Single HLA Class I Mismatches with High Peptide Divergence in the Graft-Versus-Host Direction Are Associated with Inferior Survival after 9/10 HLA-Matched UD-HCT: A Retrospective Study from the CIBMTR

Abstract Introduction Unrelated donor (UD)-recipient disparity for human leukocyte antigen (HLA) class I adversely affects outcome of hematopoietic cell transplantation (UD-HCT). HLA polymorphisms in the peptide antigen binding groove can affect the repertoire of presented peptides. We have recently shown that the degree of peptide divergence between mismatched HLA-DP allotypes is related to T-cell alloreactivity and clinical permissiveness after UD-HCT (Meurer et al Blood 2021). Here, we hypothesized that the clinical tolerability also of HLA class I mismatches in UD-HCT might depend on the divergence of their respective peptide repertoires. Methods We studied 2,562 patients after 9/10 HLA-A, -B, -C, -DRB1, -DQB1-matched UD-HCT for acute myeloid or lymphocytic leukemia, or myelodysplastic syndrome, between 2008 and 2018, and 14,426 10/10 HLA-matched UD-HCT with similar characteristics. Peptide divergence of the mismatched HLA allotypes was predicted based on hierarchical clustering of experimentally determined peptide binding motifs (PBM) (Bassani-Sternberg et al Front Immunol 2018), with 21 different PBM groups identified in 122 HLA class I allotypes (44, 63 and 18 for HLA-A, -B and -C, respectively). The mismatched cohort was stratified into PBM-matches or PBM-mismatches, and within the latter into host-versus-graft (HvG), graft-versus-host (GvH) or bidirectional PBM-mismatches (Figure 1A). The primary study endpoint was overall survival (OS); secondary endpoints included treatment-related mortality (TRM), GVHD and relapse. P-value<0.01 was considered statistically significant. Results The available PBM data allowed us to classify 1,629/2,562 (63.6%) of our pairs. Of these, 386 (23.7%) were PBM-matched and 1,243 (76.3%) were PBM-mismatched, and in the latter, 254 (20.5%), 238 (19.1%) and 751 (60.4%) had HvG, GvH or bidirectional PBM-mismatches, respectively. Transplants were performed mainly with peripheral blood stem cells (78%), myeloablative conditioning (65%) and tacrolimus-based graft-versus-host disease (GvHD) prophylaxis (74%). About half of the 9/10 HLA-matched HCT were performed using in vivo T-cell depletion by anti-thymocyte globulin or Campath, and none used post-transplant cyclophosphamide. Multivariable analyses showed that 10/10 HLA-matched transplants had significantly higher OS, lower TRM and aGvHD 3-4 compared to 9/10 HLA-matched transplants but relapse was similar (Figure 1B,C). There were no significant differences between the PBM-matched and aggregate PBM-mismatched group (Figure 1B). In further analysis, pairs with a bidirectional or only GvH PBM-mismatch had significantly worse OS, compared to pairs in the PBM-matched group or with only a unidirectional HvG (hazards ratio [HR] 0.76, 95% confidence interval [CI] 0.63-0.92, P = 0.0036; Figure 1C). The hazards of TRM and aGvHD 3-4 were lower for the HvG or PBM-matched group compared to the reference (HR 0.78, 95% CI 0.65-0.95, P = 0.0135 and HR 0.79, 95% CI 0.65-0.95, P = 0.0126, respectively), although these were not statistically significant (Figure 1C). Conclusion We show that single HLA class I PBM-mismatches with high peptide divergence in the unidirectional or bidirectional GvH directions are significantly associated with worse survival after 9/10 HLA-matched UD-HCT compared to PBM-matched or unidirectional mismatching in the HvG direction. These data suggest that the mechanistic role of peptide-diversity for T-cell alloreactivity we previously observed for HLA-DPB1 disparity (Meurer et al., Blood 2021), is also a relevant to class I mismatches, providing a new rationale for selecting permissive donors in the setting of 9/10 HLA-matched UD-HCT. Avoiding class I PBM mismatches in the GvH direction is associated with better survival. Figure 1 Figure 1. Disclosures Paczesny: Medical University of South Carolina: Patents & Royalties: inventor on the ST2 bispecific antibody patent application. Lee: Amgen: Research Funding; AstraZeneca: Research Funding; Incyte: Research Funding; Janssen: Other; Kadmon: Research Funding; National Marrow Donor Program: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees, Research Funding; Pfizer: Research Funding; Syndax: Research Funding; Takeda: Research Funding.

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Human CD83 Targeted Chimeric Antigen Receptor T Cell for the Prevention of Graft Versus Host Disease and Treatment of Myeloid Leukemia

Blood ◽

10.1182/blood-2019-124031 ◽

2019 ◽

Vol 134 (Supplement_1) ◽

pp. 196-196

Author(s):

Bishwas Shrestha ◽

Kelly Walton ◽

Jordan Reff ◽

Elizabeth M. Sagatys ◽

Nhan Tu ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Board Of Directors ◽

Research Funding ◽

Advisory Committees ◽

Graft Versus Host ◽

Car T Cells ◽

Car T Cell ◽

Car T ◽

Fund Research

Distinct from pharmacologic immunosuppression, we designed a programmed cytolytic effector T cell that prevents graft versus host disease (GVHD). CD83 is expressed on allo-activated conventional T cells (Tconv) and pro-inflammatory dendritic cells (DCs), which are implicated in GVHD pathogenesis. Therefore we developed a novel human CD83 targeted chimeric antigen receptor (CAR) T cell for GVHD prophylaxis. Here we demonstrate that human CD83 CAR T cells eradicate cell mediators of GVHD, significantly increase the ratio of regulatory T cells (Treg) to allo-activated Tconv, and provide lasting protection from xenogeneic GVHD. Further, we show human, acute myeloid leukemia (AML) expresses CD83 and can be targeted by CD83 CAR T cells. A 2nd generation CD83 CAR was generated with CD3ζ and 41BB costimulatory domain that was retrovirally transduced in human T cells to generate CD83 CAR T cells. The CD83 CAR construct exhibited a high degree of transduction efficiency of about 60%. The CD83 CAR T cells demonstrated robust IFN-γ and IL-2 production, killing, and proliferation when cultured with CD83+ target cells. To test whether human CD83 CAR T cells reduce alloreactivity in vitro, we investigated their suppressive function in allogeneic mixed leukocyte reactions (alloMLR). CD83 CAR T cells were added to 5-day alloMLRs consisting of autologous T cells and allogeneic monocyte-derived DCs at ratios ranging from 3:1 to 1:10. The CD83 CAR T cells potently reduced alloreactive T cell proliferation compared to mock transduced and CD19 CAR T cells. We identified that CD83 is differentially expressed on alloreactive Tconv, compared to Tregs. Moreover, the CD83 CAR T cell efficiently depletes CD83+ Tconv and proinflammatory DCs with 48 hours of engagement. To test the efficacy of human CD83 CAR T cells in vivo, we used an established xenogeneic GVHD model, where mice were inoculated with human PBMCs (25x106) and autologous CD83 CAR (1-10x106) or mock transduced T cells. The CD83 CAR T cells were well tolerated by the mice, and significantly improved survival compared to mock transduced T cells (Figure 1A). Mice treated with CD83 CAR T cells exhibited negligible GVHD target organ damage at day +21 (Figure 1B). Mice inoculated with CD83 CAR T cells demonstrated significantly fewer CD1c+, CD83+ DCs (1.7x106 v 6.2x105, P=0.002), CD4+, CD83+ T cells (4.8x103 v 5.8x102, P=0.005), and pathogenic Th1 cells (3.1x105 v 1.1x102, P=0.005) at day +21, compared to mice treated with mock transduced T cells. Moreover, the ratio of Treg to alloreactive Tconv (CD25+ non-Treg) was significantly increased among mice treated with CD83 CAR T cells (78 v 346, P=0.02), compared to mice injected with mock transduced T cells. Further, CD83 appears to be a promising candidate to target myeloid malignancies. We observed CD83 expression on malignant myeloid K562, Thp-1, U937, and MOLM-13 cells. Moreover, the CD83 CAR T cells effectively killed AML cell lines. Many AML antigens are expressed on progenitor stem cells. Thus, we evaluated for stem cell killing in human colony forming unit (CFU) assays, which demonstrated negligible on-target, off-tumor toxicity. Therefore, the human CD83 CAR T cell is an innovative cell-based approach to prevent GVHD, while providing direct anti-tumor activity against myeloid malignancies. Figure Disclosures Blazar: Kamon Pharmaceuticals, Inc: Membership on an entity's Board of Directors or advisory committees; Five Prime Therapeutics Inc: Co-Founder, Membership on an entity's Board of Directors or advisory committees; BlueRock Therapeutics: Membership on an entity's Board of Directors or advisory committees; Abbvie Inc: Research Funding; Leukemia and Lymphoma Society: Research Funding; Childrens' Cancer Research Fund: Research Funding; KidsFirst Fund: Research Funding; Tmunity: Other: Co-Founder; Alpine Immune Sciences, Inc.: Research Funding; RXi Pharmaceuticals: Research Funding; Fate Therapeutics, Inc.: Research Funding; Magenta Therapeutics and BlueRock Therapeuetics: Membership on an entity's Board of Directors or advisory committees; Regeneron Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees. Davila:Atara: Research Funding; Celgene: Research Funding; Precision Biosciences: Consultancy; Bellicum: Consultancy; GlaxoSmithKline: Consultancy; Adaptive: Consultancy; Anixa: Consultancy; Novartis: Research Funding.

