scholarly journals Characterization of Tumor Reactivity of Human Vγ9Vδ2 γδ T Cells In Vitro and in SCID Mice In Vivo

2004 ◽  
Vol 173 (11) ◽  
pp. 6767-6776 ◽  
Author(s):  
Dieter Kabelitz ◽  
Daniela Wesch ◽  
Elke Pitters ◽  
Margot Zöller
Keyword(s):  
T Cells ◽  
Γδ T Cells ◽  
Scid Mice

γδ T cells for immune therapy of patients with lymphoid malignancies

Blood ◽  
2003 ◽  
Vol 102 (1) ◽  
pp. 200-206 ◽  
Author(s):  
Martin Wilhelm ◽  
Volker Kunzmann ◽  
Susanne Eckstein ◽  
Peter Reimer ◽  
Florian Weissinger ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Antitumor Activity ◽  
Low Dose ◽  
Γδ T Cells ◽  
Low Grade ◽  
Treatment Schedule ◽  
Γδ T Cell

Abstract There is increasing evidence that γδ T cells have potent innate antitumor activity. We described previously that synthetic aminobisphosphonates are potent γδ T cell stimulatory compounds that induce cytokine secretion (ie, interferon γ [IFN-γ]) and cell-mediated cytotoxicity against lymphoma and myeloma cell lines in vitro. To evaluate the antitumor activity of γδ T cells in vivo, we initiated a pilot study of low-dose interleukin 2 (IL-2) in combination with pamidronate in 19 patients with relapsed/refractory low-grade non-Hodgkin lymphoma (NHL) or multiple myeloma (MM). The objectives of this trial were to determine toxicity, the most effective dose for in vivo activation/proliferation of γδ T cells, and antilymphoma efficacy of the combination of pamidronate and IL-2. The first 10 patients (cohort A) who entered the study received 90 mg pamidronate intravenously on day 1 followed by increasing dose levels of continuous 24-hour intravenous (IV) infusions of IL-2 (0.25 to 3 × 106 IU/m2) from day 3 to day 8. Even at the highest IL-2 dose level in vivo, γδ T-cell activation/proliferation and response to treatment were disappointing with only 1 patient achieving stable disease. Therefore, the next 9 patients were selected by positive in vitro proliferation of γδ T cells in response to pamidronate/IL-2 and received a modified treatment schedule (6-hour bolus IV IL-2 infusions from day 1-6). In this patient group (cohort B), significant in vivo activation/proliferation of γδ T cells was observed in 5 patients (55%), and objective responses (PR) were achieved in 3 patients (33%). Only patients with significant in vivo proliferation of γδ T cells responded to treatment, indicating that γδ T cells might contribute to this antilymphoma effect. Overall, administration of pamidronate and low-dose IL-2 was well tolerated. In conclusion, this clinical trial demonstrates, for the first time, that γδ T-cell–mediated immunotherapy is feasible and can induce objective tumor responses. (Blood. 2003;102:200-206)

scholarly journals Vγ9Vδ2 T cells as a promising innovative tool for immunotherapy of hematologic malignancies

2011 ◽  
Vol 4 (4) ◽  
pp. 211
Author(s):  
Serena Meraviglia ◽  
Carmela La Mendola ◽  
Valentina Orlando ◽  
Francesco Scarpa ◽  
Giuseppe Cicero ◽  
...  
Keyword(s):  
T Cells ◽  
T Lymphocytes ◽  
Ex Vivo ◽  
Γδ T Cells ◽  
Hematologic Malignancies ◽  
Cytolytic Activity ◽  
Cell Therapies ◽  
Stressed Cells

The potent anti-tumor activities of γδ T cells, their ability to produce pro-inflammatory cytokines, and their strong cytolytic activity have prompted the development of protocols in which γδ agonists or ex vivo-expanded γδ cells are administered to tumor patients. γδ T cells can be selectively activated by either synthetic phosphoantigens or by drugs that enhance their accumulation into stressed cells as aminobisphosphonates, thus offering new avenues for the development of γδ T cell-based immunotherapies. The recent development of small drugs selectively activating Vγ9Vδ2 T lymphocytes, which upregulate the endogenous phosphoantigens, has enabled the investigators to design the experimental approaches of cancer immunotherapies; several ongoing phase I and II clinical trials are focused on the role of the direct bioactivity of drugs and of adoptive cell therapies involving phosphoantigen- or aminobisphosphonate-activated Vγ9Vδ2 T lymphocytes in humans. In this review, we focus on the recent advances in the activation/expansion of γδ T cells in vitro and in vivo that may represent a promising target for the design of novel and highly innovative immunotherapy in patients with hematologic malignancies.<br />

scholarly journals Reduced Immune Response to Borrelia burgdorferi in the Absence of γδ T Cells

