t cell expansion
Recently Published Documents


TOTAL DOCUMENTS

496
(FIVE YEARS 105)

H-INDEX

56
(FIVE YEARS 7)

2021 ◽  
Vol 13 (625) ◽  
Author(s):  
Qian Zhang ◽  
Morgan E. Hresko ◽  
Lora K. Picton ◽  
Leon Su ◽  
Michael J. Hollander ◽  
...  

Cell Reports ◽  
2021 ◽  
Vol 37 (10) ◽  
pp. 110083
Author(s):  
Hanfei Zhao ◽  
Ying Liu ◽  
Lixia Wang ◽  
Gang Jin ◽  
Xiaocui Zhao ◽  
...  

2021 ◽  
Vol 9 (12) ◽  
pp. e003339
Author(s):  
Huaishan Wang ◽  
Hui Chen ◽  
Shujing Liu ◽  
Jie Zhang ◽  
Hezhe Lu ◽  
...  

BackgroundGamma delta (γδ) T cells are attractive effector cells for cancer immunotherapy. Vδ2 T cells expanded by zoledronic acid (ZOL) are the most commonly used γδ T cells for adoptive cell therapy. However, adoptive transfer of the expanded Vδ2 T cells has limited clinical efficacy.MethodsWe developed a costimulation method for expansion of Vδ2 T cells in PBMCs by activating γδ T-cell receptor (γδTCR) and Toll-like receptor (TLR) 7/8 using isopentenyl pyrophosphate (IPP) and resiquimod, respectively, and tested the functional markers and antitumoral effects in vitro two-dimensional two-dimensional and three-dimensional spheroid models and in vivo models. Single-cell sequencing dataset analysis and reverse-phase protein array were employed for mechanistic studies.ResultsWe find that Vδ2 T cells expanded by IPP plus resiquimod showed significantly increased cytotoxicity to tumor cells with lower programmed cell death protein 1 (PD-1) expression than Vδ2 T cells expanded by IPP or ZOL. Mechanistically, the costimulation enhanced the activation of the phosphatidylinositol 3-kinase (PI3K)–protein kinase B (PKB/Akt)–the mammalian target of rapamycin (mTOR) pathway and the TLR7/8–MyD88 pathway. Resiquimod stimulated Vδ2 T-cell expansion in both antigen presenting cell dependent and independent manners. In addition, resiquimod decreased the number of adherent inhibitory antigen-presenting cells (APCs) and suppressed the inhibitory function of APCs by decreasing PD-L1 and cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) expression in these cells during in vitro Vδ2 T-cell expansion. Finally, we showed that human Vδ2 T cells can be expanded from PBMCs and spleen of humanized NSG mice using IPP plus resiquimod or ZOL, demonstrating that humanized mice are a promising preclinical model for studying human γδ T-cell development and function.ConclusionsVδ2 T cells expanded by IPP and resiquimod demonstrate improved anti-tumor function and have the potential to increase the efficacy of γδ T cell-based therapies.


Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 3874-3874
Author(s):  
Kirubel Gebre ◽  
Bradley Drumheller ◽  
Sophia Faude ◽  
Jane Wei ◽  
Jared McFerran ◽  
...  