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Impact of HLA Molecular Mismatch on Haploidentical Hematopoietic Stem Cell Transplantation: A Center for International Blood and Marrow Transplant Research Study

Blood ◽

10.1182/blood-2021-146847 ◽

2021 ◽

Vol 138 (Supplement 1) ◽

pp. 3917-3917

Author(s):

Jun Zou ◽

Tao Wang ◽

Yung-Tsi Bolon ◽

Shahinaz M. Gadalla ◽

Steven G.E. Marsh ◽

...

Keyword(s):

Stem Cell ◽

Board Of Directors ◽

Research Funding ◽

Hla Class I ◽

Marrow Transplant ◽

Class I ◽

Advisory Committees ◽

Hematopoietic Stem ◽

Blood And Marrow Transplant ◽

Neutrophil Engraftment

Abstract ABSTRACT BACKGROUND The number of haploidentical hematopoietic stem cell transplantations (haplo-HSCT) being performed has substantially increased in recent years. Single-center studies have previously used in silico algorithms to quantitively measure HLA disparity and shown an association of the number of HLA molecular mismatches with relapse protection and/or increased risk of acute graft-versus-host disease (GVHD) in haplo-HSCT. However, inconsistent results from small studies have made it difficult to understand the full clinical impact of molecular mismatch in haplo-HSCT. OBJECTIVE In the current study, we investigated the potential of the HLA class I and II mismatched eplet (ME) score measured by HLAMatchmaker, as well as ME load at a specific locus to predict outcomes in a registry-based cohort of haplo-HSCT recipients. STUDY DESIGN We analyzed data from patients (n= 1,287) who underwent their first haplo-HSCT for acute lymphoblastic leukemia, acute myeloid leukemia, or myelodysplastic syndrome between 2013 and 2017, as reported to the Center for International Blood and Marrow Transplant Research database. ME load at each HLA locus and total Class-I and -II were scored using the HLAMatchmaker module incorporated in HLA Fusion software v4.3, which identifies predicted eplets based on the crystalized HLA molecule models and identifies ME by comparing donor and recipient eplets. RESULTS In the cohort studied, ME scores derived from total HLA Class I or Class II loci or individual HLA loci were not associated with overall survival, disease-free survival, non-relapse mortality, relapse, acute or chronic GVHD (P< .01). An unexpected strong association was identified between total class II ME load in the GVH direction and slower neutrophil engraftment (HR 0.82; 95% CI, 0.75 - 0.91; P < .0001) and platelet engraftment (HR 0.80; 95% CI, 0.72 - 0.88; P < .0001). This was likely attributable to ME load at the HLA-DRB1 locus, which was similarly associated with slower neutrophil engraftment (HR 0.82; 95% CI, 0.73 - 0.92; P = .001) and slower platelet engraftment (HR 0.76; 95% CI, 0.70 - 0.84; P < .0001). Additional analyses suggested that this effect is attributable to matched vs. mismatched in the GVH direction and not to ME load, as there was no dose effect identified. CONCLUSION These findings contradict those of prior relatively small studies reporting that ME load, as quantified by HLAMatchmaker, was associated with haplo-HSCT outcomes. As the study failed to demonstrate the predictive value of ME from HLA molecules for major clinical outcomes, other molecular mismatch algorithms in haplo-HSCT settings should be tested. Disclosures Lee: Pfizer: Research Funding; Novartis: Membership on an entity's Board of Directors or advisory committees, Research Funding; National Marrow Donor Program: Membership on an entity's Board of Directors or advisory committees; Incyte: Research Funding; Janssen: Other; Takeda: Research Funding; Syndax: Research Funding; AstraZeneca: Research Funding; Kadmon: Research Funding; Amgen: Research Funding.

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Lenalidomide/Rituximab Maintenance After Induction With Fludarabine/Rituximab In Combination With Escalating Doses Of Lenalidomide In Previously Untreated Chronic Lymphocytic Leukemia (CLL): The Revlirit CLL5 AGMT Phase I/II Study, Final Results

Blood ◽

10.1182/blood.v122.21.4164.4164 ◽

2013 ◽

Vol 122 (21) ◽

pp. 4164-4164 ◽

Cited By ~ 6

Author(s):

Alexander Egle ◽

Michael Steurer ◽

Franz Josef Gassner ◽

Roland Geisberger ◽

Thomas Melchardt ◽

...

Keyword(s):