2011 ◽  
Vol 79 (10) ◽  
pp. 3940-3946 ◽  
Author(s):  
Cuixia Shi ◽  
Bikash Sahay ◽  
Jennifer Q. Russell ◽  
Karen A. Fortner ◽  
Nicholas Hardin ◽  
...  
Keyword(s):  
Immune Response ◽  
T Cells ◽  
Borrelia Burgdorferi ◽  
Adaptive Immune Response ◽  
Γδ T Cells ◽  
Content Type ◽  
Adaptive Immune ◽  
Cytokines And Chemokines

ABSTRACTLittle is known regarding the function of γδ T cells, although they accumulate at sites of inflammation in infections and autoimmune disorders. We previously observed that γδ T cellsin vitroare activated byBorrelia burgdorferiin a TLR2-dependent manner. We now observe that the activated γδ T cells can in turn stimulate dendritic cellsin vitroto produce cytokines and chemokines that are important for the adaptive immune response. This suggested thatin vivoγδ T cells may assist in activating the adaptive immune response. We examined this possibilityin vivoand observed that γδ T cells are activated and expand in number duringBorreliainfection, and this was reduced in the absence of TLR2. Furthermore, in the absence of γδ T cells, there was a significantly blunted response of adaptive immunity, as reflected in reduced expansion of T and B cells and reduced serum levels of anti-Borreliaantibodies, cytokines, and chemokines. This paralleled a greaterBorreliaburden in γδ-deficient mice as well as more cardiac inflammation. These findings are consistent with a model of γδ T cells functioning to promote the adaptive immune response during infection.

scholarly journals In Vitro Expanded γδ T Cells Derived from Cord Blood Induce Potent and Specific Cytotoxicity Against Human Acute Myeloid Leukemia Cells

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 2168-2168
Author(s):  
Alice MS Cheung ◽  
Kar Wai Tan ◽  
Dian Yan Guo ◽  
Su-Ann Goh ◽  
Amanda SY Lau ◽  
...  
Keyword(s):  
T Cells ◽  
Myeloid Leukemia ◽  
Γδ T Cells ◽  
Leukemic Cell ◽  
In Vitro Cytotoxicity ◽  
Nsg Mice ◽  
Specific Cytotoxicity ◽  
Post Infusion

