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To test the hypothesis that the outcome of hematopoietic stem cell (HSC) grafts is at least partially determined by the cellular composition of the graft, the National Marrow Donor Program (NMDP) analyzed the correlation of cellular phenotypes of unrelated grafts with graft outcome. Samples from 94 bone marrow (BM) and 181 peripheral blood progenitor cell (PBPC) grafts for transplantations at 40 U.S. transplant centers between 2003 and 2005 were analyzed at a single immunophenotyping reference laboratory. Samples were shipped from transplant centers upon receipt of graft. Graft cellular composition included analysis of leukocyte total cell numbers, and subsets of myeloid [CD34+, CD34+ CD38−], lymphoid [CD3+, CD3+ CD4+, CD3+ CD8+], and activated lymphoid cells [CD3+ CD25+, CD3+ CD69+, CD3+ HLA-DR+] coexpressing CD3+. There was substantial variability in the cellular composition of BM and PBPC grafts before and after graft processing by red blood cell (RBC) removal or plasma depletion in preparation for transplant. With BM grafts, cellular composition was not associated with hematopoietic recovery, graft-versus-host disease (GVHD), or survival. With PBPC grafts, survival rates were higher with CD34+ >5×106/kg, 59% compared to 34% with CD34+ ≤5×106/kg at 1 year. Platelet recovery was higher with PBPC containing CD3+ CD8+ >8×107/kg. Neutrophil recovery or GVHD could not be predicted by any cellular subsets of PBPC grafts. Although survival was superior with PBPC grafts containing >5×106 CD34+/kg, an optimal graft mix of myeloid, lymphoid, and activated lymphoid subsets was not identified. [Copyright &y& Elsevier]

We examined the effect of donor characteristics on graft failure (<5% donor chimerism within 3 months after transplantation), acute and chronic graft-versus-host disease (aGVHD, cGVHD), and survival after unrelated donor reduced-intensity conditioning (RIC) transplantation in 709 patients with hematologic malignancies. Donor–recipient pairs were HLA typed at HLA-A, -B, -C, and -DRB1 (allele-level). A total of 501 patients were >95% donor chimerism, 145 patients were 5% to 95%, and 63 patients were <5%. The only donor characteristic associated with transplantation outcome was donor–recipient HLA matching. One- or 2-loci mismatched transplants led to higher grade 2-4 (relative risk [RR] = 1.27, P = .035) and grade 3-4 (RR = 1.85, P < .001) aGVHD and 2-loci mismatched transplants higher mortality (RR = 2.22, P < .001). Graft failure was higher after transplantation of bone marrow (RR = 2.33, P = .002). Donor age, parity, and donor sex match were not associated with transplantation outcome. Donor–recipient HLA matching is the only donor characteristic predictive for survival after RIC regimens for hematologic malignancies. [ABSTRACT FROM AUTHOR]

Abstract: We evaluated effects of graft transport time on outcomes after transplantation of 938 unrelated donor bone marrow (BM) or 507 peripheral blood progenitor cells (PBPC) in patients with acute or chronic leukemia and myelodysplastic syndrome (MDS). BM grafts were collected at 107 centers and PBPC, 89 centers. Median time from end of collection to infusion was 14 hours for BM and 15 hours for PBPC. Platelet recovery was less likely in BM recipients when the interval from end of collection to receipt at transplant center was ≥20 hours (odds ratio 0.47, P = .010) and when the interval from receipt to infusion was ≥6 hours (odds ratio 0.57, P = .001). Mortality rates were higher in recipients of HLA-matched BM when the interval from end of collection to receipt at transplant center was ≥20 hours (relative risk 2.67, P < .001) after adjustment for other significant prognostic factors. Mortality after HLA-mismatched BM transplants was not associated with transport time. Transport times had no demonstrable effect on outcomes after PBPC transplants. These data support a general review of current transport procedures, especially for BM grafts requiring longer transport time and every effort made to minimize time from collection to infusion. [Copyright &y& Elsevier]