High Risk ◽

Board Of Directors ◽

Research Funding ◽

Maintenance Treatment ◽

Maintenance Phase ◽

Study Endpoint ◽

Primary Study ◽

Advisory Committees ◽

Skin Reactions ◽

Tumor Lysis

Abstract Introduction Lenalidomide has shown encouraging activity in monotherapy trials in CLL, but tumor lysis and tumor flare presented obstacles in development. We and others previously presented first data on combinations of lenalidomide with standard treatment regimens in CLL. As reported at ASH 2011 we combined Lenalidomide safely and efficaciously with the combination of Fludarabine and Rituximab, achieving early reduction of tumor load without tumor lysis or tumor flare and with high response rates. We also uncovered a patient population unable to tolerate higher Lenalidomide doses and marked by an exhausted T cell subset, measured pre-treatment. We now report final results of this trial, including the maintenance phase. Study design In induction Lenalidomide was combined with Fludarabine (40mg/m2 po d1-3 q28d) and Rituximab (375mg/m2 iv d4 cycle 1; 500mg/m2 iv d1 cycles 2-6, q28d). In cycle 1 Lenalidomide was added day 7-21 at 2.5 mg. Toxicity permitting, Lenalidomide dose was escalated to 5, 10, 15, 20 and 25mg day 1-21 over cycles 2-6. Subsequent maintenance treatment was two-monthly Rituximab at 375mg/m2 and Lenalidomide at the last dose tolerated in combination in a 28 day cycle without interruption for 6 months. The main goal of this treatment phase was to establish safety and efficacy as a secondary endpoint to the study. Results Patient characteristics of 45 recruited patients were previously reported: median age was 66 years and at least one molecular high risk feature was present in 64% of patients. No systematic toxicity determining an MTD in induction, the primary study endpoint, was found. The median daily dose in cycle 6 was 15mg in 40 evaluable patients, with 3 patients receiving the last cycle without lenalidomide. Toxicity and efficacy of the induction regimen were reported previously. Maintenance treatment was started in all 40 patients finishing induction. Three patients that finished without lenalidomide received only Rituximab. The median starting dose for all 40 patients was 15mg daily and 70% started with 10mg upwards. In total, 46% needed dose reductions, with prolonged neutropenia being the main reason, but 47% received doses above 10mg up to cycle 6 of maintenance. Interestingly, 9/13 patients receiving 25mg as maintenance were able to receive the treatment uninterrupted for 6 months, suggesting that a biologically select group may tolerate very high doses. As alluded to the major toxicity was neutropenia with 45% and 27% reaching G3 and G4, respectively. Surprisingly this did not translate into a relevant signal for infections. Grade 3, but no G4 infections, were observed in 5% of patients and all other G3/4 toxicities remained below 5%. Compared to the reported incidence of skin reactions in induction, we did not observe a significant signal in the maintenance phase. Improvement of response from PR after induction to CR at the end of maintenance was observed in 25%. The overall best response in ITT to the regimen during induction and maintenance was CR in 67% and PR in 29%. Median follow up of the study is now 35 months, at which point PFS is 89% and observed median PFS is currently 46 months. Exploratory analyses show no significant influences of age>65, mutation state or CD38 risk on PFS, but undetectable MRD after induction and high risk cytogenetics showed borderline effects (both p=0.08), the latter (2 cases with del17p and 9 with del11q) being driven by relapses in patients with del11q with a median PFS of 42 months in this group. Conclusions A combination of Lenalidomide with FR followed my maintenance with Lenalidomide and Rituximab proved clinically feasible. While initial dose-finding was complicated by highly individual levels of tolerance to lenalidomide in the combination (with skin toxicity being a major problem), the main toxicity in maintenance was neutropenia. Although the important myelotoxicity did not translate into a rate of infection above that expected after induction treatment, our judgment is, that a more moderate approach to dosing may be warranted in maintenance, since a majority of patients had to be dose-adjusted. Finally, the PFS observed with this induction and maintenance regimen seems encouraging by comparison with other first line regimens. While the exploratory nature of the trial clearly limits this conclusion, we further explore combination approaches with lenalidomide. Disclosures: Egle: Celgene: Honoraria, Membership on an entity’s Board of Directors or advisory committees, Research Funding; Roche: Honoraria, Research Funding. Off Label Use: Lenalidomide in CLL. Pleyer:Celgene: Honoraria, Research Funding. Fridrik:Roche: Honoraria. Thaler:Roche: Honoraria, Research Funding. Greil:Roche: Honoraria, Membership on an entity’s Board of Directors or advisory committees, Research Funding; Celgene: Honoraria, Research Funding.

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Multicenter Phase II Trial of Bendamustine, Lenalidomide and Dexamethasone (BRd) As 2nd-Line Therapy for Multiple Myeloma

Blood ◽

10.1182/blood.v124.21.4779.4779 ◽

2014 ◽

Vol 124 (21) ◽

pp. 4779-4779

Author(s):

Ulrich JM Mey ◽

Wolfram Brugger ◽

Mario Bargetzi ◽

Heike Schwarb ◽

Andreas Schwarzer ◽

...

Keyword(s):

Board Of Directors ◽

Induction Therapy ◽

Research Funding ◽

Study Endpoint ◽

Primary Study ◽

Induction Treatment ◽

High Quality ◽

Advisory Committees ◽

Line Therapy ◽

Grade 3

Abstract Background High quality response [very good partial remission (VGPR), complete remission (CR)] is associated with improved long-term outcome in multiple myeloma (MM) therapy. Lenalidomide (Len)/ dexamethasone (Dex) (Rd) is considered a standard 2nd-line regimen. However, after 1st-line MM therapy containing a “novel agent” like thalidomide, only a relatively small fraction (21.3%) of patients (pts) achieved VGPR or better when RD was used in relapsed/refractory MM (RRMM)1. Bendamustine (Ben) is an alkylating agent with superior activity compared to melphalan/prednisone. The combination of Ben, Len and Dex (BRd) in pts with advanced RRMM resulted in dose-limiting hematotoxicity, which restricted its efficacy in extensively pretreated pts2. However, recent phase I experience demonstrated the feasibility of BRd as 2nd- and 3rd-line myeloma therapy3. We therefore evaluated the efficacy and toxicity of the BRd-regimen as 2nd line therapy for RRMM. Patients and methods This multicenter Phase II study was designed to enroll 50 pts with RRMM undergoing 2nd-line MM therapy [including also pts with prior autologous stem cell transplantation (ASCT)]. Pts had to have measurable disease, ECOG performance status 0-2, adequate hematological values and a creatinine clearance > 50 ml/min. Study treatment consisted of Ben 75 mg/m2 i.v. day (d) 1 & 2 and Len 25 mg p.o. d 1-21 for a total of six 28-day induction cycles. Dex (40 and 20 mg p.o. for pts <= or > 75 years of age, respectively) was given on d 1, 8, 15, and 22. Pegfilgrastim 6 mg s.c. was administered on d 3 in case of severe neutropenia, treatment delay due to neutropenia or febrile neutropenia according to predefined application rules. Induction treatment was followed by 12 cycles (28 d) of maintenance therapy with Rd at the same dose. The primary study endpoint was the CR/VGPR rate after induction therapy, based on standard IMWG criteria. A Simon's two stage design was used to differentiate between 20% (considered uninteresting) vs 40 % (considered promising, power of 80%, p=5%) of high quality responses. At least 13 pts with VGPR or better after induction therapy were required to meet the statistical threshold predefined for promising activity of the BRd regimen in this study. Results 50 pts were enrolled between 04/2012 and 07/2014 (median age 68 years [46-84y], 73% men). 49% pts had ISS stage II/III, 42.5% had undergone prior SCTs, and 13% had known high risk cytogenetic aberrations. 91% of pts had received novel agents (thalidomide, bortezomib) during 1st-line therapy. At the time of abstract submission, data from 38 pts having completed induction treatment were available. Final analysis of the primary study endpoint including all patients will be updated and presented at the meeting. Of the currently 38 evaluable pts, 20 (52.6%, Table 1) achieved a CR/VGPR after a median of 3.3 induction cycles. Only 1 patient experienced disease progression (PD) during induction phase. 77% of pts received pegfilgrastim. Dose reduction during induction therapy was required in 7.7% of pts. 42.5% received < 6 scheduled induction cycles (50% due to toxicities, 50% for other reasons). 49% had treatment-related SAEs. Grade 3/ 4 neutropenia and thrombocytopenia occurred in 51%/25.5% and 23.4%/8.5% of pts, respectively. Only 1 patient developed CTC grade 3 febrile neutropenia. Most common grade 3/4 non-hematologic toxicities were infections (14%), rash (9.5%), and diarrhea (9.5%). Anaphylactic reaction grade 4 related to Ben and pulmonary embolism grade 3 occurred in 1 patient each. A cerebral insult grade 4 occurred in a patient non-compliant with the anti-thrombotic prophylaxis required per protocol. One death due to respiratory failure (considered to be unlikely related to study treatment) was reported for an 83 year old male. Conclusion BRd is a safe and efficacious regimen for 2nd line treatment of RRMM patients whose 1st-line therapy included thalidomide or bortezomib. High quality responses (>= VGPR) can be achieved in a considerable proportion (52.6%) of these pts. Our study suggests that the fraction of patients achieving VGPR or better after BRd treatment may be substantially higher than attainable with Rd alone. References 1 Wang et al. Blood, 2008, 112: 4445-51 2 Lentzsch et al. Blood, 2012, 119: 4608-13 3 Poenisch et al., Br J Haematol, 2013, 162, 202–9 Table 1 Best response after induction CR VGPR PR MR SD total (n=38) 3 17 15 1 2 prior ASCT (n=16) 2 6 7 1 0 no prior ASCT (n=22) 1 11 8 0 2 Disclosures Mey: Mundipharma: Honoraria, Research Funding; Amgen: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding. Off Label Use: Use of Bendamustine in relapsed/refractory Multiple Myekoma. To investigate the efficacy and safety of Bendamustine in combination with the standard backbone of Lenalidomide/ Dexamethasone in patients with replaced/refractory MM.. Bargetzi:Celgene: Membership on an entity's Board of Directors or advisory committees. Taverna:Celgene: Consultancy, Membership on an entity's Board of Directors or advisory committees. Schmid:Celgene: Honoraria. Knauf:Celgene: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Mundipharma: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees. von Moos:Amgen: Consultancy, Honoraria, Research Funding. Hiendlmeyer:Celgene: Research Funding; Mundipharma: Research Funding; Amgen: Research Funding. Hitz:Celgene: Research Funding. Driessen:Mundipharma: Membership on an entity's Board of Directors or advisory committees, Research Funding; Amgen: Membership on an entity's Board of Directors or advisory committees.

