Immunotherapy with natural killer cells: a possible approach for the treatment of Acute Myeloid Leukemia also in Brazil / Imunoterapia com células natural killer: um caminho possível para o tratamento da Leucemia Mielóide Aguda também no Brasil

SUMMARY The allogeneic hematopoietic stem cell transplantation (HSCT) can cure intermediate and high-risk acute myeloid leukemia. Even with the development of strategies to reduce HSCT toxicity, this is still a complex treatment with high morbidity and mortality. Knowledge of the graft versus leukemia effect of HSCT has prepared the way for the development of Adoptive Immunotherapy or in vitro expansion of activated lymphocytes without alloreactivity, with subsequent intravenous infusion. The infusion of genetically modified T lymphocytes and haploidentical natural killer cells has been tested as an alternative to HSCT with very interesting results worldwide and in Brazil, as we not only have the technology of in vitro expansion of clinical grade lymphocytes available, but also do it according to the Good Manufacturing Practices that have been determined internationally.

RESUMO O transplante de células-tronco hematopoéticas (TCTH) alogênico é curativo para leucemia mielóide aguda de risco intermediário e alto. Mesmo com o desenvolvimento de estratégias para minorar a toxicidade do TCTH, este ainda é um tratamento complexo com elevada morbi-mortalidade. O conhecimento sobre o efeito enxerto contra leukemia do TCTH pavimentou o caminho para o desenvolvimento da Imunoterapia Adotiva ou expansão in vitro de linfócitos ativados, sem alo-reatividade, com posterior infusão endovenosa. A infusão de Linfócitos T geneticamente modificados e de células Natural Killer haploidenticas tem sido testada como alternativa ao TCTH com resultados bastante interessantes no mundo e no Brazil já que não apenas dominamos a tecnologia de expansão in vitro de linfócitos em grau clínico, como o fazemos segundo as Boas Práticas de Manufatura determinadas internacionalmente.

Effect of decitabine on immune regulation in patients with acute myeloid leukemia after allogeneic hematopoietic stem cell transplantation / 中国实验血液学杂志

Based on the representative articles in recent years, the different mechanisms of decitabine on immune regulation in patients with acute myeloid leukemia (AML) after allogeneic hematopoietic stem cell transplantation (HSCT) are summarized. Decitabine improves the expression of WT1 gene to stimulate specific cytotoxic T cells which can enhance graft versus leukemia effect (GVL) and improve the expression of FOXP3 gene to stimulate regulatory T cells so as to inhibit the acute graft versus host disease (GVHD). Through the above-mentimed mechanisms, decitabine can improve both therapeutic effect and quality of life in the patients with AML after allogeneic HSCT.

Implication of Early Lymphocyte Recovery after Allogeneic Hematopoietic Stem Cell Transplantation in Children with Leukemia

PURPOSE: The repopulating lymphocytes after allogeneic hematopoietic stem cell transplantation have an important role not only on the prevention of serious infections in the early transplantation period, but also on the killing of residual leukemic cells by graft-versus-leukemia effect. The aim of this study was to analyze the impact of lymphocyte recovery after allogeneic stem cell transplantation in children with hematologic malignancies. MATERIALS AND METHODS: We evaluated 69 children transplanted for acute lymphoblastic leukemia (ALL) (n=34), acute myeloid leukemia (AML) (n=26), chronic leukemia (n=7) and juvenile myelomonocytic leukemia (n=2) between 1996 and 2008 at the Chonnam National University Hospital, Korea. The patients were grouped based on absolute lymphocyte counts (ALC) or =500/microL at D+21 and D+30 after transplant. RESULTS: Patients with a High ALC at D+21 and D+30 had a faster neutrophil and platelet engraftment. The High at D+30 group had a better 5 year overall survival (71% vs. 53%, p=0.043) and event-free survival (72% vs. 53%, p=0.065) than the Low at D+30 group. The incidence of grade II-IV acute and chronic graft-versus-host disease (GVHD), and relapse rate did not differ by the ALC counts. However, the Low at D+30 group had a significantly increased risk for transplant-related mortality (p=0.019). The univariate analysis showed that the factors associated with decreased survival were a Low ALC at D+30, patients with high risk ALL, and grade II-IV aGVHD in patients with ALL and AML. CONCLUSION: Early posttransplant serial lymphocyte measurement would be a simple but useful method for predicting transplant outcomes.

