Abstract: Hodgkin lymphoma relapsing after autologous transplantation (autoSCT) has a dismal outcome. Allogeneic transplantation (alloSCT) using reduced intensity conditioning (RIC) is a salvage option, but its effectiveness is still unclear. To evaluate the role of RIC alloSCT, we designed a retrospective study based on the commitment of attending physicians to perform a salvage alloSCT; thus, only Hodgkin lymphoma patients having human leukocyte antigen-typing immediately after the failed autoSCT were included. Of 185 patients, 122 found an identical sibling (55%), a matched unrelated (32%) or a haploidentical sibling (13%) donor; 63 patients did not find any donor. Clinical features of both groups did not differ. Two-year progression-free (PFS) and overall survival (OS) were better in the donor group (39.3% vs 14.2%, and 66% vs 42%, respectively, P < .001) with a median follow-up of 48 months. In multivariable analysis, having a donor was significant for better PFS and OS (P < .001). Patients allografted in complete remission showed a better PFS and OS. This is the largest study comparing RIC alloSCT versus conventional treatment after a failed autoSCT, indicating a survival benefit for patients having a donor.
Abstract: Recently vancomycin-resistant and sporadically linezolid-resistant Enterococcus species have been described in adults. We report what we believe to be the first case of a child with prolonged bone marrow aplasia following haematopoietic stem cell transplantation developing a fatal sepsis caused by Enterococcus faecium resistant to glycopeptides and linezolid.
Abstract: IMPORTANCE OF THE FIELD: Primary central nervous system lymphomas (PCNSL) are rare but potentially curable tumours. The overall outcome for PCNSL patients is unsatisfactory and several therapeutic questions remain open. Modest progress in outcome reflects difficulties in conducting randomized trials and scarce molecular and biological knowledge. AREAS COVERED IN THIS REVIEW: This review describes conventional and investigational treatments for PCNSL and focuses on the main questions for future clinical trials. PubMed and the authors' own files were utilized for references search. The terms 'PCNSL', 'primary AND CNS lymphoma', and 'CNS AND lymphoma' were used for PubMed queries. All papers published in English before November 2009 were considered. WHAT THE READER WILL GAIN: This review illustrates how the paradigm for PCNSL treatment changed during the 1990s from radiotherapy alone to the establishment of high-dose methotrexate-cytarabine combination as standard approach. We present promising data from Phase II studies and discuss questions for randomized trials. Finally, we offer a 5-year scenario for the management of PCNSL. TAKE-HOME MESSAGE: The methotrexate-cytarabine combination should currently be considered as the reference treatment for PCNSL. Well-designed randomized trials and biological studies deriving from the use of novel technologies will be crucial to further improve outcome in these patients.
Abstract: Graft-versus-host disease (GvHD) is one of the major complications of allogeneic hematopoietic stem cell transplantation, an otherwise highly effective therapeutic modality for patients affected by hematological diseases. The main inducers of GvHD are alloreactive donor T cells, which recognize host antigens presented by recipient cells. The critical role of lymphocytes in GvHD is well documented by the observation that T-cell depletion from the graft prevents GvHD. Unfortunately, the removal of donor lymphocytes from the graft increases the incidence of disease relapse and life-threatening infectious complications. Gene transfer technologies are promising tools to manipulate donor T-cell immunity to enforce graft-versus-tumor/graft-versus-infection while preventing or controlling GvHD. For this purpose, several cell and gene transfer approaches have been investigated at the preclinical level and implemented in clinical trials.Gene Therapy advance online publication, 27 May 2010; doi:10.1038/gt.2010.83.