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Neoantigens Are Typically Associated with Intact HLA Class I Presentation in Early-Stage Follicular Lymphoma

Blood ◽

10.1182/blood-2020-135954 ◽

2020 ◽

Vol 136 (Supplement 1) ◽

pp. 37-38 ◽

Cited By ~ 1

Author(s):

Hennes Tsang ◽

Ann-Marie Patch ◽

Colm Keane ◽

Soi C. Law ◽

Jay Gunawardana ◽

...

Keyword(s):

Follicular Lymphoma ◽

Antigen Presentation ◽

Board Of Directors ◽

Tumor Antigen ◽

Research Funding ◽

Early Stage ◽

Hla Class I ◽

Class I ◽

Advisory Committees ◽

Tissue Samples

Neoantigens are generated by mutations acquired in malignant cells. To be immunogenic, neoantigen peptides must be processed and then presented by HLA molecules on the tumor cell surface to evoke a T cell response. Emerging evidence suggests neoantigens are promising new targets of personalised cancer immunotherapies, provided that antigen presentation remains intact. Recently, we (Tobin, JCO 2019) demonstrated the link between the tumor microenvironment (TME) and clinical outcome in patients with advanced-stage Follicular Lymphoma (ASFL). In particular, ASFL patients with favorable outcomes had high levels of immune infiltration (best stratified by PD-L2 expression) with increased intratumoral CD8+ T cell clonotypes suggestive of expanded antigen-specific T cells in response to presentation by HLA class I neoantigens. This is supported by HLA class I pathway genetic aberrations being infrequent (unlike HLA class II) in FL. However, to date there is minimal data on the frequency and nature of putative neoantigens, their relationship with HLA class I antigen presentation and with the TME in any stage of FL. In particular, the immunobiology of early-stage FL (ESFL) has been largely neglected. Here, we present detailed characterization of these parameters in patients with early-stage FL (ESFL) from the TROG99.03 prospective clinical trial (MacManus, JCO 2018). Digital multiplex gene expression (770 gene PanCancer Immune Panel, NanoString) and targeted NGS (365 genes recurrently mutated in hematological malignancies) was performed on 97 FFPE diagnostic tissue samples. Intratumoral CD8A expression cut-off derived by maximally selected rank statistics was associated with >2-fold differential median PFS (CD8Ahi: 11.5 years vs. CD8Alo: 4.9 years, p=0.0042, Figure 1A). In keeping with a relationship between HLA class I immune effector infiltration within the TME and FL tumor antigen presentation/processing, expression of NLRC5 (a transcriptional activator of HLA class I) was also associated with differential median PFS (NLRC5hi: 10.8 years vs. NLRC5lo: 4.9 years, p=0.021, Figure 1B). The impact of expression of CD8A and NLRC5 were independent of mutations in BCL-2,TP53 and all but of one the m7-FLIPI genes. The exception was EZH2, mutations of which were enriched in cases with reduced CD8A (p=0.0066) and NLRC5 (p<0.0001), broadly consistent with prior observations in DLBCL (Ennishi, Can Disc 2019). Next, we investigated the expression of HLA class I and predicted neoantigens. HLA genotyping was performed using a custom sequencing panel on matched germline peripheral blood DNA for 71 TROG99.03 diagnostic samples. For the corresponding somatic coding variants, the strongest HLA class I binding efficiencies were calculated using 8 epitope prediction algorithms. 41% of samples had no detectable neoantigens, whereas 59% of samples had ≥1 neoantigens detected, with 116 neoantigens total. Importantly, unsupervised hierarchical clustering showed 2-fold enrichment of samples with neoantigens among those with elevated HLA A/B expression vs. samples with low HLA A/B (p=0.0023, Figure 1C). Interestingly, roughly one third of neoantigens were due to indels causing frameshift events resulting in the generation of 'non-self' peptides with strong predicted binding affinity. Recently TCGA data has been used to show that frameshift variants occurring at different sites in a gene can converge to generate a handful neo open reading frame peptides ('NOPs') common to multiple cancer patients. Therefore, it has been postulated that these NOPs could have a considerable therapeutic potential as a novel pan-cancer 'semi-personalized' immunotherapy. In keeping with data derived in solid tumors, in ESFL we observed frameshift variants were most frequently observed in KMT2D. Finally, 5 relapsed/transformed FL samples were compared to their paired diagnostic tissue samples. Not only did neoantigens present at diagnosis typically remain detectable, but in several cases additional neoantigens were also found. In summary, we demonstrate for the first time in ESFL that neoantigens are typically associated with intact HLA class I presentation; CD8 infiltration within the TME is related to tumor antigen processing and associated with favorable outcomes; and that a subset of FL tumors have NOPs. These data have important implications for the design of innovative next-generation immunotherapies in FL. Disclosures Keane: BMS: Research Funding; Celgene: Honoraria, Other: Travel; Roche: Honoraria, Other: Travel, Speakers Bureau; MSD Oncology: Honoraria, Other: Travel; Gilead: Honoraria, Other: Travel, Speakers Bureau. Blombery:Amgen: Consultancy; Novartis: Consultancy; Janssen: Honoraria; Invivoscribe: Honoraria. Tobin:Gilead: Research Funding. Seymour:Nurix: Honoraria; Morphosys: Consultancy, Honoraria; Mei Pharma: Consultancy, Honoraria; Gilead: Consultancy; AstraZeneca: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Janssen: Consultancy, Honoraria, Research Funding; F. Hoffmann-La Roche: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Celgene: Consultancy, Honoraria, Research Funding; AbbVie: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau. Gandhi:Celgene: Research Funding; Bristol-Myers Squibb: Research Funding; Merck Sharp & Dohme: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Amgen: Honoraria; Gilead Sciences: Honoraria; Genentech: Honoraria; Mater Research: Current Employment; Janssen-Cilag: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Roche: Other: Travel, accommodation, expenses .

