- Email: [email protected]
C029 HLA-identical umbilical cord blood transplantation from a sibling donor in juvenile myelomonocytic leukemia
S48
Oral Communications
respectively) suggesting a non-toxic effect of this concentration. However, in the presence of higher doses (1.0mg/mL30mg/mL) the proportion of apoptotic BMMCs increased at a dose-dependent manner in both patient and control groups although not at a statistically significant level (p = 0.4018 and p = 0.1377, respectively). A pooled analysis of patient and control data showed a statistically significant increase in the proportion of apoptotic BMMCs in the presence of SB-497115 at a dose 1mg/mL (1.0mg/mL-30mg/mL), compared to baseline (p = 0.0402, one-way ANOVA) These preliminary data suggest that SB-497115 at a dose of 0.1mg/mL displays a beneficial effect on megakaryopoiesis of MDS patients with low/intermediate-I IPSS without any adverse effect on BMMC survival. These encouraging in vitro data indicate a possible clinical beneficial effect of Eltrombopag in this group of patients. Studies investigating the in vitro effect of lower SB-497115 concentrations are in progress.
C028 Lenalidomide abrogates the clonal advantage of del(5q) MDS stem cells via alteration of niche interactions C. Scharenberg ° , M. Jadersten, J. Palmquist, L. Forsblom, M. Jansson, E. Hellstr¨om-Lindberg. Karolinska Institute, Stockholm, Sweden *E-mail: [email protected] Background: The myelodysplastic syndromes (MDS) constitute a heterogeneous group of clonal disorders of hematopoietic stem cells (HSC) leading to ineffective hematopoiesis in one or more lineages in the bone marrow. A range of cytogenetic abnormalities can be identified in around 50% of cases, with the most frequent cytogenetic entity being the 5q−syndrome, characterized by loss of the 5q31−33 region. Recently, the thalidomide analogue lenalidomide has been demonstrated to induce cytogenetic response and restored erythropoiesis in this group of patients. While inhibition of angiogenesis, cell adhesion, and proliferation, as well as modulation of pro-apoptotic cytokines, have all been put forward as putative modes of action of lenalidomide, the exact mechanism has remained unclear. Interestingly, the more primitive hematopoietic compartment seems to possess a clonal advantage where del(5q) HSC are able to outcompete non-5q HSC. Whether the molecular mechanism for clonal advantage is intrinsic to the stem cell clone or a result of a compromised niche is unclear. However, we have previously demonstrated that lenalidomide is able to abrogate this clonal advantage. Aims: Our previous results implicated the matri-cellular protein SPARC to be upregulated in CD34+ del(5q) cells in response to lenalidomide. Thus, we sought to test the hypothesis that a putative decrease in SPARC expression of a clonal hematopoietic progenitor should give rise to a population of cellular descendants that exhibit an increased
adhesion to the microenvironment and more active rates of proliferation, thus explaining the clonal advantage of del(5q) MDS stem cells. Methods: We therefore studied the basal adhesion of del(5q) CD34+ progenitor cells to the murine stromal cell line MS-5 using a gravitational-force upon-inversion assay. Using multi-parameter flow cytometry, we studied proliferation and apoptosis in del(5q) and normal HSC and how these parameters are affected by lenalidomide and recombinant SPARC-protein. Results: Our data show that del(5q) CD34+ progenitors exhibited increased adhesion to the murine stromal cell line MS-5 compared to normal CD34+ cells. In short-term in vitro cultures, lenalidomide and recombinant SPARC were able to abrogate this increased adhesion. Multiparameter flow cytometry revealed a decreased rate of spontaneous apoptosis in del(5q) HSC compared to progenitors. Lenalidomide increased apoptosis in both compartments and this effect was specific to del(5q) cells. However, neither lenalidomide nor SPARC appeared to have any effect on the proliferation of CD34+ progenitors from normal or del(5q) bone marrow. Summary/Conclusion: These studies suggest that decreased expression of SPARC leads to increased adhesion of del(5q) HSC/progenitor cells to their microenvironment and may explain why del(5q) HSC are able to outcompete the remaining healthy HSC. Our studies suggest that lenalidomide is able to abrogate this clonal advantage partly via its increase in SPARC expression with a consecutive decrease in adhesion and increase in apoptosis.
