining. Box plot shows the mean, upper, and lower quartiles. Day 21 was selected because of patient numbers (evaluable BM samples, n = 8) and robust data at this time. Note: the pentamer is less efficient for CD4+ T-cell binding and, therefore, may underestimate this compartment.

  • Figure 3.

    Memory phenotypes of NY-ESO-1 SPEAR T cells. (A) Distribution of memory subsets with described phenotypic markers of CM, EM, EM RA+ (EMRA), stem cell memory SCM, and naïve within CD8+ pentamer+ NY-ESO-1 SPEAR T cells in BM and PBMC postinfusion were performed in patients in whom parallel collection of BM and PBMC was carried out and in whom sufficient cells were available for analysis (n = 8). Pie chart colors represent the proportions of NY‐ESO‐1 SPEAR T‐cells with described phenotypic markers of CM (red), EM (blue), EMRA (green), SCM (orange), and naïve cells (purple) within CD8+ SPEAR T‐cells. (B) Percentage of NY-ESO-1 SPEAR T cells with described phenotypic markers of CM, EM, EMRA, and SCM within CD8+ pentamer+ NY-ESO-1 SPEAR T cells in the manufactured product and posttreatment PBMC at various times from all 25 patients. The dominant populations of persisting cells were EMRA (green) and SCM (orange). Box plot shows the mean and upper and lower quartiles.

  • Figure 4.

    Functionality of NY-ESO-1 SPEAR T cells postinfusion. (A) Cytokine production: cytokine levels per cell were measured by median fluorescent intensity. Cytokines interferon γ, interleukin 2, and tumor necrosis factor α were measured by intracellular cytokine staining after antigen-specific stimulation. Data were measured in 20 of 25 patients. Line graph shows the mean and standard deviation. Median fluorescent intensity was measured from the cytokine-positive population only (ie, median fluorescent intensity values ≥2000). (B) Functional characterization by antigen-specific cytokine responses: NY-ESO-1 SPEAR T cells produced cytokine in response to antigen stimulation, the level of which correlated with the transduction percentage. Before manufacturing, and therefore, in the absence of NY-ESO-1 SPEAR T cells (ie, day –50), there was no measurable response to antigen stimulation. Transduction efficiency of manufactured product, median (range): CD8+ T cells 18.85% (5.94%-46.5%), CD4+ T cells 14.83% (2.46%-44.83%). Data were measured in 22 of 25 patients. Day –50 = baseline peripheral blood sample. Box plot shows the mean and upper and lower quartiles. (C) Polyfunctionality: the percentage of CD4+ T cells and CD8+ T cells expressing interferon γ, interleukin 2, or tumor necrosis factor α; a combination of any 2 of the 3 cytokines; or all 3 cytokines in response to ex vivo stimulation with antigen-pulsed T2 cells were evaluated in paired PBMC and BM samples from a subset of patients (n = 11). The percentage of cytokine-positive cells producing 1 or more cytokines is displayed as a percentage of all cytokine-producing cells. Box plot shows the mean and upper and lower quartiles. (D) Expression of exhaustion markers: the percentage of CD4+ pentamer+ T cells or CD8+ pentamer+ T cells were evaluated for expression of 3 exhaustion markers (LAG3, PD1, and TIM3) in paired PBMC and BM samples from a subset of patients (n = 11). Line graph shows the mean and standard deviation. MP, manufactured product; PMA-IONO, phorbol myristate acetate + ionomycin (positive control); T2, T2 cells unpulsed (negative control); T2-NY-ESO, T2 cells pulsed with NY-ESO-1 peptide.

  • Figure 5.

    TCR clonality of T-cell response postinfusion. TCR clonality was assessed via Pielou’s Evenness Index (upper) and Shannon’s Diversity Index (lower). These indices were calculated using all TCR sequences evaluated for preinfusion manufactured product and the corresponding BM samples taken at day 100 postinfusion, and are shown for all patients in whom both baseline and day 100 samples were collected. Day 100 samples were not available for 2 patients, for whom the sample collected at the point closest to day 100 was analyzed.

Tables

  • Table 1.

    Patient disposition

    Patients (N = 25)
    ITT population, n25
    mITT population, n25
    Patient status at end of infusion 1 interventional phase, n (%)*25 (100)
     Disease progression22 (88)
     Did not progress3 (12)
    Patient status at end of infusion 2 interventional phase (for patients with second infusion), n (%)*4 (16)
     Disease progression4 (16)
    • Patients completed the study if they discontinued because of disease progression or death after their last T-cell infusion, or if they were progression-free at the time of study completion (at Sponsor discretion) and moved into long-term follow-up. ITT population: all patients who were enrolled in the trial. mITT population: all patients in the ITT population who received ≥ 1 T-cell infusion.

    • * Denominator for percentages is based on the ITT population.

    • Did not progress by the time of the primary analysis; participation was terminated by the study sponsor.

    • One patient progressed before withdrawing consent.

  • Table 2.

    Patient demographic and baseline characteristics

    ParameterPatients (N = 25)
    Sex, n (%)
     Female10 (40)
     Male15 (60)
    Age, y*
     Mean (SD)59 (8)
     Median59
     Minimum, maximum45, 72
    Race, n (%)
     Black or African American5 (20)
     White19 (76)
     Multiracial1 (4)
    HLA status, n (%)
     HLA-A2+25 (100)
    NY-ESO-1 status, n (%)
     NY-ESO-1+14 (56)
     NY-ESO-111 (44)
    LAGE-1 status, n (%)
     LAGE-1+25 (100)
    ECOG performance status, n (%)
     07 (28)
     116 (64)
     22 (8)
    Previous therapies, n (%)
     15 (20)
     26 (24)
     34 (16)
     >410 (40)
    Cytogenetic abnormalities, n (%)
     ≥112 (48)
     Deletion (13q)8 (32)
     Deletion (17p)1 (5)
     Hypodiploid0 (0)
    Transduced cell dose, n (%)
     <1 × 1093 (12)
     1-5 × 10921 (84)
     ≥5 × 1091 (4)
     Median (range)3.1 × 109 (0.5-5.05 × 109)
    • ECOG, Eastern Cooperative Oncology Group; SD, standard deviation.