Abstract: Few studies have tested the benefits of using peripheral blood stem cell (PBSC) grafts versus bone marrow (BM) grafts for unrelated donor transplantation. Yet there has been a substantial change in clinical practice, with increasing numbers of adults receiving unrelated donor PBSC grafts. We compared outcomes after 331 PBSC and 586 BM transplants in adults with leukemia and myelodysplastic syndrome (MDS) who were followed for a median of 3 years after transplantation. PBSC recipients were less likely to have chronic myelogenous leukemia (CML) and more likely to have MDS, to have poor performance scores, and to be transplanted more recently. Outcomes were analyzed using Cox regression models. Rates of grades 2-4 acute graft-versus-host disease (GVHD) (58% versus 45%, P < .001) and chronic GVHD (cGVHD) (56% versus 42%, P < .001) were significantly higher with PBSC than with BM transplants. Rates of grade II-IV aGVHD were similar with PBSC and BM transplants. The 3-year probabilities of treatment-related mortality (TRM), leukemia recurrence, leukemia-free, and overall survival (OS) were similar in the 2 groups with 3-year leukemia-free survival rates of 30% and 32% after transplantation of PBSC and BM, respectively. Unlike results after HLA-matched sibling donor PBSC transplants, we did not identify a survival advantage with PBSC grafts in patients receiving unrelated donor transplants for advanced leukemia. The higher rate of cGVHD after PBSC transplants and, consequently, more frequent late adverse events warrant extended follow up of PBSC recipients. [Copyright &y& Elsevier]

Despite an array of approved agents for the treatment of multiple myeloma (MM), most patients eventually relapse after conventional treatments. The adoptive transfer of tumor-targeted T cells has demonstrated efficacy in the treatment of patients with chemo-refractory hematological malignancies including MM. While the majority of T cell-based immunotherapeutic studies in the clinic explore genetically modified T cells that target a single tumor-expressed antigen, we have developed a strategy to generate non-engineered T cell lines that simultaneously target multiple MM-expressed antigens, thereby reducing the risk of tumor immune evasion. We manufacture multiTAA-specific T cells targeting the tumor antigens PRAME, SSX2, MAGEA4, NY-ESO-1 and Survivin by culturing patient-derived PBMCs with autologous DCs loaded with pepmixes (15mer overlapping peptides) spanning all 5 target antigens in the presence of a Th1-polarizing cytokine cocktail. We have successfully generated multi-antigen-targeted lines for 19 patients, comprising a polyclonal mixture of CD4+ (28.9±7.2%) and CD8+ (56.6±7.2%) T cells reactive against 2 to 5 of the target antigens (by IFNg ELIspot), with no activity against non-malignant autologous targets (2±3% specific lysis; E:T 20:1). We assessed the clonal diversity using TCR vβ deep sequencing analysis and found that the majority (mean 79%; range: 59 to 95%) represented rare T cell clones that were unique to the ex vivo expanded cell line, thereby enabling in vivo tracking studies. To date we have infused 18 patients who had received a median of 4 lines of prior therapy at cell doses ranging from 0.5-2 × 107/m2 without prior lymphodepletion. Ten patients were refractory to their latest therapy and had active MM, while 8 were in remission at the time of infusion. At the 6 week assessment, of the 10 patients infused to treat active disease, 1 had a CR, 1 had a PR and 8 had SD. Seven of these 10 patients were infused >1 year ago. Although 2 of the 7 subsequently had disease progression, the remaining 5 continue to respond, with sustained CR (1), PR (2) or SD (2). Of the 8 patients in CR at the time of T cell infusion, all remained in CCR at the week 6 disease