4. Therapy and Experimental Models C029 HLA-identical umbilical cord blood transplantation from a sibling donor in juvenile myelomonocytic leukemia A. De Vries1 ° , R. Bredius2 , A. Lankester2 , M. Bierings3 , M. Trebo4 , P. Sedlacek5 , C. Niemeyer6 , M. Zecca7 , F. Locatelli7 , M. van den Heuvel-Eibrink1 . 1 Department of Pediatric Hematology/Oncology, ErasmusMC Sophia Childrens Hospital, Rotterdam, The Netherlands; 2 Department of Pediatrics, Leiden University Medical Center, Leiden, The Netherlands; 3 Department of Pediatric Hematology/Oncology, UMCU/Wilhelmina Childrens Hospital, Utrecht, The Netherlands; 4 Department of Pediatrics, St Anna Childrens Hospital, Vienna, Austria; 5 Department of Pediatric Hematology/Oncology, Charles University, University Hospital Motol, Prague, Czech Republic; 6 Department of Pediatrics, University of Freiburg, Freiburg, Germany; 7 IRCCS Policlinico San Matteo, Pediatric Hematology/Oncology, Pavia, Italy *E-mail: [email protected] Background: Currently, for juvenile myelomonocytic leukemia (JMML) stem cell transplantation is the only
4. Therapy and Experimental Models curative treatment option. Although successful unrelated donor umbilical cord blood transplantations (UCBTs) have been reported, series of HLA-identical sibling donor UCBTs in JMML are not available. Patients and Methods: From the European Working Group on Childhood MDS (EWOG MDS) registry 5 JMML patients who underwent a fully HLA matched sibling UCBT were identified. Results: The median age at diagnosis was 18 months (range 15−30 months). None of the patients had clinical signs of NF1. In one case a PTPN11 mutation was found. In two other cases no mutation in RAS or PTPN11 was found. In one case no mutation analysis was performed, in four cases the conditioning regimen consisted of busulfan, cyclophosphamide and melphalan and in one case of cyclophosphamide, etoposide and total body irradiation. All patients engrafted slowly (ANC > 500 ul: median 33 days, range 10−35 days, platelets >20,000/ul: median 53 days, range 38−77 days). In 3 patients acute graft versus host disease was noticed (grade 1 and 2); no chronic graft versus host disease was reported. Two patients relapsed after the initial transplantation and underwent a second transplantation with marrow of the initial donor. One of them is in second complete remission and the other died after a second relapse. One patient developed increased donor chimerism from day 42 without any clinical sign of relapse. She was treated with DLI, 6-mercaptopurine and 13-cis retinoic acid respectively. She is still in complete remission 5 years after transplantation. Conclusion: This EWOG series illustrates that, although this needs to be confirmed in larger series, transplantation with relatively immunologically naive cord blood stem cells from a HLA-matched sibling is feasible in selected cases of JMML.
C030 Long-term remission of post-transplant MDS/AML by adoptive transfer of allogeneic WT1-specific CD4+ and CD8+ T lymphocytes Y. Kim, N. Chung, S. Cho, M. Kim, E. Kim, H. Son, H. Choi, H. Kim, C. Min, S. Lee, D. Kim, W. Min, C. Kim, T. Kim ° . Catholic University of Korea, Seoul, South Korea *E-mail: [email protected] We treated a patient with post-transplant MDS/AML by adoptive transfer of Wilm’s tumor 1 (WT1)-specific cytotoxic T lymphocytes (WT1-CTL). WT1-CTL were generated by stimulating donor T cells with dendritic cells transduced with recombinant adenovirus encoding truncated WT1 (DWT1) created by deletion of the N-terminal portion of full length WT1. DWT1-CTL was infused at a cumulative dose of 8x107 cells/m2 after fludarabine-based chemotherapy and DLI/stem cell support. Although GVHD developed on skin, mouth, and eye 2 months after CTL
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therapy, steroid therapy was effective. The patient remained in sustained remission for 37 months after DWT1-CTL therapy until disease relapsed again. The frequencies of DWT1-specific T cells gradually increased over one year and then decreased in peripheral blood. IFN-g ELISPOT responses specific for DWT1 were detected in both CD4+ and CD8+ T cells at 2 years. Furthermore, IFN-g secreting CD8+ T cells specific for peptides located within DWT1 were detected with higher frequencies compared to those for peptide within the deleted N-terminal portion of WT1. These results show that in vitro cultured DWT1-specific CD4+ and CD8+ cells can be expanded and function to overcome immune tolerance in a leukemic patient, eliciting long-lasting anti-leukemic immunity. These results raise the possibility that adoptive transfer of in vitro cultured leukemia-specific CTL can be successfully applied to treat hematological malignancies in an allogeneic setting.