Abstract Conventional chemotherapeutic regimens for acute myeloid leukemia (AML) patients have demonstrated unsurpassed efficacy in the past decades, but are far from optimal with many patients experiencing multiple disease recurrence or intolerant of the intensive chemotoxicity. Variations in the treatment scheme as well as the use of alternative, targeted agents have been pursued with limited success. This is partly ascribed to the highly heterogeneous nature of the disease comprising a dynamic repertoire of evolving leukemic clones that are both molecularly and biologically diverse, making it difficult to achieve complete disease eradication without inducing adverse off-target effects. In this regard, cellular immunotherapy has emerged as a plausible alternative, leveraging on the diversity and degeneracy of the tumor antigen-recognizing receptor complex expressed by immune cells. In particular, there is a growing interest in the specific anti-leukemia efficacy of the innate-like γδ T cells, prompted by the association of an increased number of donor derived γδ T cells (specifically the Vδ1+ subtype) in allogeneic hematopoietic stem cell transplant (HSCT) patients with improved disease control in the absence of significant graft-versus-host disease (GvHD). We therefore hypothesize that these allogeneic γδ T cells exhibit potent leukemia specific cytotoxicity and serve as an effective treatment for AML. Given the rapid availability and widespread use of cord blood (CB) as an alternative for allogeneic HSCT, we first characterized and explored the potential of expanding CB-derived γδ T cells in vitro. Compared to mobilized peripheral blood (mPB), there is a significantly lower level of γδ T cell within CB mononuclear cells (MCs) (0.61% ± 0.36% in CB vs 4.95% ± 3.83% in mPB, p<0.001). However, the fraction of Vδ1+ subset within the γδ T cells in CB is >3.5-fold higher than that in mPB (56.05% ± 9.49% in CB vs 14.54% ± 12.2% in mPB, p<0.001). Importantly, while >90% of the Vδ1+ T cells in CB are of naive or central memory phenotype, more than 40% of these cells in mPB show effector memory expression. We established that optimal in vitro expansion of CB-derived γδ T cells requires direct contact to a mixture of irradiated PBMCs and Epstein-Barr virus-transformed lymphoblastoid cell line (EBV-LCL) at a fixed ratio in the gas-permeable G-Rex culture flask. Under these conditions, we were able to achieve up to 5,200-fold expansion of the starting γδ T cells over a period of 21 days. These cells exhibit potent in vitro cytotoxicity against a range of human AML cell lines, including K562, MOLM-14, MV4-11 and NOMO-1, as well as primary patient samples in a dose dependent manner. In contrast, there is minimal in vitro cytotoxicity against CD34+ cells isolated from allogeneic CB samples even at the highest effector-to-target cell (E:T) ratio tested. Infusion of the expanded γδ T cells into NOD/SCID/IL2Rγ-/- (NSG) mice at 3 weeks post-transplantation of a FLT3-ITD+ AML patient sample (P1) resulted in a significant decrease in leukemic cell engraftment in 40% of the γδ T cells-treated mice (87.46 ± 2.25% in non-treated vs 74.85 ± 1.55% in γδ T cells-treated mice, p=0.022). In a separate experiment, infusion into NSG mouse that was engrafted with low level (0.1%) of a different FLT3-ITD+ AML patient sample (P2) maintained the leukemic cell level low at 0.1% at 4 weeks post-infusion, as opposed to the >15-fold increase in leukemic burden (1.76%) seen in the untreated mouse. Consistent with our in vitro finding, infusion of up to 5 x 108 expanded CB derived γδ T cells/kg failed to induce severe GvHD symptoms in NSG mice engrafted with allogeneic human CB cells up to 8 weeks post-infusion, with no significant effect on the level of in vivo regenerated human myeloid and lymphoid cells, as well as colony-forming cells (CFCs). In summary, our data demonstrates that in vitro expanded CB derived γδ T cells show potent AML-specific cytotoxicity both in vitro and in vivo, making it a promising alternative cell source for immunotherapy. Further investigations to enhance the mechanistic understanding would be needed to seed for future clinical translation. Disclosures Hwang: Pfizer: Honoraria, Other: Travel support; MSD: Honoraria, Other: Travel support; BMS: Honoraria, Other: Travel support; Novartis: Honoraria, Other: Travel support; Celgene: Honoraria, Other: Travel support; Roche: Honoraria, Other: Travel support; Janssen: Honoraria, Other: Travel support; Sanofi: Honoraria, Other: Travel support.

scholarly journals Murine γδ T Lymphocytes Elicited duringPlasmodium yoelii Infection Respond to PlasmodiumHeat Shock Proteins

1999 ◽  
Vol 67 (1) ◽  
pp. 57-63 ◽  
Author(s):  
Jeffrey Kopacz ◽  
Nirbhay Kumar
Keyword(s):  
T Cells ◽  
Γδ T Cells ◽  
Plasmodium Yoelii ◽  
Host Responses ◽  
Shock Proteins ◽  
Cellular Immune ◽  
Proliferation In Vitro

ABSTRACT γδ T cells accumulate during Plasmodium infections in both murine and human malarias. The biological role of these cells and the antigens that they recognize are not clearly understood, although recent findings indicate that γδ T cells in general influence both innate and antigen-specific adaptive host responses. We examined the accumulation of γδ T cells elicited during infection with virulent and avirulent Plasmodium yoelii parasites in relatively susceptible and resistant strains of mice. Our results indicated that in nonlethal malaria infections, γδ T cells comprise a larger proportion of splenic T cells than in lethal infections and that only a live infection is capable of inducing an increase in the percentage of γδ T cells in vivo. Furthermore, we demonstrate that γδ T cells elicited during a P. yoelii infection respond by proliferation in vitro to P. falciparum heat shock proteins (HSPs) of 60 and 70 kDa, suggesting a possible immunological involvement of parasite HSPs in this arm of the cellular immune response during malarial infection in mice.

scholarly journals Characterization of Candidate Live Oral Salmonella typhi Vaccine Strains Harboring Defined Mutations inaroA, aroC, and htrA