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Predictors and Outcomes of Flares in Chronic Graft-Versus-Host Disease

Blood ◽

10.1182/blood-2020-136479 ◽

2020 ◽

Vol 136 (Supplement 1) ◽

pp. 47-48

Author(s):

Grigori Okoev ◽

Daniel J. Weisdorf ◽

John E Wagner ◽

Bruce R. Blazar ◽

Margaret L. MacMillan ◽

...

Keyword(s):

Regression Analysis ◽

Board Of Directors ◽

Graft Versus Host Disease ◽

Research Funding ◽

Time Dependent ◽

Advisory Committees ◽

Graft Versus Host ◽

Day Range ◽

Similar Risk ◽

Fund Research

Introduction: Chronic Graft-versus-Host Disease (cGvHD) frequently requires prolonged immune suppressive therapy (IST) with > 50% still on IST at 5 years. The IST typically involves a slow taper of steroids often with flare of cGvHD, necessitating augmentation of previous therapy or addition of new IST. Studies describing cGvHD flares are limited. We analyzed patients with cGvHD who flared during the treatment with systemic IST, their overall survival (OS) and non-relapse mortality (NRM). Methods: This study included all adult patients with cGvHD (n=145) following an allogeneic transplant (2010 - 2017) from a matched sibling donor peripheral blood stem cell transplant (MSD, n=104 (72%) or double/single umbilical cord blood transplant (UCBT, n=41 (28%). The 2014 NIH Consensus Criteria were used to classify organ/overall cGvHD severity. Flare of cGvHD was defined as progression in cGvHD manifestations (after initial response), which was less severe than at diagnosis. Multivariate regression of flares was based on the Prentice, Williams and Peterson model for ordered multiple events (flares). Time-dependent effects on OS and NRM were analyzed by Cox and Fine and Gray regression with propensity scoring to control for confounding. Results: Flares occurred in 87 patients; the cumulative incidence of flares was 60% (95% CI: 51-70%) at a median of 188 days (range 16-751) after diagnosis of cGvHD. The median dose of prednisone was 1 mg/kg/day (range 0-4.2) at diagnosis of cGvHD. At the diagnosis of flare, 36 (41%) of the patients were off prednisone, 50 (57%) were receiving 0.1-0.5 mg/kg /day, and 2 patients > 0.5 mg/kg /day. Thirty two of the 87 (36%) patients experienced multiple flares (2 to 4). The most common organs involved at cGvHD flare were skin (n=45; 51%), mouth (n=27; 31%), GI tract (n=22; 25%) and liver (n=12; 14%); often in combinations of skin/mouth in 11 cases (13%), skin/GI in 6 (7%) and liver/mouth in 4 (5%) cases. Treatment for flare was mostly increase in dose of prednisone to 0.5 mg/kg/day (range 0.3-1.0) in 77 patients (88%) plus the addition of another line of IST in 48 patients (55%). In multiple regression analysis, only donor type was significant predictor of flare in cGvHD. UCBT was associated with 2-fold lower probability of flaring (HR 0.5; 95% CI: 0.3-0.9; p=0.03) compared to MSD. cGvHD severity, organ involvement, platelet count at diagnosis and type of onset were not significant predictors of cGvHD flares. At 2 years after the initial flare, the OS was 77% (95% CI: 66-84%) and NRM 19% (95% CI: 11-28%). Multiple regression analysis evaluating OS and NRM from onset of cGvHD comparing flare to non-flare were performed using flare as a time dependent variable. Compared to cGvHD patients without flare at 2 years, those with flare of cGvHD had a similar risk of NRM (HR 1.2; 95% CI: 0.2-6.1, p=0.86) and OS (HR 0.9; 95% CI: 0.4-2.3, p=0.85). At 2 years from cGvHD onset, the cumulative incidence of resolved cGvHD (durable discontinuation of steroids for ≥ 6 consecutive months) was 31% (95% CI: 21-41%) in those who flared vs. 86% (95% CI: 75-96%) in those without flare. Conclusions: Though cGvHD patients with flare had similar risk of NRM and OS as those without a flare, patients with flare required extended steroids, along with clinical monitoring and intensified IST. cGvHD after UCBT was associated with significantly lower risk of flaring compared to MSD. The ongoing burden of IST, risk of infection and morbidity of cGvHD is substantial and needs better approaches than chronic slow taper of steroids. Disclosures Weisdorf: Incyte: Research Funding; FATE Therapeutics: Consultancy. Wagner:Novartis: Research Funding; Rocket Pharmaceuticals, Inc.: Consultancy, Current equity holder in publicly-traded company; Magenta Therapeutics: Consultancy, Research Funding; BlueRock: Research Funding; Gadeta: Membership on an entity's Board of Directors or advisory committees. Blazar:Fate Therapeutics Inc.: Research Funding; Childrens' Cancer Research Fund: Research Funding; BlueRock Therapeutics: Research Funding; BlueRock Therapeuetic: Consultancy; Magenta Therapeutics: Consultancy; KidsFirst Fund: Research Funding; Tmunity: Other: Co-founder. MacMillan:Mesoblast: Consultancy; Angiocrine Biosciences, Inc.: Consultancy; Equillium, Inc.: Consultancy; Talaris Therapeutics, Inc: Consultancy; Fate Therapeutics, Inc.: Consultancy. Holtan:Generon: Consultancy; BMS: Consultancy; CSL Behring: Other: Clinical trial data adjudication; Incyte: Consultancy. Brunstein:AlloVir: Other: Advisory board; Gamida: Research Funding; Astex: Research Funding; Magenta: Research Funding. Betts:Patent Pending: Patents & Royalties: Dr. Betts has a pending patent WO2017058950A1: Methods of treating transplant rejection. This includes the use of JAK inhibitors. Neither he nor his institution have received payment related to claims described in the patent.. Bachanova:FATE: Research Funding; Karyopharma: Membership on an entity's Board of Directors or advisory committees; Incyte: Research Funding; Gamida Cell: Membership on an entity's Board of Directors or advisory committees, Research Funding; Kite: Membership on an entity's Board of Directors or advisory committees; BMS: Research Funding. Rashidi:Synthetic Biologics: Other: DSMC member (1 trial) and related honorarium. Arora:Fate Therapeutics: Consultancy; Kadmon: Research Funding; Pharmacyclics: Research Funding; Syndax: Research Funding.