Abstract: BACKGROUND: Neurotoxicity is a recognized complication of cyclosporine A (CSA) treatment. The incidence of severe CSA-related neurological complications following hematopoietic stem cell transplantation (HSCT) is 4-11%. METHODS: We describe 6 cases of CSA related neurotoxicity out of 67 matched related HSCT performed in paediatric Middle East patients affected by haemoglobinopaties (5 beta thalassemia major, 1 sickle cell disease-SCD). Conditioning regimen consisted of iv busulphan, cyclophosphamide and graft-versus-host-disease (GvHD) prophylaxis with CSA, methylprednisolone, methotrexate and ATG. RESULTS: All 6 patients presented prodromes such as arterial hypertension, headache, visual disturbances and vomiting, one to two days before overt CSA neurotoxicity. CSA neurotoxicity consisted of generalized seizures, signs of endocranial hypertension and visual disturbances at a median day of onset of 11 days after HSCT (range +1 to +40). Brain magnetic resonance imaging (MRI) performed in all subjects showed reversible leukoencephalopathy predominantly in the posterior regions of the brain (PRES) in 5/6 patients. EEG performed in 5/6 patients was always abnormal. Neurotoxicity was not explainable by high CSA blood levels, as all patients had CSA in the therapeutic range with a median of 178 ng/ml (range 69-250). CSA was promptly stopped and switched to tacrolimus with disappearance of clinical and radiological findings. All patients are symptoms-free at a median follow up of 882 days (range 60-1065). CONCLUSIONS: Our experience suggests that paediatric patients with haemoglobinopaties have a high incidence of CSA related neurological events with no correlation between serum CSA levels and neurotoxicity. Prognosis is good following CSA removal. Specific prodromes such as arterial hypertension, headache or visual disturbances occurring in the early post-transplant period should be carefully evaluated with electrophysiological and MRI-based imaging in order to intervene promptly and avoid irreversible sequels.
Abstract: Immune-mediated refractoriness to platelet transfusion is a major problem in patients undergoing HSCT. In a cohort of 50 pediatric patients affected by beta thalassemia coming from Middle East countries, we experienced a high incidence of refractoriness because of anti-HLA antibodies during post-HSCT aplasia. In a risk factors analysis, factors predicting a negative transfusion outcome were presence of spleen and the number of anti-HLA antibodies. We adopted a policy to select platelet donors by avoiding HLA antigens against which the patient had specific antibodies. Transfusion of dedicated units resulted in 26% refractoriness compared to 74% to random units (p < 0.0001). When dedicated transfusions were used, the presence of spleen did not influence transfusion outcome. Analyzing transfusion outcome depending on the degree of HLA match and ABO compatibility, 76% successful transfusions were obtained with HLA-matched- ABO compatible followed by 67% in HLA-1mismatch- ABO compatible or HLA-matched- ABO incompatible and by 46% in HLA-1mismatch- ABO incompatible. In conclusion, we provide evidence that the selection of platelet donors according to patient characteristics, anti-HLA antibodies and ABO matching, is successful in reducing platelet refractoriness in heavily alloimmunized thalassemia patients undergoing transplantation.
Abstract: We have previously shown that hemopoietic stem cell transplant (HSCT) recipients can be stratified on day+7 as having low, intermediate or a high risk of transplant-related mortality (TRM). With the aim of reducing TRM and GVHD, intermediate and high-risk patients (n=170) were randomized to receive anti-thymocyte globulin (ATG, thymoglobuline) on day+7 (n=84) or no treatment (n=86) (controls). There was a reduction of TRM from 35% in controls to 29% in ATG patients (P=0.3), of acute GVHD III-IV from 15 to 5% (P=0.02) and of chronic GVHD from 26 to 11% (P=0.03); survival was comparable. The predictive value of the day+7 score on TRM was confirmed for controls (19 vs 42% for intermediate vs high risk, respectively, P=0.03), whereas ATG abrogated this predictive effect (29 vs 29%). ATG reduced GVHD (P=0.006) in high-risk patients, but not in patients with an intermediate risk. In conclusion, we confirm that TRM can be predicted on the basis of day+7 laboratory values, after alternative donor HSCT; in high-, but not intermediate-risk patients, the administration of ATG on day+7 reduces GVHD. These results may represent a platform for risk-adapted post transplant immune modulation.