assessment and of the 6 evaluable patients who are >1 year post T cells, only 1 has relapsed. Clinical responses correlated with the emergence and persistence (>6mths) of "line-exclusive" tumor-reactive T cells in patient peripheral blood and marrow, as assessed by TCR deep sequencing. The expansion of product-derived clones was higher among patients with active MM than those in remission. No patient had infusion-related systemic- or neuro-toxicity. Thus, autologous multiTAA-targeted T cells directed to PRAME, SSX2, MAGEA4, NY-ESO-1 and Survivin can be safely administered to patients with MM, in whom they can subsequently be detected long-term in peripheral blood and marrow and where they produce sustained tumor responses. [ABSTRACT FROM AUTHOR]

Introduction HSCT is a curative option for patients with high-risk ALL but relapse remains the major cause of treatment failure. CD19 CAR T cells have shown remarkable efficacy in treating leukemic relapse post-HSCT but their use is limited to CD19+ malignancies and antigen negative relapses are increasingly being reported. To overcome these limitations, we developed a strategy to generate donor-derived T cell lines simultaneously targeting PRAME, WT1 and Survivin (multiTAAs) expressed by both B and T ALL for adoptive transfer to high risk HSCT recipients. Methods We generated donor multiTAA-T cells by culturing PBMCs with autologous DCs loaded with a mastermix of pepmixes spanning all 3 target antigens in the presence of a Th1-polarizing/pro-proliferative cytokine cocktail. Results To date, we have generated 14 clinical multiTAA-T cell lines comprising CD3+ T cells (mean 94±9%) with a mixture of CD4+ (mean 21±28%) and CD8+ (mean 52±24 %) cells, which expressed central and effector memory markers and recognized the targeted antigens based on IFNg ELIspot analysis. None of the lines reacted against non-malignant patient-derived cells (4±3% specific lysis; E:T 20:1) - a study release criterion. Thus far we have treated 10 high-risk ALL patients to prevent disease relapse post-transplant (Table 1). Infusions were well tolerated with no dose-limiting toxicity, GVHD, CRS or other adverse events. Two patients were not evaluable per study criteria as they received >0.5mg/kg of steroids within 4 weeks of infusion and were replaced. Seven of the 8 remaining patients infused remain in CCR a median of 9 months post-infusion (range 1-23 months). We detected the expansion of tumor-reactive T cells in patient peripheral blood post-infusion against both targeted (WT1, Survivin, PRAME) and non-targeted antigens (SSX2, MAGE-A4, -A1, -A2B, -C1, MART1, AFP and NYESO1) reflecting epitope and antigen spreading. The single patient who relapsed showed no evidence of tumor-directed T cell expansion despite receiving 3 additional infusions at 4 week intervals. Conclusion Infusion of donor-derived multiTAA-T cells to patients with ALL post-HSCT is feasible, safe and as evidenced by expansion and antigen spreading in patients, may contribute to disease control. This strategy maybe a promising approach to prevent leukemic relapse after HSCT. [ABSTRACT FROM AUTHOR]

Allogeneic HSCT (alloHSCT) can be curative for AML, but >40% of patients relapse, and then have a median OS of <6mths. Donor lymphocyte infusions (DLIs) can treat relapsed disease, but durable responses are rare and associated life-threatening GvHD frequent. Thus, to enhance GvL and minimize GvHD our group developed a strategy to selectively amplify donor-derived T cells specific for the leukemia associated antigens (LAA) WT1, PRAME, Survivin and NYESO1. We are now conducting a clinical trial to test the safety and efficacy of these multiLAA-specific T cells to prevent (Arm A) or treat (Arm B) AML/MDS relapse. To date we have made 25 lines, which were polyclonal [CD4+ (35.1±0.2%) and CD8+ (41.03%±0.23)], enriched for tumor-directed clonotypes based on deep sequencing and confirmed to be LAA-reactive as determined by IFNg ELIspot [PRAME: mean 123 SFC/105, range 0-770; WT1: 57, 0-561; Survivin: 11, 0-90; NYESO1: 3, range 0-284)]. Importantly, none of the lines manifest alloreactivity against patient PHA blasts, (2±3.6% specific lysis at E:T 20:1, n=25). We have now infused 19 pts with adverse-risk AML/MDS (12 on Arm A and 7 on Arm B) at cell doses ranging from 0.5-2 × 107 cells/m2 with no immediate toxicities. All 12 pts infused as adjuvant remained in CCR at the 4 week disease assessment but 4 later relapsed (at 4, 8, 9 and 10mths post-infusion). Of these 4, 1 subsequently entered a CR with additional mLAA-T cells, 2 (with CNS relapse) were successfully treated with intrathecal chemotherapy, and the patient with bone marrow relapse was treated with systemic chemotherapy and another alloHSCT. Thus, 11/12 pts infused as adjuvant therapy are currently alive and in CR [median 12mths] while 1 pt died in CR from influenza 12 mths post-infusion. On the active disease arm all 7 pts infused had relapsed AML and were resistant to a median of 3 lines of prior therapies. One achieved a durable CR, 1 achieved a PR (enabling a second alloHSCT), and 2 patients derived transient benefit - 1 had an improvement in ANC (0 pre-T cells to >500 at wk8) with concomitant decline in blood and platelet transfusions while another had clearance of peripheral blasts (56% to 0%) with a <50% decline in marrow blasts. Three pts developed PD, failed subsequent lines of therapies and died 4-12mths post-infusion. Clinical responses correlated with the emergence and persistence (>9mths) of "line-exclusive" tumor-reactive T cells, as assessed by longitudinal clonotype tracking using deep sequencing. The expansion of product-derived clones was higher in patients who responded as compared with those that progressed post-infusion as confirmed by both ELIspot and deep sequencing. Notably, no patient had infusion-related CRS, neurotoxicity or GVHD. Thus, mLAA-targeted T cells directed to PRAME, WT1, NYESO1 and Survivin can be safely administered to patients with AML/MDS, in whom they can subsequently be detected long-term and produce sustained responses. [ABSTRACT FROM AUTHOR]

Almost all HL and some NHL express the CD30 antigen, and monoclonal antibodies (mAb) targeting CD30 (e.g. brentuximab) produce objective antitumor responses. However, mAb have limited bio-distribution and their benefits may be short-lived. We therefore expressed the antigen binding domain of a CD30 mAb as part of a chimeric antigen receptor (CAR) on T cells, coupled to the CD28 and ζ chain endodomains. We have previously published results of a phase 1 study of autologous CD30.CAR-T cells (CD30.CARTs) infused in patients with relapsed/refractory CD30+ HL or NHL without preceding cytoreductive chemotherapy (Ramos et al. , J Clin Invest 2017). Of 6 patients with relapsed active HL, 1 entered complete remission (CR) that has lasted more than 3 years, and 3 had transient stable disease. We have now incorporated cytoreductive chemotherapy (cyclophosphamide and fludarabine) prior to CD30 CAR T infusion to discover if we can enhance the in vivo expansion and anti-tumor activity of these CD30.CARTs, (RELY-30, NCT02917083). Our preliminary results suggest a substantial improvement in efficacy. We have manufactured CD30.CARTs for 18 patients using retroviral transduction. Cell products comprise >98% T cells with a final transduction efficiency of 97.6%±1.8%. Ten relapsed/refractory HL patients have received these CD30.CARTs under the RELY-30 trial, 7 of whom had relapsed or progressed after treatment with brentuximab. Three patients have been treated on dose level (DL) 1 (2 × 107 CD30.CAR+ T cells/m2), 6 on DL2 (1 × 108) and 1 on DL3 (2 × 108). All patients received preceding lymphodepleting chemotherapy (cyclophosphamide 500 mg/m2 and fludarabine 30 mg/m2 daily for 3 days). CART infusions were associated with grade 1 cytokine release syndrome in 4 of the patients, and a transient maculopapular rash in 6 of the