    • * Age at informed consent/screening.

    • One patient tested positive for LAGE-1 per the laboratory report at screening but was erroneously recorded as LAGE-1 negative in the database, and this was not corrected prior to database lock.

    • Four patients had received a prior autologous stem cell transplant.

  • Table 3.

    Overall summary of adverse events (ITT population)

    CategoryPatients (N = 25), n (%)
    Adverse events25 (100)
     Any AE related* to study intervention24 (96)
     Any AE ≥ grade 324 (96)
     AE grade 3 hematologic21 (84)
     AE grade 3 nonhematologic23 (92)
     AE grade 4 hematologic23 (92)
     AE grade 4 nonhematologic7 (28)
     Any AE related* to study intervention and ≥grade 321 (84)
    Serious adverse events13 (52)
     Any SAE related* to study intervention7 (28)
     Any SAE ≥ grade 312 (48)
     Any SAE related* to study intervention and ≥grade 37 (28)
     Any SAEs with fatal outcome0
    • Observation period: administration of high-dose melphalan (day –2) to end of intervention phase. AEs were coded using MedDRA Version 20.0.

    • SAE, serious AE.

    • * Defined (by the investigator) as definitely related, probably related, possibly related, or unlikely related to T-cell infusion.

    • Hematologic AEs were defined as blood system disorders or investigations involving an increase/decrease in blood cells; nonhematologic AEs were defined as all other system/organ class terms.

  • Table 4.

    Hematologic and nonhematologic adverse events occurring at toxicity grade 3 or 4 in ≥5% of patients by preferred term (ITT population)

    Preferred term*Patients (N = 25), n (%)
    Grade 3Grade 4
    Hematologic toxicities
     Febrile neutropenia15 (60)2 (8)
     Anemia14 (56)0 (0)
     Neutropenia4 (16)7 (28)
     Leukopenia1 (4)13 (52)
     Lymphopenia0 (0)2 (8)
     Thrombocytopenia0 (0)18 (72)
    Nonhematologic toxicities
     Diarrhea10 (40)0 (0)
     Hypophosphatemia6 (24)0 (0)
     Hypocalcemia4 (16)1 (4)
     Graft-versus-host disease3 (12)0 (0)
     Hypokalemia3 (12)0 (0)
     Rash3 (12)0 (0)
     Stomatitis3 (12)0 (0)
     Abdominal pain2 (8)0 (0)
     Fatigue2 (8)0 (0)
     Hyponatremia2 (8)0 (0)
     Hypoxia2 (8)0 (0)
     Mucosal inflammation2 (8)0 (0)
     Neutropenic colitis2 (8)0 (0)
     Nausea2 (8)0 (0)
     Esophagitis2 (8)0 (0)
     Pain in extremity2 (8)0 (0)
     Atrial fibrillation1 (4)1 (4)
     Hypotension1 (4)1 (4)
     Pneumonitis1 (4)1 (4)
    • Patients were counted once for each preferred term.

    • Observation period: administration of high-dose melphalan at day –2 to end of intervention phase. AEs were coded using MedDRA Version 20.0. Definition of hematologic/nonhematologic AE terms are provided in Table 3.

    • * AE data are listed in descending order of frequency of grade 3 events.

  • Table 5.

    ORR per IMWG 2011 criteria by time points of interest (ITT population)

    Time pointORR (N = 25), n (%)95% Clopper-Pearson CI
    Day 4220 (80)0.59-0.93
    Day 100*19 (76)0.55-0.91
    Day 18016 (64)0.43-0.82
    Day 27013 (52)0.31-0.72
    Year 111 (44)0.24-0.65
    • ORR is defined as the proportion of patients who have a positive response by IMWG 2011 criteria (sCR, CR, VGPR, PR) relative to the total number of patients in the population.

    • CR, complete response; PR, partial response; sCR, stringent complete response; VGPR, very good partial response.

    • * Day 100 responders (n = 19): 1 sCR, 12 VGPR, and 6 PR.

    • Year 1 responders (n = 11): 1 sCR, 1 CR, 8 VGPR, and 1 PR.

  • Table 6.

    BOR by time (ITT population)

    Time and parameter/category or criterion, IMWG 2011 response, n (%)Patients (N = 25), n (%)
    Day 42
     sCR1 (4)
     CR0
     VGPR10 (40)
     PR9 (36)
     SD5 (20)
     PD0
    Day 100
     sCR1 (4)
     CR0
     VGPR12 (48)
     PR6 (24)
     SD5 (20)
     PD1 (4)
    Day 180
     sCR2 (8)
     CR1 (4)
     VGPR11 (44)
     PR2 (8)
     SD3 (12)
     PD2 (8)
    Day 270
     sCR2 (8)
     CR1 (4)
     VGPR8 (32)
     PR2 (8)
     SD2 (8)
     PD4 (16)
    Year 1
     sCR1 (4)
     CR1 (4)
     VGPR8 (32)
     PR1 (4)
     SD2 (8)
     PD1 (4)
    • Patients without a response evaluated at the visit are included in the denominator.

    • BOR, best objective response; PD, progressive disease or relapse; SD, stable disease.