Effects of immature dendritic cells genetically modified to express sTNFR I on graft-versus-host disease (GVHD) and graft-versus-leukemia (GVL) in allogeneic bone marrow transplantation mice / 中华血液学杂志

<p><b>OBJECTIVE</b>To investigate the effect of immature dendritic cells (inDC) genetically modified to express sTNFR I on acute graft-versus-host disease (aGVHD) and the graft-versus-leukemia (GVL) effect ofter allogeneic bone marrow transplantation (allo-BMT) in leukemic mice and its mechanism.</p><p><b>METHODS</b>An EL4 leukemia allo-BMT model was established with the BALB/c (H-2d) donor mice (DM)and C57BL/6 (H-2b) recipient mice (RM). The RM received DM bone marrow (BM) cells at a 1:1 ratio with spleen cells intravenously via tail vein at 4 h after TBI. Fifty DM were separated randomly into five groups: (1) Group A: total body irradiation (TBI) group, (2) Group B: lymphoma cell leukemia group, (3) Group C: allo-BMT group, (4) Group D: pXZ9-DC group, (5) Group E: sTNFR I-DC group. Acute GVHD scores, incidence of leukemic cell infiltration, histopathological analysis, survival rate, and survival rate of the recipients were estimated after allo-BMT. Enzyme-linked immunosorbent assay (ELISA) method was used to detect cytokines (INF-gamma and IL-4 ) production. Flow cytometry (FCM) analysis was used to detect allogeneic chimerism.</p><p><b>RESULTS</b>(1) The mice in group A and group B all died of the BM failure and lymphoma cell leukemia, respectively. The mice in group C developed typical clinical signs of a GVHD after BMT with an average survival time(AST) of (11.50 +/- 3.50) d. The signs of aGVHD were less evident in the group D and E, and their AST (21.70 +/- 5.80 and 25.80 +/- 5.20 days, respectively) were all longer than that in group C (P < 0.05). AST of group E was the longest (P < 0.05). The mice in group B all died of leukemia within 18 days after engraftment of EL4 cells. There was was no significant difference in groups C, D and E in the incidence of leukemia (P > 0.05). (2) Serum IFN-gamma level reached peak value. At + 12 d, then decreased gradually in group C, D, and E, and then reached the nadir at +18 d post-BMT, with the lowest in group E (P < 0.05), and the level was significantly lower in group D than in group C (P < 0.05). After BMT, serum IL-4 level slightly decreased in group C, but gradually elevated in group D and E and reached their peak at +12 d, and even more significantly increased in group E (P < 0.05). There was no statistical significance in the pair wise comparison among three group (P < 0.05). (3) The average proportion of H-2d positive cells in RM was 95%-100% on day 30 post-BMT, with complete donor-type implantation.</p><p><b>CONCLUSION</b>Immature DC can induce immuno tolerance. Immature DC genetically modified to express sTNFR I has been shown to prevent acute GVHD in lethally irradiated mice reconstituted with allogeneic bone marrow grafts while maintaining the GVL response.</p>

Relationship between WT1-specific T-cell subsets and graft-versus-host disease after nonmyeloablative allogeneic transplantation / 中华血液学杂志