C031 Maintenance treatment with 5-azacitidine for patients with high risk myelodysplastic syndrome (MDS) or acute myeloid leukemia following MDS (MDS-AML) in complete remission (CR) after induction chemotherapy M. Groevdal1 , R. Kahn1 , M. Jansson1 ° , A. Aggerholm2 , P. Antunovic3 , J. Astermark4 , P. Bernell5 , L. Engstroem6 , L. Kjeldsen7 , O. Linder8 , L. Nilsson9 , A. Olsson10 , M. Skovholm11 , J. Tangen12 . 1 Karolinska Institutet, Stockholm, Sweden; 2 Aarhus University hospital, Denmark; 3 Linkoping University hospital, Sweden; 4 Malmoe University hospital, Sweden; 5 Karolinska University hospital, Sweden; 6 University hospital of Norrland, Sweden; 7 Rigshospitalet, Denmark; 8 Orebro University hospital, Sweden; 9 Lund University hospital, Sweden; 10 Sahlgrenska University hospital, Sweden; 11 Aalborg hospital, Denmark; 12 Ullevaal University hospital, Norway *E-mail: [email protected] Around 50% of patients with high-risk MDS or MDSAML may enter CR after induction chemotherapy, but CR duration, as well as overall survival is usually short. To address this clinical problem the Nordic MDS Group designed a prospective multicenter phase II study, which assessed the clinical feasibility and utility of long-term maintenance treatment with azacitidine. Sixty patients with high-risk MDS (IPSS intermediate-2 or high) (n = 23) or AML following a previous known MDS (n = 37) were enrolled between 2004 and 2006. The mean age was 68 (54−83) and patients should not be eligible for stem cell transplantation. Induction treatment consisted of standard doses of daunorubicin and ara-C. Patients in CR received low dose azacitidine subcutaneously 5/28 days until relapse, unless unacceptable toxicity developed. Methylation status of the P15ink4b (P15), E-cadherine (CDH ) and
Oral Communications
respectively) suggesting a non-toxic effect of this concentration. However, in the presence of higher doses (1.0mg/mL30mg/mL) the proportion of apoptotic BMMCs increased at a dose-dependent manner in both patient and control groups although not at a statistically significant level (p = 0.4018 and p = 0.1377, respectively). A pooled analysis of patient and control data showed a statistically significant increase in the proportion of apoptotic BMMCs in the presence of SB-497115 at a dose 1mg/mL (1.0mg/mL-30mg/mL), compared to baseline (p = 0.0402, one-way ANOVA) These preliminary data suggest that SB-497115 at a dose of 0.1mg/mL displays a beneficial effect on megakaryopoiesis of MDS patients with low/intermediate-I IPSS without any adverse effect on BMMC survival. These encouraging in vitro data indicate a possible clinical beneficial effect of Eltrombopag in this group of patients. Studies investigating the in vitro effect of lower SB-497115 concentrations are in progress.