Abstract Introduction CAR T-cell therapies are utilized to treat relapsed/refractory B-lymphoblastic leukemia (B-ALL), diffuse large B cell lymphoma, mantle cell lymphoma, and multiple myeloma. CAR T cell expansion kinetics after infusion impact response. However, these CAR T cell products vary in their target epitope and constituent molecules that makes it difficult to validate broad molecular or flow cytometric assays for use in the clinical setting. The lack of commercially available reagents also limits the ability of clinical laboratories to validate separate CAR T cell specific assays. We investigated the utility of common hematology laboratory parameters to measure CAR T cell expansion and response. Methods Clinical hematology laboratory parameters after infusion of autologous and allogeneic CAR T cell products directed were assessed. Concurrent CBC and cell population data (CPD) parameters from the Sysmex XN 3000 automated hematology analyzer were available in 82 patients. Absolute lymphocyte count (ALC) kinetics after infusion of autologous CD19, allogeneic CD19, CD22, CD33, allogeneic CD123-directed CAR T cell products were analyzed. Patients who received bone marrow transplant served as controls. CPD parameters included X (lateral light scatter-granularity), Y (fluorescence-nuclei acid content), Z (forward scatter-size), and their distribution widths WX, WY, WZ. Archived CellaVision cell morphology images from 118 patients who received CD19-directed CAR T cell products and 25 patients who received other CAR T cell products were analyzed. Response was determined from 1 month post-CAR bone marrow disease assessment. Results Absolute Lymphocyte Counts, lymphocyte morphology and cell population data after infusion of CAR T cell products CD19-CAR, UCAR19, and CD22-CAR all showed a distinct lymphocyte expansion phase post-infusion in responders (Figure 1A) that was absent in non-responders and controls(Figure 1B). ALC showed characteristic lag, expansion, and contraction phases in responders. CD19-CAR had a peak at day 8 while CD22-CAR and allogenic Universal(U)CAR19 had relatively delayed peak times occurring near day 15 (Figure 1B). The stem cell transplant control patients did not show ALC expansion in the first two weeks and instead showed normal lymphocyte regeneration that occurs in the third to fourth weeks. CD33-CAR and allogeneic UCAR123 non responders did not show ALC expansions. CAR T cell responders showed a distinct sequence of changes in lymphocyte morphology that was absent in non-responders and stem cell transplant controls (Figure 1C). This pattern was noted uniformly across various CAR T cell products (Figure 1D-1F). The morphological changes were categorized as: early, mid, and late. Early atypical cells were noted around days 4-8 after infusion and showed immunoblastic morphology that was present in 89% (n=105) of patients. Mid atypical cells were noted around days 5-14 after infusion and showed atypical large granular lymphocyte morphology that were seen in 95% (n=112) of patients. Late atypical cells showed the typical LGL morphology and was seen in 82% (n=97) of patients. WY fluorescence, which is a measure of nucleic acid content, was most useful in assessing changes in lymphocytes after CAR T cell infusion. WY was low (mean 402, n=57) at baseline pre-infusion timepoint and peaked (mean 1207) approximately 1 week after infusion (mean 7.9 days, n = 59). Peak WY was observed 3.7 days prior to peak ALC (mean day 11.6, n = 59). Conclusion We demonstrate for the first time that common clinical laboratory parameters can be used to follow CAR T cell expansion after infusion in various autologous and allogeneic CAR T cell products. ALC expansion is a measure of CAR T cell expansion after infusion and correlates with response. Responders showed higher peak ALC compared to non-responders in all CAR T cell products. Timing of peak ALC expansion was determined by other factors such as expression of target antigen and CAR T-cell product used. Lymphocyte morphology followed ALC changes and showed a consistent sequence of changes that was seen across multiple CAR T cell products. Finally, CPD parameter WY which is a measure of nuclei acid content and activation, was an early harbinger of ALC expansion. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.


Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 3877-3877
Author(s):  
Cristiana Carniti ◽  
Chiara Monfrini ◽  
Vanessa Aragona ◽  
Martina Magni ◽  
Cristina Vella ◽  
...  