<p><b>OBJECTIVE</b>To explore the relationship between WT1-induced T-cell subsets and graft-versus-host disease (GVHD) after nonmyeloablative allogeneic hematopoietic stem cell transplantation (NST).</p><p><b>METHODS</b>Peripheral blood mononucleated cells (PBMCs) from 19 patients who expressed WT1 and developed GVHD after NST were simulated by WT1126-134 peptide in vitro, and proportions of WT1-induced-T-cell subsets (Tc1, Tc2, Th1, Th2 cells) before and after transplant were detected by intracellular cytokine staining (ICCS) assay. WT1-specific CD8(+) CTLs of 14 patients with HLA-A*0201 were detected by HLA-A*0201/WT1 pentamer.</p><p><b>RESULTS</b>(1) 17 of 19 patients developed GVHD, among whom proportions of Tc1 and Th1 cells, achieved peak value in 16 patients at occurrence of GVHD (P = 0.039); (2) The peak proportions of Tc1 and Th1 cells in patients with aGVHD above grade II were higher than those with grade I, but being no statistical difference (P = 0.900 and P = 0.140, respectively); (3) The peak proportion of Th1 cells (P = 0.004), but not Tc1 cells (P = 0.060) in patients with extensive cGVHD was significantly higher than that in patients with limited one; (4) Proportions of Tc1, Th1 and WT1(+)CD8(+)CTL in patients without GVHD were similar to those in patients with Grade I aGVHD, but lower than those in aGVHD above grade II.</p><p><b>CONCLUSION</b>GVHD promotes the generation of WT1-induced GVL effect, and the intensity of the latter maybe correlated with the intensity of GVHD, especially cGVHD. Th1 cells play a more important role in the enhancement of WT1-induced GVL effect in extensive cGVHD patient than in limited cGVHD patients.</p>

Hematopoietic Stem Cell Transplantation in Children with Leukemia: A Single Institution Experience with Respect to Donors

Aim of this study was to compare the outcomes of transplantation by donor source and to help select the best alternative donor in children with leukemia. Donor sources included matched related donor (MRD, n = 35), allele-matched unrelated donor (M-UD, n = 10) or -mismatched (MM)-UD (n = 13) or unrelated umbilical cord blood (UCB, n = 11). UCB group had a significantly higher incidence of grade II-IV acute graft versus host disease (MRD, 11.8%; M-UD, 30.0%; MM-UD, 15.4%, UCB, 54.4%, P = 0.004) but there was no difference in incidence of chronic graft versus host disease between 4 groups. The 5-yr leukemia-free survival (LFS) was 76.7%, 60.0%, 69.2%, and 45.5%, respectively (P = 0.128). MRD group showed higher LFS rate than UCB group (P = 0.022). However, LFS of M-UD and MM-UD together (65.2%) was not different from that of MRD group (76.7%, P = 0.325), or from that of UCB (45.5%, P = 0.190). The relapse incidence at 5 yr was 17.1%, 20.0%, 15.4%, and 0%, respectively (P = 0.460). The 100-day treatment-related mortality was 2.9%, 20.0%, 7.7%, and 36.4%, respectively (P = 0.011). Despite the limitations of small number of patients, unrelated donor transplants including even allele-mismatched ones, seem to be as effective in children with leukemia lacking suitable relative donors. Also, UCB transplant may serve as another possible option in urgent transplants.

The immunobiology of cord blood transplantation / 대한혈액학회지

Despite significant recent advances in the applicability and outcome following unrelated cord blood transplantation (UCBT), infections remain a major cause of mortality associated with poor immune recovery in the first 6 months after UCBT. Enhanced immune reconstitution not only could improve survival by reduced transplant related mortality, but may also favorably impact on relapse incidence by improved graft-versus-leukemia effects. This review will summarize our current understanding of the biology of immune recovery post-UCBT with an emphasis on adaptive T cell dependent immunity. New efforts to boost immunity will be also highlighted including our own laboratory, where ex vivo T cell expansion is pursued towards adoptive immunotherapy.

Influence of donor Treg cells on GVHD and GVL effects after allogeneic bone marrow transplantation in mice / 中国实验血液学杂志