Abstract: Mixed chimerism (MC) and secondary graft failure are frequent events following SCT for thalassemia. There is limited information regarding the outcome of donor lymphocyte infusion (DLI) to prevent rejection, mainly from case reports describing only successful cases. We describe a series of seven children affected by beta-thalassemia treated with escalating doses of DLI for level 2-3 MC (donor<90%) following myeloablative SCT from a matched family donor. The infusions were safe and no acute or chronic GVHD were documented; five patients experienced neutropenia and thrombocytopenia resolving spontaneously. DLI was successful in converting to full donor chimerism two patients stratified in the low-risk class (Pesaro class II). Conversely, for five high-risk patients, DLI was not effective in preventing secondary graft failure. This limited series suggests that escalating doses of DLI are safe in thalassemia patients post myeloablative therapy but efficacy may be jeopardized by rapidly growing anti-donor alloimmunity in high-risk patients. We suggest giving escalating doses of donor T cells to attempt a graft-versus-thalassemia as soon as level 2-3 MC is detected.
Abstract: beta-thalassemia is a major health problem worldwide, and stem cell transplantation (SCT) is the only curative option. Oral Busulfan (Bu) based conditioning is widely used in this setting. Due to the variability of Bu systemic exposure, intravenous (i.v.) Bu has been proposed as a standard of care, with no need for drug monitoring and dose adjustment. Patients with beta-thalassemia from countries with limited resources might be at higher risk of erratic Bu metabolism because of liver dysfunction, severe iron overload, and specific ethnic/genetic features. We studied Bu pharmacokinetics in 53 children with advanced beta-thalassemia from Middle Eastern countries who underwent a total of 57 matched related donor SCTs. Forty-two percent of the children required dose adjustment because they did not achieve the therapeutic window after the first dose. With a Bu dose-adjustment policy, regimen-related toxicity was limited. At a median follow-up of 564 days, the probabilities of 2-year survival, current thalassemia-free survival, rejection, and treatment-related mortality were 96%, 88%, 21%, and 4%, respectively. Conditioning with i.v. Bu and dose adjustment is feasible and well tolerated, although recurrence of thalassemia remains an unsolved problem in children with advanced disease.
Abstract: Allogeneic hematopoietic stem cell transplantation (allo-SCT) from an HLA-matched related or unrelated donor is a curative option for patients with high-risk hematological diseases. In the absence of a matched donor, patients have been offered investigational transplantation strategies such as umbilical cord blood SCT or family haploidentical SCT. Besides the activity of the conditioning regimen, most of the antileukemic potential of allo-SCT relies on alloreactivity, promoted by donor lymphocytes reacting against patient-specific antigens, such as minor and major histocompatibility antigens, ultimately translating into cancer immunotherapy. Unfortunately, alloreactivity is also responsible for the most serious and frequent complication of allo-SCT: graft-versus-host-disease (GvHD). The risk of GvHD increases with the level of HLA disparity between host and donor, and leads to impaired quality of life and reduced survival expectancy, particularly among patients receiving transplants from HLA-mismatched donors. Gene transfer technologies are promising tools to manipulate donor T cell immunity to enforce the graft-versus-tumor effect, to promote functional immune reconstitution (graft vs. infection), and to prevent or control GvHD. To this purpose, several cell and gene transfer approaches have been investigated at the preclinical level, and are being implemented in clinical trials. Suicide gene therapy is to date the most extensive clinical application of T cell-based gene therapy. In several phase I-II clinical studies conducted worldwide this approach proved highly feasible, safe, and effective in promoting a dynamic and patient-specific modulation of alloreactivity. This review focuses on this approach.