Abstract Background: CD19-directed CAR-T cell therapy has shown promising efficacy in relapsed/refractory (R/R) B-cell malignancies in clinical trials resulting in the approval and commercialization of two products (tisagenlecleucel/Tisa-cel and axicabtagene ciloleucel/Axi-cel) for R/R diffuse large B cell lymphoma (DLBCL) and primary mediastinal large B cell lymphoma (PMBCL). However, relapses occur in 60-65% of patients (pts) and thus a better understanding of the early determinants of response is critical to improve long-term survival in the real-world scenario. Aims of the study: To assess whether CAR-T cell expansion after infusion represents a crucial determinant to sustain effective anti-tumor responses to both Tisa-cel and Axi-celTo evaluate differences in CAR-T cell kinetics due to the use of CD28 or 4-1BB costimulatory moleculesTo identify immune phenotypic features of infusion products accounting for CAR-T cell expansion and survival probability Methods: We analyzed samples from 43 pts [29 DLBCL, 8 high grade B-cell lymphoma (HGBCL) and 6 PMBCL] treated with Axi-cel (n=22) and Tisa-cel (n=21) at the Fondazione IRCCS Istituto Nazionale Tumori prospectively collected between November 2019 and April 2021. CAR-T cells were monitored in the peripheral blood (PB) on days 0, 4, 7, 10, 14, 21, 28 and monthly post infusion by flow cytometry (FCM). Cells were stained with CD19 CAR Detection Reagent (Miltenyi), CD3, CD4, CD8, CD45, CD14, CD45RO, CD62L, CD197, CD279, CD223 and CD366. Residual cells obtained from washings of 32 infused commercial CAR-T bags (10 Tisa-cel and 22 Axi-cel) were also analyzed by FCM. Data were acquired on a BD FACSCanto II (BD Biosciences) and a MACSQuant® Analyzer MQ10 (Miltenyi) and analyzed using FlowJo software, version 10. Results: The median time to maximal expansion of CAR-T cells was at day 10 post infusion with no differences between Axi-cel and Tisa-cel [median concentration at day 10 (C 10) 25 for Axi-cel vs 26 CAR-T cells/µl for Tisa-cel; p, ns], nor among the different histologies (median C 10 33 for DLBCL vs 19 for HGBCL vs 18 CAR-T cells/µl for PMBCL; p, ns). On the contrary, CAR-T peak concentration (C max) was higher in responders at 3 months post infusion (RE, n=28) (defined as pts achieving complete or partial response by PET/CT) than in non responders (NR, n=13) (median C max 87 in RE vs 26 in NR CAR-T cells/µl; p<0.01; Fig 1A). Consistently, the magnitude of CAR-T cell expansion in the first 30 days was higher in RE than in NR [median area under the curve (AUC 0-30) 189 vs 50; p<0.005; Fig 1B]. Circulating CAR-T cells were enriched in subpopulations representing naïve T cells (CD8+ T N; CD45RO−/CD62L+) in RE (median 0.4% in RE vs 0.04% in NR, p<0.05) while NR had significantly higher levels of effector memory T cells (CD8+ T EM; CD45RO+/CD62L+) (median 26.5% in RE vs 66.2% in NR, p<0.05). Additionally, the extent of CAR-T cell expansion predicted the progression free survival (PFS), but not the overall survival (OS), irrespective of the product used (Fig 2, p<0.05) and the overall survival was improved by salvage treatment with bispecifc antibodies. Finally, we evaluated whether CAR-T cell expansion was influenced by the immune phenotypic attributes of the infused products. A significant enrichment of central memory populations (CD8+ T CM; CD45RO−/CCR7+/CD62L+) among CAR-T cells within the infusion products of pts with longer PFS was documented, as compared with those with shorter PFS (CD8+ T CM; median 15.2% vs 3.1%; p<0.005). Conclusion: To the best of our knowledge, this is the first study assessing the clinical utility of early CAR-T cell monitoring in lymphoma pts receiving both commercial anti-CD19 CAR-T cell therapies. We provide evidence that in pts treated with Axi-cel and Tisa-cel: i) the in vivo kinetics of the CAR-T cell products are similar, consistent with the fact that no differences in the outcome of Axi-cel and Tisa-cel treated pts were detected; ii) CAR-T cell expansion is critical for efficacy and predicts the PFS; iii) circulating CAR-T cells in responders have a naïve phenotype; iv) a memory signature in the CAR-T cell product before infusion is associated with in vivo expansion and survival. Figure 1 Figure 1. Disclosures Chiappella: Celgene Bristol Myers Squibb: Other: lecture fee, advisory board; Incyte: Other: lecture fee; Novartis: Other: lecture fee; Astrazeneca: Other: lecture fee; Servier: Other: lecture fee; Takeda: Other: advisory board; Gilead Sciences: Other: lecture fee, advisory board; Clinigen: Other: lecture fee, advisory board; Roche: Other: lecture fee, advisory board; Janssen: Other: lecture fee, advisory board. Corradini: AbbVie, ADC Theraputics, Amgen, Celgene, Daiichi Sankyo, Gilead/Kite, GSK, Incyte, Janssen, KyowaKirin, Nerviano Medical Science, Novartis, Roche, Sanofi, Takeda: Consultancy; AbbVie, ADC Theraputics, Amgen, Celgene, Daiichi Sankyo, Gilead/Kite, GSK, Incyte, Janssen, KyowaKirin, Nerviano Medical Science, Novartis, Roche, Sanofi, Takeda: Honoraria; KiowaKirin; Incyte; Daiichi Sankyo; Janssen; F. Hoffman-La Roche; Kite; Servier: Consultancy; Novartis; Gilead; Celgene: Consultancy, Other: Travel and accommodations; BMS: Other: Travel and accommodation; Sanofi: Consultancy, Honoraria; Amgen; Takeda; AbbVie: Consultancy, Honoraria, Other: Travel and accommodations; Incyte: Consultancy; Novartis, Janssen, Celgene, BMS, Takeda, Gilead/Kite, Amgen, AbbVie: Other: travel and accomodations.