In order to explore the influence of purified donor regulatory T cells (Treg) infused after allogeneic bone marrow transplantation (allo-BMT) on GVHD and GVL effect in mice, an EL4 leukemia allo-BMT model of BALB/c-->C57BL/6 mice was established. The CD4(+)CD25(+)T cells were purified by positive selection using MACS. The recipients were injected with CD4(+)CD25(+)T cells or CD4(+)CD25(-)T cells within 4 hours respectively along with allo-BMT. Survival time, clinical GVHD score or histopathological features were observed after allo-BMT. The results showed that the mean survival time in leukemia control group was (17.9 +/- 0.7) days and all mice died of leukemia. The mean survival times in transplantation control group and CD4(+)CD25(-)T group were (23.2 +/- 1.6) and (22.3 +/- 1.9) days. Histopathological analysis in several target organs (skin, liver and small intestine) confirmed the presence of severe GVHD. The mean survival time in Treg group was (47.3 +/- 6.5) days. 70% of recipients were alive until day 60 without features of GVHD or tumor progression. Clinical GVHD scores in Treg group significantly decreased as compared to transplantation control group and CD4(+)CD25(-)T group (p < 0.05). It is concluded that allo-BMT combined with injection of donor CD4(+)CD25(+)Treg cells can efficiently prevent recipients from lethal GVHD with preserving GVL effect during allo-BMT.

Clinical study on the relapsd leukemia patients with graft versus host disease after allogeneic hematopoietic stem cell transplantation / 中华血液学杂志

<p><b>OBJECTIVE</b>To explore the dissociation of graft-versus-leukemia (GVL) effects from graft-versus-host disease (GVHD) in the patients who experienced GVHD during leukemia relapse after allogeneic hematopoietic stem cell transplantation (allo-HSCT).</p><p><b>METHODS</b>The primary disease, disease status, GVHD, response to donor lymphocyte infusion (DLI) and prognosis were analysed in 11 leukemia patients who relapsed with GVHD after allo-HSCT.</p><p><b>RESULTS</b>Of the 11 relapsed, 5 were acute lymphoblastic leukemia and 6 acute myeloid leukemia. Five received DLI before relapse and all developed post-DLI GVHD, including 2 grade II acute GVHD (aGVHD), 1 limited chronic GVHD (cGVHD) plus grade II aGVHD, and 2 extensive cGVHD. After relapse of the 5 patients, 2 received Chemo-DLI, one achieved CR with extensive cGVHD and then relapsed again, the other didn't achieved CR. The other 6 patients didn't received DLI before relapse and also developed post-HSCT GVHD while relapsing, including 3 extensive cGVHD, 1 grade I aGVHD and 2 grade II-IV aGVHD. After relapse, these 6 patients received Chemo-DLI, 2 achieved CR and then relapsed again, 4 didn't achieved CR.</p><p><b>CONCLUSION</b>The elicited GVHD after allo-HSCT may not accompany effective GVL effects inhibiting leukemic relapse.</p>

Papel do Interferon-γ na resposta anti-leucemia associada à Doença Enxerto contra Hospedeiro Experimental

A utilização do Transplante alogeneico de Células Tronco Hematopoiéticas para o tratamento de malignidades é em grande parte prejudicada pela ocorrência da Doença Enxerto contra Hospedeiro (DECH). A grande maioria dos esforços para reduzir a incidência da DECH resultou na diminuição do efeito Enxerto contra Leucemia (ECL), com aumento na recaída pós-transplante, sugerindo que os efeitos enxerto contra Hospedeiro (ECH) e ECL estão intimamente ligados. Para separar esses efeitos, é preciso caracterizar os mecanismos imunológicos envolvidos com cada um deles. Neste trabalho, foi caracterizado um sistema capaz de discriminar e quantificar os efeitos ECH e ECL: hospedeiros BALB/c ou F1(B6XDBA/2) irradiados letalmente receberam medula óssea (MO) de C57BL/6 administrada junto com células T esplênicas de camundongos C57BL/6 normais ou deficientes em Interferon-γ (GKO), acompanhadas de células do mastocitoma P815 expressando GFP. O efeito ECH foi avaliado pela mortalidade, por parâmetros clínicos (peso, pele, diarréia, postura e atividade) e por histopatologia (pele, fígado e intestino), e o efeito ECL foi avaliado pela quantidade de células leucêmicas no sangue, baço, fígado, linfonodos e medula óssea, utilizando-se citometria de fluxo. Foi verificado que o efeito ECL em órgãos linfóides secundários (OLS) era independente da produção de IFN-γ. No fígado, o efeito ECL foi dependente de IFN-γ apenas quando o efeito ECH era parcial. Na MO, o ECL foi estritamente dependente de IFN-γ, independente da magnitude do ECH. A avaliação da produção de citocinas nos OLS mostrou que as células T aloreativas produziam IL-17A apenas nos receptores de células leucêmicas, independente de IFN-γ. Em contraste com receptores normais, em receptores leucêmicos de células T GKO a patologia intestinal é moderada, e os danos no fígado ocorrem principalmente no parênquima, com preservação dos espaços vasculares. O efeito ECH observado na ausência de IFN-γ correlaciona-se com o efeito ECL em todos os órgãos avaliados exceto na MO. Esses resultados levantam importantes questões relacionadas ao controle da doença residual mínima e à resposta imune na MO, um santuário de micrometástases. IL-17A, presente apenas em receptores de células leucêmicas e independente de IFN-γ, não foi suficiente para eliminar as células malignas da MO. Nossos resultados sugerem que a resposta anti-tumoral é sítioespecífica, provavelmente refletindo diferentes condições ambientais impostas por diferentes órgãos. Nós acreditamos que o estudo dos mecanismos indutores e efetores dos efeitos ECH e ECL, nos diferentes órgãos alvo, nos possibilitará desenvolver novos tratamentos que inibam a DECH enquanto mantenham o efeito ECL.