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Automated Manufacture of Matched Donor-Derived Allogeneic CD19 CAR T-Cells for Relapsed/Refractory B-ALL Following Allogeneic Stem Cell Transplantation: Toxicity, Efficacy and the Important Role of Lymphodepletion

Blood ◽

10.1182/blood-2019-129640 ◽

2019 ◽

Vol 134 (Supplement_1) ◽

pp. 776-776

Author(s):

Claire Roddie ◽

Maeve A O'Reilly ◽

Maria A V Marzolini ◽

Leigh Wood ◽

Juliana Dias Alves Pinto ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Board Of Directors ◽

Research Funding ◽

Advisory Committees ◽

Car T Cells ◽

Car T Cell ◽

Car T ◽

Peak Car ◽

Matched Donor

Introduction: 2nd generation CD19 CAR T cells show unprecedented efficacy in B-ALL, but several challenges remain: (1) scaling manufacture to meet patient need and (2) feasibility of generating products from lymphopenic patients post allogeneic stem cell transplant (allo-SCT). To overcome these issues we propose: (1) use of the CliniMACS Prodigy (Miltenyi Biotec), a semi-automated cGMP platform that simplifies CAR T cell manufacture and (2) the use of matched donor T cells to overcome the challenge posed by patient lymphopenia, albeit this may come with a heightened risk of graft versus host disease (GvHD). CARD (NCT02893189) is a Phase I study of matched donor derived CD19 CAR T cells generated on the CliniMACS Prodigy in 14 adult patients with relapsed/refractory (r/r) B ALL following allo-SCT. We additionally explore the requirement for lymphodepletion (LD) in the allogeneic CAR T cell setting and report on the incidence of GvHD with this therapy. Methods: Manufacturing: CARD utilises non-mobilised matched donor leucapheresate to manufacture 2nd generation CD19CAR T cells using a closed CliniMACS® Prodigy/ TransACTTM process. Study design: Eligible subjects are aged 16-70y with r/r B ALL following allo SCT. Study endpoints include feasibility of CD19CAR T cell manufacture from allo-SCT donors on the CliniMACS Prodigy and assessments of engraftment and safety including GvHD. To assess the requirement for LD prior to CD19CAR T cells in lymphopenic post-allo-SCT patients, the study is split into Cohort 1 (no LD) and Cohort 2 (fludarabine (30 mg/m2 x3) and cyclophosphamide (300mg/m2 x3)). To mitigate for the potential GvHD risk, cell dosing on study mirrors conventional donor lymphocyte infusion (DLI) schedules and is based on total CD3+ (not CAR T) cell numbers: Dose 1=1x106/kg CD3+ T cells; Dose 2= 3x106/kg CD3+ T cells; Dose 3= 1x107/kg CD3+ T cells. Results: As of 26 July 2019, 17 matched allo SCT donors were leukapheresed and 16 products were successfully manufactured and QP released. Patient demographics are as follows: (1) median patient age was 43y (range 19-64y); (2) 4/17 had prior blinatumomab and 5/17 prior inotuzumab ozogamicin; (3) 7/17 had myeloablative allo SCT and 10/17 reduced intensity allo SCT of which 6/17 were sibling donors and 12/17 were matched unrelated donors. No patients with haploidentical transplant were enrolled. To date, 12/16 patients have received at least 1 dose of CD19CAR T cells: 7/16 on Cohort 1 and 5/16 on Cohort 2 (2/16 are pending infusion on Cohort 2 and 2/16 died of fungal infection prior to infusion). Median follow-up for all 12 patients is 22.9 months (IQR 2.9-25.9; range 0.7 - 25.9). At the time of CAR T cell infusion, 7/12 patients were in morphological relapse with >5% leukemic blasts. Despite this, CD19CAR T cells were administered safely: only 2/12 patients experienced Grade 3 CRS (UPenn criteria), both in Cohort 1, which fully resolved with Tocilizumab and corticosteroids. No patients experienced ≥Grade 3 neurotoxicity and importantly, no patients experienced clinically significant GvHD. In Cohort 1 (7 patients), median peak CAR expansion by flow was 87 CD19CAR/uL blood whereas in Cohort 2 (5 patients to date), median peak CAR expansion was 1309 CD19CAR/uL blood. This difference is likely to reflect the use of LD in Cohort 2. CAR T cell persistence by qPCR in Cohort 1 is short, with demonstrable CAR in only 2/7 treated patients at Month 2. Data for Cohort 2 is immature, but this will also be reported at the meeting in addition to potential mechanisms underlying the short persistence observed in Cohort 1. Of the 10 response evaluable patients (2/12 pending marrow assessment), 9/10 (90%) achieved flow/molecular MRD negative CR at 6 weeks. 2/9 responders experienced CD19 negative relapse (one at M3, one at M5) and 3/9 responders experienced CD19+ relapse (one at M3, one at M9, one at M12). 4/10 (40%) response evaluable patients remain on study and continue in flow/molecular MRD negative remission at a median follow up of 11.9 months (range 2.9-25.9). Conclusions: Donor-derived matched allogeneic CD19 CAR T cells are straightforward to manufacture using the CliniMACS Prodigy and deliver excellent early remission rates, with 90% MRD negative CR observed at Week 6 in the absence of severe CAR associated toxicity or GvHD. Peak CAR expansion appears to be compromised by the absence of LD and this may lead to a higher relapse rate. Updated results from Cohorts 1 and 2 will be presented. Disclosures Roddie: Novartis: Consultancy; Gilead: Consultancy, Speakers Bureau; Celgene: Consultancy, Speakers Bureau. O'Reilly:Kite Gilead: Honoraria. Farzaneh:Autolus Ltd: Equity Ownership, Research Funding. Qasim:Autolus: Equity Ownership; Orchard Therapeutics: Equity Ownership; UCLB: Other: revenue share eligibility; Servier: Research Funding; Bellicum: Research Funding; CellMedica: Research Funding. Linch:Autolus: Membership on an entity's Board of Directors or advisory committees. Pule:Autolus: Membership on an entity's Board of Directors or advisory committees. Peggs:Gilead: Consultancy, Speakers Bureau; Autolus: Membership on an entity's Board of Directors or advisory committees.

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Identification of Two CAR T-Cell Populations Associated with Complete Response or Progressive Disease in Adult Lymphoma Patients Treated with Axi-Cel

Blood ◽

10.1182/blood-2019-130815 ◽

2019 ◽

Vol 134 (Supplement_1) ◽

pp. 779-779 ◽

Cited By ~ 2

Author(s):

Zinaida Good ◽

Jay Y. Spiegel ◽

Bita Sahaf ◽

Meena B. Malipatlolla ◽

Matthew J. Frank ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Board Of Directors ◽