patients, starting approximately one week after administration. The molecular signal from CARTs, assessed by Q-PCR in peripheral blood, peaked at 1-2 weeks following infusion, but dropped progressively after 4 weeks and decreased to the limit of detection by 6 months. The signal level was dose dependent, with a peak average of 19,371 copies/mg DNA in patients treated on DL2 versus 7,132 copies/mg for DL1; this expansion was 45 and 119-fold higher, respectively, than patients receiving the same CART dose without prior cytoreduction in our previous trial. Nine patients have been evaluated at 6 weeks after infusion. Six have had a CR lasting up to >9 months, while 3 patients had disease progression. Hence, infusion of CD30.CARTs after cytoreductive chemotherapy is well tolerated at the doses used. Inclusion of cytoreduction pre-infusion substantially improves CD30.CART expansion and appears associated with superior anti-tumor activity in relapsed patients (6/9 CR versus 1/6 CR, P = 0.04). [ABSTRACT FROM AUTHOR]

Immunotherapy is emerging as a potent treatment for a range of hematologic malignancies including lymphomas. Indeed adoptive transfer of T cells genetically engineered to express the CD19 CAR has now received FDA approval for the treatment of refractory diffuse large B cell lymphomas (DLBCL). We have developed a non-engineered T cell therapy to treat patients with Hodgkin's and non-Hodgkin's lymphoma (HL/NHL) using single T cell lines that simultaneously target the tumor-associated antigens (TAAs) PRAME, SSX2, MAGEA4, NY-ESO-1 and Survivin. We can consistently prepare these lines by culturing PBMCs in the presence of a Th1-polarizing/pro-proliferative cytokine cocktail, and adding autologous pepmix-loaded DCs as APCs. The use of whole antigen overcomes the HLA restriction imposed by the use of transgenic TCRs specific for single peptides, while targeting multiple antigens simultaneously should reduce the risk of tumor immune evasion. We have generated 42 clinical-grade multiTAA T cell lines, comprising CD3+ T cells (mean 98±1.1%) with a mixture of CD4+ (mean 48±4.3%) and CD8+ (mean 37±4%) T cells that recognize the targeted antigens PRAME, SSX2, MAGEA4, NY-ESO-1 and Survivin (range 0-463, 0-496, 0-330, 0-379 and 0-304 SFU/2 × 105, respectively in IFNg ELIspot). None of the lines reacted against non-malignant autologous cells (3±3.8% specific lysis; E:T 20:1). We have treated 33 patients: 13 with HL, 17 with aggressive NHL (DLBCL, mantle cell, T cell lymphomas) and 3 with indolent NHLs (FL, marginal zone lymphoma). Patients were infused with 0.5-2 × 107 multiTAA-T cells/m2 and did not receive lymphodepleting chemotherapy. Of 18 patients who were infused as adjuvant all but 2 remain in remission (range 3-42 months). Fifteen patients have received multiTAA-specific T cells to treat active disease, all of whom had failed a median of 4 lines of prior therapy. Of these, 5 had transient disease stabilization followed by disease progression, 4 have ongoing stable disease at 3-18 months post-multiTAA-specific T cells while the remaining 6 (3 with HL and 3 with DLBCL) had complete and durable responses (4 to 41 months), as assessed by PET. These clinical responses correlated with the detection of tumor-reactive T cells in patient peripheral blood post-infusion directed against both targeted antigens as well as non-targeted TAAs including MAGEA2B and MAGE C1, indicating induction of antigen/epitope spreading. Notably, no patient, including the complete responders, had infusion-related systemic- or neuro-toxicity. Thus, infusion of autologous multiTAA-targeted T cells directed to PRAME, SSX2, MAGEA4, NY-ESO-1 and Survivin has been safe, and despite concerns that endogenous T cells directed against shared/cancer testis antigens would be of insufficient affinity to produce prolonged benefit our multiTAA-specific T cells were able to induce durable complete responses in patients with lymphomas. [ABSTRACT FROM AUTHOR]