The clinical use of allogeneic stem cell transplantation for cancer treatment is seriously hampered by the occurrence of graft-versus-host disease (GVHD). For the most part, efforts to reduce the incidence of GVHD have also diminished graft-versusleukemia (GVL) responses, with increased tumor relapse, suggesting that the Graftversus- Host (GVH) and the GVL effects are intimately linked. To separate these effects, the immunological mechanisms related to each one must be characterized. In this work, a system able to discriminate and quantify the GVH and GVL effects was characterized: Radiation bone marrow (BM) chimeras were prepared using BM cells from C57BL/6 administered into BALB/c or F1(B6XDBA/2) hosts, together with splenic T cells from C57BL/6 or Interferon-γ (IFN- γ) knockout mice. These chimeras were injected IV with P815 mastocytoma cells transduced with GFP. The GVH effect was evaluated by the mortality rate, clinical parameters (weight, skin, diarrhea, hunching and activity) and histopathology (skin, liver and intestine), and the GVL effect was evaluated by the amount of leukemia cells in the blood, spleen, liver, lymph nodes and bone marrow, using flow cytometry. It was verified that the GVL effect was present in secondary lymphoid organs (SLOs) regardless IFN-γ production. In the liver, the GVL was dependent on IFN-γ only when the GVH effect was partial. In the BM, the GVL was strictly dependent on IFN- γ, and independent of the GVH magnitude. Evaluation of the cytokine production in SLOs showed that alloreactive T cells produced IL-17A only in leukemic recipients, independently of IFN- γ. By contrast with normal recipients, in leukemic recipients of GKO T cells the intestinal pathology was mild and the liver damage was mainly in the parenchyma, preserving the vascular spaces. The GVH effect observed in the absence of IFN-γ correlates with the GVL effect in every organ except in the BM. These results raise important questions related to the control of minimal residual disease and the immune response in the BM, a sanctuary of micrometastasis. IL17A, present only in leukemic recipients and independent of IFN- γ, was not enough to eliminate malignant cells from the BM. Our results suggest that the anti-tumor response is site specific, probably reflecting different environmental conditions imposed by different organs. We believe that studying the inductive and effector mechanisms of GVH and GVL effects in different target organs will enable us to design new treatments to prevent GVHD while maintaining the GVL effect.