Research Funding ◽

Advisory Board ◽

Advisory Committees ◽

Car T Cells ◽

Scientific Advisory Board ◽

Car T ◽

Scientific Advisory

Axicabtagene ciloleucel (Axi-cel) is an autologous anti-CD19 chimeric antigen receptor (CAR) T-cell therapy approved for the treatment of relapsed or refractory diffuse large B-cell lymphoma (r/r DLBCL). Long-term analysis of the ZUMA-1 phase 1-2 clinical trial showed that ~40% of Axi-cel patients remained progression-free at 2 years (Locke et al., Lancet Oncology 2019). Those patients who achieved a complete response (CR) at 6 months generally remained progression-free long-term. The biological basis for achieving a durable CR in patients receiving Axi-cel remains poorly understood. Here, we sought to identify CAR T-cell intrinsic features associated with CR at 6 months in DLBCL patients receiving commercial Axi-cel at our institution. Using mass cytometry, we assessed expression of 33 surface or intracellular proteins relevant to T-cell function on blood collected before CAR T cell infusion, on day 7 (peak expansion), and on day 21 (late expansion) post-infusion. To identify cell features that distinguish patients with durable CR (n = 11) from those who developed progressive disease (PD, n = 14) by 6 months following Axi-cel infusion, we performed differential abundance analysis of multiparametric protein expression on CAR T cells. This unsupervised analysis identified populations on day 7 associated with persistent CR or PD at 6 months. Using 10-fold cross-validation, we next fitted a least absolute shrinkage and selection operator (lasso) model that identified two clusters of CD4+ CAR T cells on day 7 as potentially predictive of clinical outcome. The first cluster identified by our model was associated with CR at 6 months and had high expression of CD45RO, CD57, PD1, and T-bet transcription factor. Analysis of protein co-expression in this cluster enabled us to define a simple gating scheme based on high expression of CD57 and T-bet, which captured a population of CD4+ CAR T cells on day 7 with greater expansion in patients experiencing a durable CR (mean±s.e.m. CR: 26.13%±2.59%, PD: 10.99%±2.53%, P = 0.0014). In contrast, the second cluster was associated with PD at 6 months and had high expression of CD25, TIGIT, and Helios transcription factor with no CD57. A CD57-negative Helios-positive gate captured a population of CD4+ CAR T cells was enriched on day 7 in patients who experienced progression (CR: 9.75%±2.70%, PD: 20.93%±3.70%, P = 0.016). Co-expression of CD4, CD25, and Helios on these CAR T cells highlights their similarity to regulatory T cells, which could provide a basis for their detrimental effects. In this exploratory analysis of 25 patients treated with Axi-cel, we identified two populations of CD4+ CAR T cells on day 7 that were highly associated with clinical outcome at 6 months. Ongoing analyses are underway to fully characterize this dataset, to explore the biological activity of the populations identified, and to assess the presence of other populations that may be associated with CAR-T expansion or neurotoxicity. This work demonstrates how multidimensional correlative studies can enhance our understanding of CAR T-cell biology and uncover populations associated with clinical outcome in CAR T cell therapies. This work was supported by the Parker Institute for Cancer Immunotherapy. Figure Disclosures Muffly: Pfizer: Consultancy; Adaptive: Research Funding; KITE: Consultancy. Miklos:Celgene: Membership on an entity's Board of Directors or advisory committees; BMS: Membership on an entity's Board of Directors or advisory committees; Kite-Gilead: Membership on an entity's Board of Directors or advisory committees, Research Funding; AlloGene: Membership on an entity's Board of Directors or advisory committees; Precision Bioscience: Membership on an entity's Board of Directors or advisory committees; Miltenyi Biotech: Membership on an entity's Board of Directors or advisory committees; Becton Dickinson: Research Funding; Adaptive Biotechnologies: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees, Research Funding; Juno: Membership on an entity's Board of Directors or advisory committees. Mackall:Vor: Other: Scientific Advisory Board; Roche: Other: Scientific Advisory Board; Adaptimmune LLC: Other: Scientific Advisory Board; Glaxo-Smith-Kline: Other: Scientific Advisory Board; Allogene: Equity Ownership, Membership on an entity's Board of Directors or advisory committees; Apricity Health: Equity Ownership, Membership on an entity's Board of Directors or advisory committees; Unum Therapeutics: Equity Ownership, Membership on an entity's Board of Directors or advisory committees; Obsidian: Research Funding; Lyell: Consultancy, Equity Ownership, Other: Founder, Research Funding; Nektar: Other: Scientific Advisory Board; PACT: Other: Scientific Advisory Board; Bryologyx: Other: Scientific Advisory Board.

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Pre-Existing T Cell Subsets Determine Anti-PD1 Blockade Response in Richter's Transformation

Blood ◽

10.1182/blood-2020-138601 ◽

2020 ◽

Vol 136 (Supplement 1) ◽

pp. 42-43

Author(s):

Prajish Iyer ◽

Lu Yang ◽

Zhi-Zhang Yang ◽

Charla R. Secreto ◽

Sutapa Sinha ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Single Cell ◽

Board Of Directors ◽

Peripheral Blood ◽

Research Funding ◽

Gene Signature ◽

T Cell Subsets ◽

Rna Seq ◽

Advisory Committees

Despite recent developments in the therapy of chronic lymphocytic leukemia (CLL), Richter's transformation (RT), an aggressive lymphoma, remains a clinical challenge. Immune checkpoint inhibitor (ICI) therapy has shown promise in selective lymphoma types, however, only 30-40% RT patients respond to anti-PD1 pembrolizumab; while the underlying CLL failed to respond and 10% CLL patients progress rapidly within 2 months of treatment. Studies indicate pre-existing T cells in tumor biopsies are associated with a greater anti-PD1 response, hence we hypothesized that pre-existing T cell subset characteristics and regulation in anti-PD1 responders differed from those who progressed in CLL. We used mass cytometry (CyTOF) to analyze T cell subsets isolated from peripheral blood mononuclear cells (PBMCs) from 19 patients with who received pembrolizumab as a single agent. PBMCs were obtained baseline(pre-therapy) and within 3 months of therapy initiation. Among this cohort, 3 patients had complete or partial response (responders), 2 patients had rapid disease progression (progressors) (Fig. A), and 14 had stable disease (non-responders) within the first 3 months of therapy. CyTOF analysis revealed that Treg subsets in responders as compared with progressors or non-responders (MFI -55 vs.30, p=0.001) at both baseline and post-therapy were increased (Fig. B). This quantitative analysis indicated an existing difference in Tregs and distinct molecular dynamic changes in response to pembrolizumab between responders and progressors. To delineate the T cell characteristics in progressors and responders, we performed single-cell RNA-seq (SC-RNA-seq; 10X Genomics platform) using T (CD3+) cells enriched from PBMCs derived from three patients (1 responder: RS2; 2 progressors: CLL14, CLL17) before and after treatment. A total of ~10000 cells were captured and an average of 1215 genes was detected per cell. Using a clustering approach (Seurat V3.1.5), we identified 7 T cell clusters based on transcriptional signature (Fig.C). Responders had a larger fraction of Tregs (Cluster 5) as compared with progressors (p=0.03, Fig. D), and these Tregs showed an IFN-related gene signature (Fig. E). To determine any changes in the cellular circuitry in Tregs between responders and progressors, we used FOXP3, CD25, and CD127 as markers for Tregs in our SC-RNA-seq data. We saw a greater expression of FOXP3, CD25, CD127, in RS2 in comparison to CLL17 and CLL14. Gene set enrichment analysis (GSEA) revealed the upregulation of genes involved in lymphocyte activation and FOXP3-regulated Treg development-related pathways in the responder's Tregs (Fig.F). Together, the greater expression of genes involved in Treg activation may reduce the suppressive functions of Tregs, which led to the response to anti-PD1 treatment seen in RS2 consistent with Tregs in melanoma. To delineate any state changes in T cells between progressors and responder, we performed trajectory analysis using Monocle (R package tool) and identified enrichment of MYC/TNF/IFNG gene signature in state 1 and an effector T signature in state 3 For RS2 after treatment (p=0.003), indicating pembrolizumab induced proliferative and functional T cell signatures in the responder only. Further, our single-cell results were supported by the T cell receptor (TCR beta) repertoire analysis (Adaptive Biotechnology). As an inverse measure of TCR diversity, productive TCR clonality in CLL14 and CLL17 samples was 0.638 and 0.408 at baseline, respectively. Fifty percent of all peripheral blood T cells were represented by one large TCR clone in CLL14(progressor) suggesting tumor related T-cell clone expansion. In contrast, RS2(responder) contained a profile of diverse T cell clones with a clonality of 0.027 (Fig. H). Pembrolizumab therapy did not change the clonality of the three patients during the treatment course (data not shown). In summary, we identified enriched Treg signatures delineating responders from progressors on pembrolizumab treatment, paradoxical to the current understanding of T cell subsets in solid tumors. However, these data are consistent with the recent observation that the presence of Tregs suggests a better prognosis in Hodgkin lymphoma, Follicular lymphoma, and other hematological malignancies. Figure 1 Disclosures Kay: Pharmacyclics: Membership on an entity's Board of Directors or advisory committees, Research Funding; Oncotracker: Membership on an entity's Board of Directors or advisory committees; Rigel: Membership on an entity's Board of Directors or advisory committees; Juno Theraputics: Membership on an entity's Board of Directors or advisory committees; Agios Pharma: Membership on an entity's Board of Directors or advisory committees; Cytomx: Membership on an entity's Board of Directors or advisory committees; Astra Zeneca: Membership on an entity's Board of Directors or advisory committees; Morpho-sys: Membership on an entity's Board of Directors or advisory committees; Tolero Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees, Research Funding; Bristol Meyer Squib: Membership on an entity's Board of Directors or advisory committees, Research Funding; Acerta Pharma: Research Funding; Sunesis: Research Funding; Dava Oncology: Membership on an entity's Board of Directors or advisory committees; Abbvie: Research Funding; MEI Pharma: Research Funding. Ansell:AI Therapeutics: Research Funding; Takeda: Research Funding; Trillium: Research Funding; Affimed: Research Funding; Bristol Myers Squibb: Research Funding; Regeneron: Research Funding; Seattle Genetics: Research Funding; ADC Therapeutics: Research Funding. Ding:Astra Zeneca: Research Funding; Abbvie: Research Funding; Octapharma: Membership on an entity's Board of Directors or advisory committees; MEI Pharma: Membership on an entity's Board of Directors or advisory committees; alexion: Membership on an entity's Board of Directors or advisory committees; Beigene: Membership on an entity's Board of Directors or advisory committees; DTRM: Research Funding; Merck: Membership on an entity's Board of Directors or advisory committees, Research Funding. OffLabel Disclosure: pembrolizumab