Abstract: PURPOSE: Short imatinib pulses were added to chemotherapy to improve the long-term survival of adult patients with Philadelphia chromosome (Ph) -positive acute lymphoblastic leukemia (ALL), to optimize complete remission (CR) and stem-cell transplantation (SCT) rates. PATIENTS AND METHODS: Of 94 total patients (age range, 19 to 66 years), 35 represented the control cohort (ie, imatinib-negative [IM-negative] group), and 59 received imatinib 600 mg/d orally for 7 consecutive days (ie, imatinib-positive [IM-positive] group), starting from day 15 of chemotherapy course 1 and from 3 days before chemotherapy during courses 2 to 8. Patients in CR were eligible for allogeneic SCT or, alternatively, for high-dose therapy with autologous SCT followed by long-term maintenance with intermittent imatinib. RESULTS: CR and SCT rates were greater in the IM-positive group (CR: 92% v 80.5%; P = .08; allogeneic SCT: 63% v 39%; P = .041). At a median observation time of 5 years (range, 0.6 to 9.2 years), 22 patients in the IM-positive group versus five patients in the IM-negative group were alive in first CR (P = .037). Patients in the IM-positive group had significantly greater overall and disease-free survival probabilities (overall: 0.38 v 0.23; P = .009; disease free: 0.39 v 0.25; P = .044) and a lower incidence of relapse (P = .005). SCT-related mortality was 28% (ie, 15 of 54 patients), and postgraft survival probability was 0.46 overall. CONCLUSION: This imatinib-based protocol improved long-term outcome of adult patients with Ph-positive ALL. With SCT, post-transplantation mortality and relapse remain the major hindrance to additional therapeutic improvement. Additional intensification of imatinib therapy should warrant a better molecular response and clinical outcome, both in patients selected for SCT and in those unable to undergo this procedure.
Abstract: BACKGROUND: Although hematopoietic stem cell transplantation (HSCT) has been widely used to treat pediatric patients with beta-thalassemia major, there is lack of evidence to determine whether this treatment improves health related quality of life (HRQoL). DESIGN AND METHODS: We used child-self and parent-proxy reports to prospectively evaluate HRQoL in 28 thalassemia children from Middle Eastern countries who underwent allogeneic HSCT in Italy. The PedsQL 4.0 Generic Core Scales were administered to patients and their parents 1 month before and 3, 6, 18 months after transplantation. RESULTS: Two-year overall survival, thalassemia-free survival, mortality and rejection were 89.3%, 78.6%, 10.9% and 14.3%, respectively. The cumulative incidence of acute and chronic graft-versus host disease (GVHD) was 36% and 18%, respectively. Physical functioning significantly declined (p = 0.02) from baseline (median score 81.3) at 3 months after HSCT (median score 62.5). Major improvements (p = 0.04) were seen afterwards up to 18 months after HSCT (median score 93.7). Agreement between child- and parent proxy-ratings was high. Chronic GVHD was the most significant factor associated with lower HRQoL scores over time (p = 0.02). CONCLUSIONS: The child-self and parent-proxy reports showed an improvement in the HRQoL of thalassemia children after HSCT. Overall, our study provides preliminary evidence-based data to further support clinical decision-making in this area.
Abstract: Use of allogeneic transplantation for patients with acute myeloid leukemia (AML) depends mainly on the risk of the disease, and HLA matched donor availability. In patients with high-risk leukemia, in the absence of a HLA (human leukocyte antigen) matched donor, alternative donors such as unrelated umbilical cord blood (UCB) or haploidentical donor (haplo) have been currently used. Both strategies have important advantages such as shorter time to transplant, which is particularly relevant to patients requiring urgent transplantation, and tolerance of HLA mismatched graft that make possible that a donor can be found for virtually all patients. However, in spite of higher incidence of graft failure in UCB transplatation recipients and higher relapse incidence after haplo transplants, final outcomes seem to be comparable with HLA matched unrelated hematopoietic stem cell transplantation (bone marrow or peripheral blood). Therefore, the complexity of choosing the best alternative donor will depend on urgency of the transplantation, status and risk of the disease, donor criteria and center experience. Here we review the current status of UCBT and haplo transplants to treat adults with high-risk acute myeloid leukemia and we discuss the main issues associated with the use of both hematopoietic stem cell transplant approaches.