The action of donor-derived NK cell in leukemic mice MHC haplotype-mismatched bone marrow transplantation / 中华血液学杂志

<p><b>OBJECTIVE</b>To study the role of donor-derived NK cells in haploidentical bone marrow transplantation (BMT) in leukemic mice.</p><p><b>METHODS</b>CB6F1(H-2b/d) mice model of EL9611 (H-2d) erythroleukemia was established by injection of EL9611 (H-2d) cells via tail vein. CB6F1(H-2b/d) mice were used as recipient, and C57BL/6(H-2b) mice as donor. Five days later, 70 CB6F1(H-2b/d) mice were randomly divided into 7 groups (10 mice per group) as follows: group 1: without treatment; group 2: simple-irradiated group; group 3: treated with cytarabine (Ara-C) at 50 mg/kg x6 d; group 4: simple BMT; group 5: haploidentical BMT with graft-versus-host disease (GVHD) that injected with bone marrow cells and spleen cells of C57BL/6(H-2b) mice 4 hours after irradiation; group 6: after irradiated with 9 Gy, mice were injected with C57BL/6(H-2b) NK cells (1 x 10(6)) and 4 hours later with BM cells, group 7: after irradiation of 9 Gy, mice were injected with C57BL/6(H-2b) NK cells (1 x 10(6)) and 4 hours later with BM cells and spleen cells. The blood routine test, survival time, body weight, and histopathology in the recipients were observed and compared among these group.</p><p><b>RESULTS</b>The survival time was (10.1 +/- 0.9), (9.8 +/- 0.9), (22.7 +/- 3.2) and (20.1 +/- 1.7) days in groups 1, 2, 3, and 5 respectively; was (30.1 +/- 16.0) days in group 4, out of which 2 mice survived for more than 30 days. The survival time was (39.1 +/- 18.1) and (49.3 +/- 17.2) days in groups 6 and 7 respectively, out of which 4 mice in group 6 and 7 mice in group 7 survived for more than 30 days. The survival time in group 6 was much longer than that in group 1, 2, 3 and 5 (P < 0.01). The survival time in group 7 was much longer than that in other groups (P < 0.05). The liver and spleen enlargement, organ destruction and infiltration with leukemic cells were observed in mice died from leukemia. The chimerism of Y chromosome appeared (80%-90%) in long-term survival mice in groups 6 and 7.</p><p><b>CONCLUSION</b>Donor-derived NK cells have the antileukemia ability and reduce GVHD in haploidentical BMT in erythroleukemia mice (EL9611, H-2d).</p>

Factors influencing graft versus leukemia effect -- review / 中国实验血液学杂志

In treating of leukemia and controlling of minimal-residual disease (MRD), graft versus leukemia effect (GVL) plays a critical role, and complicated mechanisms are involved in this immunology process. When graft cells are infused into recipients, the evoked GVL effect must be inevitably influenced by many factors derived from allogeneic effect between donor and receptor. To utilize GVL more efficiently in future clinical practice and to improve the curative effect of allo-HSCT, it is necessary to recognize these factors. Some potential factors influencing GVL such as chimerism patterns, autocytotoxic cells, dynamics of immune cells in patients, the cytokines and so on are reviewed in this article.

Graft-versus-Leukemia Effect of Nonmyeloablative Stem Cell Transplantation

Nonmyeloablative stem cell transplantation (NST) is increasingly used with beneficial effects because it can be applied to older patients with hematological malignancies and those with various complications who are not suitable for conventional myeloablative stem cell transplantation (CST). Various conditioning regimens differ in their myeloablative and immunosuppressive intensity. Regardless of the type of conditioning regimen, graft-versus- host disease (GVHD) in NST occurs almost equally in CST, although a slightly delayed development of acute GVHD is observed in NST. Although graft-versus-hematological malignancy effects (i.e., graft-versus-leukemia effect, graft-versus-lymphoma effect, and graft-versus-myeloma effect) also occur in NST, completely eradicating residual malignant cells through allogeneic immune responses is insufficient in cases with rapidly growing disease or uncontrolled progressive disease. Donor lymphocyte infusion (DLI) is sometimes combined to support engraftment and to augment the graft-versus-hematological malignancy effect, such as the graft-versus-leukemia effect. DLI is especially effective for controlling relapse in the chronic phase of chronic myelogenous leukemia, but not so effective against other diseases. Indeed, NST is a beneficial procedure for expanding the opportunity of allogeneic hematopoietic stem cell transplantation to many patients with hematological malignancies. However, a more sophisticated improvement in separating graft-versus-hematological malignancy effects from GVHD is required in the future.