1999 ◽  
Vol 67 (2) ◽  
pp. 700-707 ◽  
Author(s):  
David C. Lowe ◽  
Tor C. Savidge ◽  
Derek Pickard ◽  
Lars Eckmann ◽  
Martin F. Kagnoff ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Salmonella Typhi ◽  
Scid Mice ◽  
Intestinal Epithelial ◽  
Human Intestinal Epithelial Cells ◽  
U937 Macrophage ◽  
Ht 29

ABSTRACT The properties of two candidate Salmonella typhi-based live oral typhoid vaccine strains, BRD691 (S. typhi Ty2 harboring mutations in aroA and aroC) and BRD1116 (S. typhi Ty2 harboring mutations inaroA, aroC, and htrA), were compared in a number of in vitro and in vivo assays. BRD1116 exhibited an increased susceptibility to oxidative stress compared with BRD691, but both strains were equally resistant to heat shock. Both strains showed a similar ability to invade Caco-2 and HT-29 epithelial cells and U937 macrophage-like cells, but BRD1116 was less efficient at surviving in epithelial cells than BRD691. BRD1116 and BRD691 were equally susceptible to intracellular killing within U937 cells. Similar findings were demonstrated in vivo, with BRD1116 being less able to survive and translocate to secondary sites of infection when inoculated into the lumen of human intestinal xenografts in SCID mice. However, translocation of BRD1116 to spleens and livers in SCID mice occurred as efficiently as that of BRD691 when inoculated intraperitonally. The ability of BRD1116 to increase the secretion of interleukin-8 following infection of HT-29 epithelial cells was comparable to that of BRD691. Therefore, loss of the HtrA protease inS. typhi does not seem to alter its ability to invade epithelial cells or macrophages or to induce proinflammatory cytokines such as IL-8 but significantly reduces intracellular survival in human intestinal epithelial cells in vitro and in vivo.

Quantitative and Functional Alterations of the Gamma Delta T-Cell Subset within Follicular Lymphoma Microenvironment.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 2601-2601
Author(s):  
Sophie de Guibert ◽  
Jean-Baptiste Thibert ◽  
Céline Bonnaventure ◽  
Patricia Ame-Thomas ◽  
Céline Pangault ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Follicular Lymphoma ◽  
Peripheral Blood ◽  
Γδ T Cells ◽  
Γδ T Cell ◽  
Ifn Γ ◽  
Nonpeptide Antigens

Abstract T cells carrying a γδ TCR account for less than 5% of CD3pos T cells in healthy individuals but are key effectors of innate immunity through the recognition of some unprocessed nonpeptide antigens of both self and foreign origin. Whereas the Vδ2 subpopulation represents more than 70% of peripheral blood γδ T cells, the Vδ1 subset is mainly located in the mucosal tissue. Increasing evidence suggest that γδ T cells have potent antitumor activity and are implicated in the defense against some haematological and epithelial malignancies. Moreover, Vδ2 T cells constitute an attractive immunotherapy strategy since they could be expanded and activated both in vivo and in vitro using synthetic phosphoantigens and aminobiphosphonates. Such strategies are currently tested in preliminary clinical trials, notably in follicular lymphoma (FL). However, an exhaustive phenotypic and functional characterisation of γδ T cells in this disease, including tumor infiltration, is still lacking. We first explored the composition of FL microenvironment using a multicolour flow cytometry analysis. We observed a significant decrease in the percentage of myeloid (LinnegCD11cposHLADRpos) and plasmacytoid (LinnegCD123posHLADRpos) dendritic cells (P = .0011 and P &lt; .0001, respectively) in FL compared to normal secondary lymphoid organs. In addition, among CD3pos T cells, the proportion of follicular helper T cells (CD4posCXCR5posICOShi) was increased (P = .001) whereas regulatory T-cell (CD4posCD25posfoxp3pos) frequency was not altered. When considering the γδ T-cell compartment, we first highlighted a reduction of the Vδ2 subset in normal tonsils (Vδ2 = 23.48 ± 0.15% of γδ T cells, n = 11) when compared with peripheral blood. Remaining non-δ2 γδT cells were predominantly δ1 T cells. More importantly, infiltrating γδ T cells were significantly decreased in lymph node biopsies from FL patients (mean = 0.48 ± 0.4% of CD3pos T cells; n = 27) when compared both to normal tonsils (mean = 2.49 ± 1.6% of CD3pos T cells; n = 33) (P &lt; .0001) and reactive lymph nodes (mean = 2.64 ± 2.6% of CD3pos T cells; n = 9) (P = .0009). This reduction affected both the Vδ1 and Vδ2 T-cell subsets. The functionality of γδ T cells was then assessed by the measurement of cell expansion and production of IFN-γ upon stimulation with the isopentenyl pyrophosphate (IPP) phosphoantigen. Amplification rate in vitro reached 14.6 ± 4.6 fold in tonsils (n = 10) but only 4.36 ± 1.9 fold in FL samples (n = 7) (P &lt; .002) after 5 days of culture in the presence of IPP + IL-2 + IL-15. When focusing on the δ2 subset, this difference was further increased with a 40-fold amplification in tonsil and a 3-fold amplification in FL samples (P = .0004). Evaluation of IFN-γ production using ELISPOT assay revealed a high heterogeneity among tumor samples since 1 to 40% of δ2 T cells were able to respond to IPP stimulation (n = 7). Preliminary data argued for an association between the quantity and the functionality of γδ T cells in FL tumors. In conclusion, we reported an alteration of γδ T cell frequency and functionality within FL tumor niche. The next purpose will be to correlate these in vitro defects with in vivo clinical responses to immunotherapy strategies targeting γδ T cells.