Abstract: BACKGROUND:: The use of positron emission tomography (PET) scanning in Hodgkin lymphoma (HL) and aggressive non-Hodgkin lymphoma (HG-NHL) has recognized prognostic value in patients who are receiving chemotherapy or undergoing autologous stem cell transplantation (SCT). In contrast, the role of PET before reduced-intensity conditioning (RIC) and followed by allogeneic SCT has not been investigated to date. METHODS:: PET was used to assess 80 patients who had chemosensitive disease (34 patients with HG-NHL and 46 patients with HL) before they underwent allogeneic SCT: 42 patients had negative PET studies, and 38 patients had positive PET studies. Patients underwent allograft from matched related siblings (n = 41) or alternative donors (n = 39). RESULTS:: At the time of the last follow-up, 48 patients were alive (60%), and 32 had died. The 3-year cumulative incidence of nonrecurrence mortality and disease recurrence was 17% and 40%, respectively. The cumulative incidence of disease recurrence was significantly lower in the PET-negative patients (25% vs 56%; P = .007), but there was no significant difference between the patients with or without chronic graft-versus-host disease (P = .400). The patients who had negative PET studies before undergoing allogenic SCT also had significantly better outcomes in terms of 3-year overall survival (76% vs 33%; P = .001) and 3-year progression-free survival (73% vs 31%; P = .001). On multivariate analysis, overall survival was influenced by PET status (hazard ratio [HR], 3.35), performance status (HR, 5.15), and type of donor (HR, 6.26 for haploidentical vs sibling; HR, 1.94 for matched unrelated donor vs sibling). CONCLUSIONS:: The current results indicated that PET scanning appears to be an accurate tool for assessing prognosis in patients who are eligible for RIC allografting. Cancer 2010. (c) 2010 American Cancer Society.
Abstract: In the past years, in the field of beta cell replacement for diabetes therapy, the easy availability of bone marrow (BM) and the widely consolidated clinical experience in the field of haematology have contributed to the development of strategy to achieve donor-specific transplantation tolerance. Recently, the potential role of BM in diabetes therapy has been reassessed from a different point of view. Diverse groups investigated the contribution of BM cells to beta cell replacement as direct differentiation into insulin-producing cells. More importantly, while direct differentiation is highly unlikely, a wide array of experimental evidences indicates that cells of BM origin are capable of facilitating the survival or the endogenous regeneration of beta cells through an as yet well-defined regeneration process. These new experimental in-vitro and in-vivo data will expand in the next future the clinical trials involving BM or BM-derived cells to cure both type 1 or type 2 diabetes in humans. In this review we recapitulate the history of use of BM in diabetes therapy and we provide the clinically relevant actual information about the participation of BM and BM-derived stem cells in islet cell regeneration processes. Furthermore, new aspects as employing BM as "feeder tissue" for pancreatic islet and new clinical use of BM in diabetes therapy are discussed.
Abstract: SCT from an HLA-compatible sibling donor is an adoptive immunotherapy for cytokine-refractory, metastatic clear-cell renal cell cancer (RCC). However, the recent introduction of targeted therapy compounds has reduced the interest in this therapeutic strategy. We have reanalyzed our series with the aim to assess long-term benefit from allografting. Twenty-five RCC patients received a reduced-intensity allograft from an HLA-identical sibling donor. All patients received a thiotepa, fludarabine and CY conditioning regimen, and a cyclosporine-based GVHD prophylaxis. Best response to allograft was evaluable in 24 patients: 1 CR, 4 PR, 12 minor response/stable disease, 7 progressive disease. One-year survival was 48%, and five-year survival was 20%. At a median observation time of 65 months, five patients are alive, one in CR, one in PR and three with stable disease. By multivariate analysis, C-reactive protein value before transplant, the number of CD34 + infused cells and disease status at day +90 significantly correlated with survival. Survival of patients at favorable/intermediate-risk according to the MSKCC score that underwent allografting was better in comparison to the survival predicted by historical controls. We conclude that 20% of cytokine-refractory RCC patients are alive long-term after allografting. Transplantation is able to induce long-term disease control in a fraction of relapsed RCC patients.