Effect of recipient mouse splenocytes taken orally by donor mouse on GVHD after splenocyte transplantation / 中国实验血液学杂志

This study was aimed to explore whether the GVHD in mice can be ameliorated and the GVL effect in mice can be reserved by transfusion of lymphocytes of donors fed with recipient splenocytes effect. Male (DBA-2) mice (H-2(d)) as donors were fed with BALB/c splenocytes, DBA-2 splenocytes, bovine serum albumin, or regular chow, every other day. Induction of tolerance was assessed by a mixed lymphocyte reaction (MLR). Female (BALB/c) mice (H-2(d)) as recipients received total body irradiation (TBI) of 6.0 Gy ((60)Cogamma-ray) followed by inoculation of 3 x 10(3) P388 mouse leukemia cells on the same day. Subsequently, tail vein injection of 2 x 10(7) splenocytes supplied by DBA-2 was undertaken. Control groups were fed identically without leukemia cell inoculation. The results showed that GVHD was significantly ameliorated and CD4(+)/CD8(+) ratio increased in recipient-mice transplanted with splenocytes of tolerated donors, compared with control animals. There was no significant difference in survival rate between different groups of recipients inoculated with leukemia cell. It is concluded that the peroral recipient-mouse splenocytes can ameliorate GVHD without hampering effect on GVL.

Experimental study on IL-2- and IL-15 application in allogeneic hematopoietic stem cell transplantation / 中华血液学杂志

<p><b>OBJECTIVE</b>To explore the impact of IL-2- and IL-15-activated donor natural killer (NK) cell infusion on graft-versus-host-disease (GVHD) and graft-versus-leukemia (GVL) effect post allogeneic hematopoietic stem cell transplantation (allo-HSCT).</p><p><b>METHODS</b>The C57BL/6 mice splenic NK cells were selected by microbeads, and then expanded in the media containing IL-2 and IL-15. The killing activity of NK cells was detected. In the leukemia mouse model, recipients (BALB/c) were intravenously inoculated with EL9611 leukemia cells 8 days before transplantation. Lethally irradiated BALB/c recipient mice were transplanted with 5 x 10(6) bone marrow cells (BMCs), or 5 x 10(6) BMCs plus 1 x 10(7) splenocytes with or without 1 x 10(7) activated NK cells. Additionally, NK cell infusion group mice were intraperitoneally injected with a mixture of IL-2 and IL-15 post transplant. Survival time, GVHD occurrence, lineage chimerism, TRBV spectra-typing were observed post transplant.</p><p><b>RESULTS</b>The purity of isolated splenic NK cells was 95.7% - 97.1%. The killing activity of NK cells after activation was increased by 3 times. GVHD did not occurred in allogeneic BMCs infusion group, whereas did from 1 week after transplant in allogeneic BMCs + splenocytes infusion group. The severity of GVHD in total body irradiation (TBI) experimental group was significantly lower than in splenocytes infusion group (P < 0.05). The survival time was 9.5 - 14.0 d in TBI alone conditioning group. In leukemia mouse model, 100 day survival rate was 10% the rest of them were died of leukemia while in experimental group, the more than 100 days survival rate was 80% (P < 0.01). PB NK cells at 2 week post-transplant were 4.8% in experimental group and 2.8% in control group. NK cells recovery in experimental group was earlier than that in control group (P < 0.05). TRBV reconstitution was faster in experimental group than in control group, moreover, the number of TRBV family expression was more in experimental group than in control group which mainly expressed monoclone or oligo-clone.</p><p><b>CONCLUSIONS</b>Donor alloreactive NK cells can be efficiently expanded and activated with IL-2 and IL-15. Donor activated NK cell infusion and IL-2, IL-15 treatment can promote immune reconstitution, mitigate GVHD and reduce leukemia relapse.</p>

Effects of NKG2D and its ligands RAE-1 and H60 on graft-versus-tumor response / 中国实验血液学杂志