In Vivo Eradication of Human Clonigenic Acute Lymphoblastic Leukemic Cells Expressing the Wilms Tumor Protein, WT-1, by HLA Restricted WT-1 Specific T-Cells in NOD/SCID Mice Monitored by Bioluminescent Imaging.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 581-581
Author(s):  
Ekaterina Doubrovina ◽  
Mikhail Doubrovin ◽  
Elena Kanaeva ◽  
Richard J. O’Reilly
Keyword(s):  
T Cells ◽  
Wilms Tumor ◽  
Scid Mice ◽  
Leukemic Cells ◽  
Luciferase Reporter ◽  
Bioluminescent Imaging ◽  
Acute Leukemias ◽  
Post Infusion

Abstract WT-1 is expressed in 60–80% of acute leukemias, CML and high risk forms of MDS. Its expression has been hypothesized to be critical to the growth or survival of leukemic stem cells. Previously, alloreactive HLAA0201− T-cells recognizing a complex of WT-1 peptide and HLA A0201 were reported to prevent growth of leukemic HLA A0201+ CD34+ Ph+CML progenitor cells in NOD/ SCID mice (Transplantation, vol 75, No9, 2003). In this study, we have assessed the capacity of HLA-restricted, WT-1 peptide specific CTL (WT1-CTL) lacking alloreactivity to prevent the outgrowth of a human acute preB-lymphocytic leukemia (B-ALL)in NOD/SCID mice. This leukemia contained 65% of the blasts expressed WT-1 as determined by FACS analysis. For these studies the leukemic cells were transduced to express a luciferase reporter gene, permitting sequential monitoring of growth in vivo by bioluminescent imaging. WT-1 specific T-cells were generated from normal HLA A0201+ donor PBMC by in vitro sensitization with autologous dendritic cells loaded with the immunogenic HLA A0201 binding WT-1 peptide, RMFPNAPYL, and shown to be selectively cytotoxic against HLA A0201+WT-1+ leukemias and peptide loaded PHA blasts. T-cells from the same donor sensitized with autologous EBV BLCL and exhibiting HLA A0201 restricted EBV-specific cytotoxic activity served as controls. WT-1-CTL or EBV CTL were co-incubated in vitro with the WT-1+ HLA A0201+ BALL-LUC at a 4:1 effector target ratio for 7 hours at 37°C. Thereafter, separate groups of 5 NOD/SCID mice received intravenous infusions of cells from each of the co-cultures, at doses providing 12 × 106 WT1 CTL or EBVCTL and 3 × 106 BALL-LUC cells/mouse. A third group received 3×106 BALL-LUC alone. Leukemia growth was monitored at 2–3 day intervals from day 1–45 post infusion. In all 3 groups, BALL-LUC could be detected in the thorax by imaging at day 1. In mice treated with BALL-LUC alone or together with EBV-CTL, signal accumulation in the thorax increased steadily through 45 days of observation. By day 17, BALL-LUC were also detected throughout the head, abdomen and pelvis, and thereafter also increased until sacrifice at day 45. Autopsy confirmed presence of leukemic nodules in the lung and leukemic cells in blood, spleen and marrow as well as other organs. In contrast, in mice treated with WT1-CTL+ BALL LUC, signal intensity in lung decreased by day 4. In 4/5 of these mice, BALL-LUC could not be detected thereafter. In one mouse from this group, BALL-LUC were first detected in the head 31 days post infusion. At autopsy on day 45, this mouse had detectable BALL in the skull but in no other sites. WT-1 expression of residual leukemic cells is being analyzed. The other mice treated with WT-1 CTL had no detectable residual disease. These results suggest that clonogenic BALL cells express WT-1 and are susceptible to eradication in vivo by WT-1 peptide specific cytotoxic T-cells. The elimination of such clonogenic leukemic cells is sufficient to prevent subsequent development of leukemia.