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Increased Early Mortality after Fludarabine and Melphalan Conditioning with Peripheral Blood Grafts in Haploidentical SCT with Post-Transplant Cyclophosphamide

Blood ◽

10.1182/blood-2019-129872 ◽

2019 ◽

Vol 134 (Supplement_1) ◽

pp. 4496-4496 ◽

Cited By ~ 1

Author(s):

Luke Eastburg ◽

David A. Russler-Germain ◽

Ramzi Abboud ◽

Peter Westervelt ◽

John F. DiPersio ◽

...

Keyword(s):

Bone Marrow ◽

Stem Cell ◽

T Cell ◽

Board Of Directors ◽

Research Funding ◽

Early Mortality ◽

Cell Transplant ◽

Advisory Committees ◽

Post Transplant ◽

Equity Ownership

The use of post-transplant cyclophosphamide (PTCy) in the context of haploidentical stem cell transplant (haplo-SCT) has led to drastically reduced rates of Graft-vs-Host (GvH) disease through selective depletion of highly allo-reactive donor T-cells. Early trials utilized a reduced-intensity Flu/Cy/TBI preparative regimen and bone marrow grafts; however, relapse rates remained relatively high (Luznik et al. BBMT. 2008). This led to the increased use of myeloablative (MA) regimens for haplo-SCT, which have been associated with decreased relapse rates (Bashey et al. J Clin Oncol. 2013). Most studies have used a MA total body irradiation (TBI) based regimen for haplo-SCT. Preparative regimens using fludarabine and melphalan (FluMel), with or without thiotepa, ATG, and/or low dose TBI have also been reported using bone marrow grafts. Reports on the safety and toxicity of FluMel in the haplo-SCT setting with PTCy and peripheral blood stem cell (PBSC) grafts are lacking. In this two-center retrospective analysis, the safety/toxicity of FluMel as conditioning for haplo-SCT was evaluated. We report increased early mortality and toxicity using standard FluMel conditioning and PBSC grafts for patients undergoing haplo-SCT with PTCy. 38 patients at the University of Rochester Medical Center and the Washington University School of Medicine underwent haplo-SCT with FluMel conditioning and PBSC grafts between 2015-2019. Outcomes were measured by retrospective chart review through July 2019. 34 patients (89.5%) received FluMel(140 mg/m2). Two patients received FluMel(100 mg/m2) and two patients received FluMel(140 mg/m2) + ATG. The median age at time of haplo-SCT was 60 years (range 21-73). 20 patients were transplanted for AML, eight for MDS, two for PMF, two for NHL, and five for other malignancies. The median Hematopoietic Cell Transplantation-specific Comorbidity Index (HCT-CI) score was 4 (≥3 indicates high risk). 11 patients had a history of prior stem cell transplant, and 16 patients had active disease prior to their haplo-SCT. Seven patients had sex mismatch with their stem cell donor. Median donor age was 42 (range 21-71). 20 patient deaths occurred by July 2019 with a median follow up of 244 days for surviving patients. Nine patients died before day +100 (D100, "early mortality"), with a D100 non-relapse mortality (NRM) rate of 24%. Median overall and relapse free survival (OS and RFS, respectively) were 197 days (95% CI 142-not reached) and 180 days (95% CI 141-not reached), respectively, for the entire cohort. The 1 year OS and NRM were 29% and 50%. The incidence of grades 2-4cytokine release syndrome (CRS) was 66%, and 52% of these patients were treated with tocilizumab. CRS was strongly associated with early mortality, with D100 NRM of 36% in patients with grade 2-4 CRS compared to 0% in those with grade 0-1. The incidence of acute kidney injury (AKI) was 64% in patients with grade 2-4 CRS, and 8% in those without (p < 0.001). 28% of patients with AKI required dialysis. Grade 2-4 CRS was seen in 54% of patients in remission prior to haplo-SCT and in 92% of those with active disease (p = 0.02). Of the 9 patients with early mortality, 89% had AKI, 44% needed dialysis, and 100% had grade 2-4 CRS, compared to 31%, 10%, and 55% in those without early mortality (p = 0.002, p = 0.02, p = 0.01). Early mortality was not significantly associated with age, HCT-CI score, second transplant, disease status at transplant, total dose of melphalan, volume overload/diuretic use, or post-transplant infection. In conclusion, we observed a very high rate of NRM with FluMel conditioning and PBSC grafts for haplo-SCT with PTCy. The pattern of toxicity was strongly associated with grade 2-4 CRS, AKI, and need for dialysis. These complications may be mediated by excessive inflammation in the context of allo-reactive donor T-cell over-activation. Consistent with this, multiple groups have shown that FluMel conditioning in haplo-SCT is safe when using bone marrow or T-cell depleted grafts. Based on our institutional experiences, we would discourage the use of FluMel as conditioning for haplo-SCT with PTCy with T-cell replete PBSC grafts. Alternative regimens or variations on melphalan-based regimens, such as fractionated melphalan dosing or inclusion of TBI may improve outcomes but further study and randomized controlled trials are needed. This study is limited in its retrospective design and sample size. Figure Disclosures DiPersio: WUGEN: Equity Ownership, Patents & Royalties, Research Funding; Karyopharm Therapeutics: Consultancy; Magenta Therapeutics: Equity Ownership; Celgene: Consultancy; Cellworks Group, Inc.: Membership on an entity's Board of Directors or advisory committees; NeoImmune Tech: Research Funding; Amphivena Therapeutics: Consultancy, Research Funding; Bioline Rx: Research Funding, Speakers Bureau; Macrogenics: Research Funding, Speakers Bureau; Incyte: Consultancy, Research Funding; RiverVest Venture Partners Arch Oncology: Consultancy, Membership on an entity's Board of Directors or advisory committees. Liesveld:Onconova: Other: Data safety monitoring board; Abbvie: Membership on an entity's Board of Directors or advisory committees.

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