The study was purposed to explore the effects of NKG2D receptor and its ligands RAE-1 and H60 on graft-versus-tumor (GVT) response induced by MHC haploidentical bone marrow/spleen cell transplantation. Female (BALB/c x C57BL/6) F1 mice (CB6F1, H-2K(b/d)) inoculated with H22 cells to develop a solid tumor model were the recipients, and bone marrow mixed with spleen cells of the healthy male C57BL/6 (H-2K(b)) mice were the donor cells. GVT response was observed after transplantation that from donor cells T and NK cells were purged with anti-CD3 and anti-NK monoclonal antibody, and the NKG2D receptor was blocked with anti-NKG2D monoclonal antibody, the expression levels of RAE-1 and H60 mRNA in tumor tissue were measured by means of semi-quantitative reverse transcription polymerase chain reaction (RT-PCR) at different time points after transplantation. The results showed that the GVT response of transplantation was reduced after in vitro depletion of T and NK cells or blocking NKG2D receptor in donor cells of the graft, the expression levels of RAE-1 and H60 mRNA in tumor tissue increased after transplantation of haploidential bone marrow mixed with spleen cells. It is concluded that NKG2D and its ligands RAE-1 and H60 may play important roles in GVT response.

Investigation of plasmacytoid dendritic cell reconstitution and its relationship with cGVHD and relapse after haploidentical hematopoietic stem cell transplantation / 中国实验血液学杂志

To investigate the characteristics and significance of reconstitution of peripheral blood plasmacytoid dendritic cell (PDC) precursors after allogeneic human leukocyte antigen mismatched/haploidentical hematopoietic stem cell transplantation, and its relationship with chronic graft versus host disease (cGVHD) and relapse, 19 patients with leukemia were enrolled for this study. Peripheral blood dendritic cell (DC) subsets of patients and healthy controls were detected by flow cytometry, and the correlations between reconstitution of DC and cGVHD, relapse were analyzed. The results showed that compared with healthy subjects, patients with leukemia had a significantly decreased proportion and absolute number of myeloid dendritic cell (MDC), MDC1, DC and the ratio of MDC/PDC (P<0.05). There were not statistically different in MDC2 and PDC between patients and healthy subjects. After transplant, all the proportion of WBC and absolute numbers of DC reached to healthy controls levels at 9 months (P>0.05), besides the proportion of PDC which reached to healthy controls levels at 1 year (P=0.494). Compared with levels before relapse, the proportions of MDC1, MDC, DC and the ratio of MDC/PDC were lower, but proportions of MDC2 and PDC were slightly higher after relapse. Patients with a 'high' PDC recovery profile had an improved cumulative incidence of cGVHD in contrast to patients with a 'low' PDC recovery profile on day 120 after transplantation (P=0.007). It is concluded that compared with healthy subjects, de novo leukemia patients have a significantly decreased proportion and absolute number of DC and the ratio of MDC/PDC before haploidentical hematopoietic stem cell transplantation; while ratio of MDC/PDC can be normalized with relative rapidity, the proportions of all DC subsets reached to normal levels on the whole at 9 months after transplantation, and also recovery level of DCs is correlated with occurrence of cGVHD and relapse.

Hepatocyte growth factor and its immunoregulatory activity - review / 中国实验血液学杂志

Hepatocyte growth factor (HGF) is a pleiotropic cytokine, its roles in the physiology and pathology of immune system, have been investigated thoroughly, great deal of data have been documented on its immunoregulatory activity. In this review, according to advance of study on HGF in recent years, the role of HGF in the immune regulation, such as immunoregulatory effects of HGF on T lymphocytes, B lymphocytes and dendritic cell, modulation of HGF on specific humoral and cellular immune response, control of acute GVHD and acceleration of myeloid and immunologic reconstitution in allogenetic bone marrow transplantation models, promotion of tissue repair and regeneration, and alleviation of immune rejection in allogeneic organ transplantation including the heart, liver and kidney transplantation, prevention of grafts from injury as well as applicably useful of HGF in the therapy of autoimmune disorders were summarized.