C028 Lenalidomide abrogates the clonal advantage of del(5q) MDS stem cells via alteration of niche interactions C. Scharenberg ° , M. Jadersten, J. Palmquist, L. Forsblom, M. Jansson, E. Hellstr¨om-Lindberg. Karolinska Institute, Stockholm, Sweden *E-mail: [email protected] Background: The myelodysplastic syndromes (MDS) constitute a heterogeneous group of clonal disorders of hematopoietic stem cells (HSC) leading to ineffective hematopoiesis in one or more lineages in the bone marrow. A range of cytogenetic abnormalities can be identified in around 50% of cases, with the most frequent cytogenetic entity being the 5q−syndrome, characterized by loss of the 5q31−33 region. Recently, the thalidomide analogue lenalidomide has been demonstrated to induce cytogenetic response and restored erythropoiesis in this group of patients. While inhibition of angiogenesis, cell adhesion, and proliferation, as well as modulation of pro-apoptotic cytokines, have all been put forward as putative modes of action of lenalidomide, the exact mechanism has remained unclear. Interestingly, the more primitive hematopoietic compartment seems to possess a clonal advantage where del(5q) HSC are able to outcompete non-5q HSC. Whether the molecular mechanism for clonal advantage is intrinsic to the stem cell clone or a result of a compromised niche is unclear. However, we have previously demonstrated that lenalidomide is able to abrogate this clonal advantage. Aims: Our previous results implicated the matri-cellular protein SPARC to be upregulated in CD34+ del(5q) cells in response to lenalidomide. Thus, we sought to test the hypothesis that a putative decrease in SPARC expression of a clonal hematopoietic progenitor should give rise to a population of cellular descendants that exhibit an increased
adhesion to the microenvironment and more active rates of proliferation, thus explaining the clonal advantage of del(5q) MDS stem cells. Methods: We therefore studied the basal adhesion of del(5q) CD34+ progenitor cells to the murine stromal cell line MS-5 using a gravitational-force upon-inversion assay. Using multi-parameter flow cytometry, we studied proliferation and apoptosis in del(5q) and normal HSC and how these parameters are affected by lenalidomide and recombinant SPARC-protein. Results: Our data show that del(5q) CD34+ progenitors exhibited increased adhesion to the murine stromal cell line MS-5 compared to normal CD34+ cells. In short-term in vitro cultures, lenalidomide and recombinant SPARC were able to abrogate this increased adhesion. Multiparameter flow cytometry revealed a decreased rate of spontaneous apoptosis in del(5q) HSC compared to progenitors. Lenalidomide increased apoptosis in both compartments and this effect was specific to del(5q) cells. However, neither lenalidomide nor SPARC appeared to have any effect on the proliferation of CD34+ progenitors from normal or del(5q) bone marrow. Summary/Conclusion: These studies suggest that decreased expression of SPARC leads to increased adhesion of del(5q) HSC/progenitor cells to their microenvironment and may explain why del(5q) HSC are able to outcompete the remaining healthy HSC. Our studies suggest that lenalidomide is able to abrogate this clonal advantage partly via its increase in SPARC expression with a consecutive decrease in adhesion and increase in apoptosis.
4. Therapy and Experimental Models C029 HLA-identical umbilical cord blood transplantation from a sibling donor in juvenile myelomonocytic leukemia A. De Vries1 ° , R. Bredius2 , A. Lankester2 , M. Bierings3 , M. Trebo4 , P. Sedlacek5 , C. Niemeyer6 , M. Zecca7 , F. Locatelli7 , M. van den Heuvel-Eibrink1 . 1 Department of Pediatric Hematology/Oncology, ErasmusMC Sophia Childrens Hospital, Rotterdam, The Netherlands; 2 Department of Pediatrics, Leiden University Medical Center, Leiden, The Netherlands; 3 Department of Pediatric Hematology/Oncology, UMCU/Wilhelmina Childrens Hospital, Utrecht, The Netherlands; 4 Department of Pediatrics, St Anna Childrens Hospital, Vienna, Austria; 5 Department of Pediatric Hematology/Oncology, Charles University, University Hospital Motol, Prague, Czech Republic; 6 Department of Pediatrics, University of Freiburg, Freiburg, Germany; 7 IRCCS Policlinico San Matteo, Pediatric Hematology/Oncology, Pavia, Italy *E-mail: [email protected] Background: Currently, for juvenile myelomonocytic leukemia (JMML) stem cell transplantation is the only
4. Therapy and Experimental Models curative treatment option. Although successful unrelated donor umbilical cord blood transplantations (UCBTs) have been reported, series of HLA-identical sibling donor UCBTs in JMML are not available. Patients and Methods: From the European Working Group on Childhood MDS (EWOG MDS) registry 5 JMML patients who underwent a fully HLA matched sibling UCBT were identified. Results: The median age at diagnosis was 18 months (range 15−30 months). None of the patients had clinical signs of NF1. In one case a PTPN11 mutation was found. In two other cases no mutation in RAS or PTPN11 was found. In one case no mutation analysis was performed, in four cases the conditioning regimen consisted of busulfan, cyclophosphamide and melphalan and in one case of cyclophosphamide, etoposide and total body irradiation. All patients engrafted slowly (ANC > 500 ul: median 33 days, range 10−35 days, platelets >20,000/ul: median 53 days, range 38−77 days). In 3 patients acute graft versus host disease was noticed (grade 1 and 2); no chronic graft versus host disease was reported. Two patients relapsed after the initial transplantation and underwent a second transplantation with marrow of the initial donor. One of them is in second complete remission and the other died after a second relapse. One patient developed increased donor chimerism from day 42 without any clinical sign of relapse. She was treated with DLI, 6-mercaptopurine and 13-cis retinoic acid respectively. She is still in complete remission 5 years after transplantation. Conclusion: This EWOG series illustrates that, although this needs to be confirmed in larger series, transplantation with relatively immunologically naive cord blood stem cells from a HLA-matched sibling is feasible in selected cases of JMML.