Author(s):  
Mengyan Li ◽  
Scott J. Garforth ◽  
Kaitlyn E. O’Connor ◽  
Hang Su ◽  
Danica M. Lee ◽  
...  

2021 ◽  
Vol 9 (Suppl 1) ◽  
pp. A16-A16
Author(s):  
V Blumenberg ◽  
S Völkl ◽  
G Busch ◽  
S Baumann ◽  
M Winkelmann ◽  
...  

BackgroundCD19. CAR T-cells for the treatment of relapsed and refractory (r/r) Diffuse Large B-Cell Lymphoma (DLBCL) demonstrated complete responses in 40%-58% of the patients. Recently, others could associate high tumor volume and low CAR T-cell expansion in vivo with poor outcome. We hypothesize, that the expansion and immunphenotype of (CAR) T cells in vivo determine treatment response and depend on patient- and disease associated factors.Materials and MethodsPatients with r/r DLBCL (n=34) were treated with either Axi-cel or Tisa-cel at the University Hospitals of Erlangen and Munich (LMU). The CAR T-cell product and peripheral blood were collected on day 0, 4, 7, 14, 30, 60 and 90 post transfusion. CAR T-cells were detected through flow cytometry utilizing a two step-staining with a biotinylated CD19 protein. Effector:Target (E:T) Ratios were estimated as absolute peak expansion of CAR T-cells (/ul) per tumorvolume (cm3). Responder (R, complete or partial remission) were compared to Non-Responder (NR, stable or progressive disease) according to response assessment with PET-CT three months after transfusion.ResultsCAR T-cell expansion peaked between day 7 and day 14 after transfusion with a greater expansion of CD8+compared to CD8- CAR T-cells on day 14 (59.27% vs 37.42%, p=0.021). The ratio of CD8+ and CD8- CAR T-cells did not differ between R and NR, however R exhibited higher E:T ratios of CD3+ CAR T-cells compared to NR (20.94 vs 12.81, p=0.015) and an increased E:T ratio of CD8+ CAR T-cells correlated with better progression-free survival (p=0.033). Interestingly, high CRP and ferritin levels at baseline were inversely associated with the E:T ratio (p=0.048 and p=0.017). CD3+ CAR T-cells of R showed earlier peak expression of PD-1 than NR (day 7 vs day 21). Further, peak expansion of CD3+ CAR T-cells correlated with higher PD-1 expression in R but not in NR (p=0.003 vs p=0.12). In addition, R revealed an increased relative frequency of effector memory differentiated CD3+ CAR T-cells (CCR7-CD45RA-, p=0.02), whereas CAR T-cells in NR showed an increased relative frequency of a naïve phenotype (CCR7+CD45RA+, p=0.001) on day 7 post infusion.ConclusionsFlow-based immunomonitoring with longitudinal characterization of CAR T-cells demonstrated a correlation of the E:T ratio with treatment response and survival. Increased inflammatory conditions at baseline correlated with diminished E:T ratios. Notably, in R CAR peak expansion was positively associated with higher PD-1 expression suggestive for superior CAR T-cell activation. In addition, greater memory differentiation was associated with efficacy during the time of peak expansion. Multiparameter analysis with other clinical covariates will show, whether CAR T-cell expansion and immunphenotypes can predict patient outcome.Disclosure InformationV. Blumenberg: B. Research Grant (principal investigator, collaborator or consultant and pending grants as well as grants already received); Significant; Novartis, Gilead Sciences, Janssen, BMS/Celgene. S. Völkl: None. G. Busch: None. S. Baumann: None. M. Winkelmann: None. B. Tast: None. D. Nixdorf: None. G. Hänel: None. L. Frölich: None. C. Schmidt: None. R. Jitschin: None. F. Vettermann: None. W. Kunz: None. D. Mougiakakos: None. M. von Bergwelt: None. V. Bücklein: None. A. Mackensen: None. M. Subklewe: None.


Sign in / Sign up

Export Citation Format

Share Document