Cytotoxicity Is Not Required for the Anti-Leukemia Efficacy of Human H-Y-Specific CD4+ T Lymphocytes In Vivo.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 3553-3553
Author(s):  
Attilio Bondanza ◽  
Lothar Hambach ◽  
Zohara Aghai ◽  
Monica Casucci ◽  
Bart Nijmeijer ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Lymphocytes ◽  
Cell Clone ◽  
Scid Mice ◽  
Helper T Cell ◽  
T Cell Clone ◽  
Control Clone

Abstract Abstract 3553 Poster Board III-490 Introduction Minor histocompatibility antigens (mHag) play a major role in the graft-versus-leukemia (GvL) effect following HLA-matched allogeneic hemopoietic cell transplantation (allo-HCT). Clinically, the GvL effect coincides with the emergence of mHag-specific CD8+ cytotoxic T lymphocytes (CTL). Experimentally, targeting a single mHag with human CD8+ CTL has a major anti-leukemia effect in NOD/scid mice. Altogether, these observations suggest that mHag-specific cytotoxicity by CD8+ T cells is an important component of the GvL effect. In contrast, little is known on the contribution of mHag-specific CD4+ T cells. Female-to-male allo-HCT is characterized by a low rate of leukemia relapse, indicating that H-Y-encoded mHag are potent leukemia-regression antigens. Earlier, we described a DRB3*0301-restricted H-Y mHag epitope inducing CD4+ helper T-cell responses in H-Y-mismatched HLA-matched allo-HCT. Aim: The aim of this study is to elucidate the role of mHag-specific human CD4+ T lymphocytes on the GvL effect. Methods The ALL-CM leukemia cell line, derived from a male (i.e. H-Y+) HLA-A0201+, DRB30301+ patient, reproducibly engrafts in NOD/scid mice after administration of 10×106 cells. Both an HLA-A0201-restricted H-Y-specific CD8+ CTL clone and the DRB30301-restricted H-Y-specific CD4+ helper T-cell clone that we earlier described were used to investigate the anti-leukemia efficacy of CD8+ and CD4+ T cells in NOD/scid mice. Results In vitro, the CD8+ H-Y specific CTL clone was highly cytotoxic against the ALL-CM leukemia. The H-Y specific CD4+ helper T-cell clone did not lyse the leukemia, but produced IFN-γ upon recognition. Infusion of the H-Y-specific CD8+ CTL clone (25×106 cells/mouse) 3 days after ALL-CM leukemia challenge significantly delayed leukemia progression by 3 weeks compared to a CMV-specific CD8+ CTL control clone (p<0,001). Despite no measurable in vitro cytotoxicity, the H-Y-specific CD4+ helper T-cell clone (25×106 cells/mouse) delayed leukemia progression by 2 weeks compared to a leukemia non-reactive HLA-DR1-specific CD4+ helper T-cell control clone (p<0,001). In vitro co-incubation of the H-Y-specific CD4+ helper T-cell clone did not influence leukemia proliferation but induced up-regulation of MHC-class I and II, CD80, CD86 and CD40. In vitro, pre-incubation of leukemia cells with the H-Y-specific CD4+ helper T-cell clone irradiated did not improve the in vivo anti-leukemia efficacy of the H-Y-specific CD8+ CTL clone. Co-infusion of the H-Y specific CD4+ helper T-cell clone did not augment the in vivo persistence of the H-Y-specific CD8+ CTL T-cell clone. Nevertheless, the co-infusion resulted in a delay in leukemia progression of approximately 5 weeks, suggesting an additive, non overlapping anti-leukemia mechanism. Conclusions Minor Hag-specific human CD4+ T lymphocytes may contribute to the GvL effect through a direct, non cytotoxic mechanism, which could be additive to that of CD8+ CTL. The nature of this non cytotoxic GvL effect is currently under investigation. A.B. and L.H. equally contributed to this study. Disclosures: No relevant conflicts of interest to declare.

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