C030 Long-term remission of post-transplant MDS/AML by adoptive transfer of allogeneic WT1-specific CD4+ and CD8+ T lymphocytes Y. Kim, N. Chung, S. Cho, M. Kim, E. Kim, H. Son, H. Choi, H. Kim, C. Min, S. Lee, D. Kim, W. Min, C. Kim, T. Kim ° . Catholic University of Korea, Seoul, South Korea *E-mail: [email protected] We treated a patient with post-transplant MDS/AML by adoptive transfer of Wilm’s tumor 1 (WT1)-specific cytotoxic T lymphocytes (WT1-CTL). WT1-CTL were generated by stimulating donor T cells with dendritic cells transduced with recombinant adenovirus encoding truncated WT1 (DWT1) created by deletion of the N-terminal portion of full length WT1. DWT1-CTL was infused at a cumulative dose of 8x107 cells/m2 after fludarabine-based chemotherapy and DLI/stem cell support. Although GVHD developed on skin, mouth, and eye 2 months after CTL
S49
therapy, steroid therapy was effective. The patient remained in sustained remission for 37 months after DWT1-CTL therapy until disease relapsed again. The frequencies of DWT1-specific T cells gradually increased over one year and then decreased in peripheral blood. IFN-g ELISPOT responses specific for DWT1 were detected in both CD4+ and CD8+ T cells at 2 years. Furthermore, IFN-g secreting CD8+ T cells specific for peptides located within DWT1 were detected with higher frequencies compared to those for peptide within the deleted N-terminal portion of WT1. These results show that in vitro cultured DWT1-specific CD4+ and CD8+ cells can be expanded and function to overcome immune tolerance in a leukemic patient, eliciting long-lasting anti-leukemic immunity. These results raise the possibility that adoptive transfer of in vitro cultured leukemia-specific CTL can be successfully applied to treat hematological malignancies in an allogeneic setting.
C031 Maintenance treatment with 5-azacitidine for patients with high risk myelodysplastic syndrome (MDS) or acute myeloid leukemia following MDS (MDS-AML) in complete remission (CR) after induction chemotherapy M. Groevdal1 , R. Kahn1 , M. Jansson1 ° , A. Aggerholm2 , P. Antunovic3 , J. Astermark4 , P. Bernell5 , L. Engstroem6 , L. Kjeldsen7 , O. Linder8 , L. Nilsson9 , A. Olsson10 , M. Skovholm11 , J. Tangen12 . 1 Karolinska Institutet, Stockholm, Sweden; 2 Aarhus University hospital, Denmark; 3 Linkoping University hospital, Sweden; 4 Malmoe University hospital, Sweden; 5 Karolinska University hospital, Sweden; 6 University hospital of Norrland, Sweden; 7 Rigshospitalet, Denmark; 8 Orebro University hospital, Sweden; 9 Lund University hospital, Sweden; 10 Sahlgrenska University hospital, Sweden; 11 Aalborg hospital, Denmark; 12 Ullevaal University hospital, Norway *E-mail: [email protected] Around 50% of patients with high-risk MDS or MDSAML may enter CR after induction chemotherapy, but CR duration, as well as overall survival is usually short. To address this clinical problem the Nordic MDS Group designed a prospective multicenter phase II study, which assessed the clinical feasibility and utility of long-term maintenance treatment with azacitidine. Sixty patients with high-risk MDS (IPSS intermediate-2 or high) (n = 23) or AML following a previous known MDS (n = 37) were enrolled between 2004 and 2006. The mean age was 68 (54−83) and patients should not be eligible for stem cell transplantation. Induction treatment consisted of standard doses of daunorubicin and ara-C. Patients in CR received low dose azacitidine subcutaneously 5/28 days until relapse, unless unacceptable toxicity developed. Methylation status of the P15ink4b (P15), E-cadherine (CDH ) and