Abstract: Central nervous system (CNS) dissemination is a rare (4-5%) but usually fatal complication of aggressive lymphomas. Prophylaxis modalities to prevent CNS dissemination in aggressive lymphomas cannot be widely applied to every lymphoma patient since it is associated with increased risk of neurotoxicity. Therefore, identification of high-risk patients as the best candidates to receive CNS prophylaxis constitutes a major endpoint in the management of these malignancies. Various risk factors and models for CNS recurrence have been described. Parameters reflecting the extent and proliferation of the disease, like elevated serum lactate dehydrogenase levels, involvement of multiple extranodal sites, advanced stage and high age-adjusted International Prognostic Index (IPI) score, as well as the involvement of specific anatomic sites, like testes, orbit, paranasal sinuses, have been identified and confirmed as important to predict CNS dissemination. Management of this complication in aggressive lymphomas with conventional-dose chemotherapy is associated with disappointing results, while some preliminary but encouraging experiences suggest a potential role of high-dose chemotherapy and stem cell transplantation. The analysis of recent clinical studies could lead to advancement in the prognosis of aggressive lymphomas, but several questions regarding the optimum chemotherapy combination, the best conditioning regimen and the role of radiation therapy and intrathecal chemotherapy remain still unanswered. The purposes of the present review are to critically analyse current data on the risk of CNS dissemination in aggressive lymphomas, the clinical presentation of secondary CNS lymphomas and the efficacy of CNS prophylaxis as well as to discuss the available therapeutic options for this devastating event.
Abstract: BACKGROUND: We investigated the long-term outcome of gene therapy for severe combined immunodeficiency (SCID) due to the lack of adenosine deaminase (ADA), a fatal disorder of purine metabolism and immunodeficiency. METHODS: We infused autologous CD34+ bone marrow cells transduced with a retroviral vector containing the ADA gene into 10 children with SCID due to ADA deficiency who lacked an HLA-identical sibling donor, after nonmyeloablative conditioning with busulfan. Enzyme-replacement therapy was not given after infusion of the cells. RESULTS: All patients are alive after a median follow-up of 4.0 years (range, 1.8 to 8.0). Transduced hematopoietic stem cells have stably engrafted and differentiated into myeloid cells containing ADA (mean range at 1 year in bone marrow lineages, 3.5 to 8.9%) and lymphoid cells (mean range in peripheral blood, 52.4 to 88.0%). Eight patients do not require enzyme-replacement therapy, their blood cells continue to express ADA, and they have no signs of defective detoxification of purine metabolites. Nine patients had immune reconstitution with increases in T-cell counts (median count at 3 years, 1.07x10(9) per liter) and normalization of T-cell function. In the five patients in whom intravenous immune globulin replacement was discontinued, antigen-specific antibody responses were elicited after exposure to vaccines or viral antigens. Effective protection against infections and improvement in physical development made a normal lifestyle possible. Serious adverse events included prolonged neutropenia (in two patients), hypertension (in one), central-venous-catheter-related infections (in two), Epstein-Barr virus reactivation (in one), and autoimmune hepatitis (in one). CONCLUSIONS: Gene therapy, combined with reduced-intensity conditioning, is a safe and effective treatment for SCID in patients with ADA deficiency. (ClinicalTrials.gov numbers, NCT00598481 and NCT00599781.)
Abstract: OBJECTIVES: To describe the first in-depth analysis of both the T-cell responses against human herpesvirus-8 (HHV-8) and the HHV-8 viral load in 1 patient who developed iatrogenic HHV-8-associated-Kaposi's sarcoma (KS) following immunosuppressive treatment for undifferentiated arthritis and to review the literature on iatrogenic KS (IKS). METHODS: T-cell responses against HHV-8 lytic and latent antigens were analyzed by ex vivo enzyme-linked immunospot (Elispot) and HHV-8 viral load was assessed by quantitative polymerase chain reaction, in sequential peripheral blood samples from a 55-year-old woman who developed skin/mucosal and visceral KS, while receiving treatment with cyclosporine, methotrexate, and methylprednisolone for undifferentiated arthritis. RESULTS: KS may result from HHV-8 infection in patients undergoing immunosuppressive treatment for rheumatic diseases and this is the first case of IKS occurring in undifferentiated arthritis. A role for immune surveillance in the pathogenesis of IKS is supported by the observation of disease regression following discontinuation of immunosuppressive therapy. In a 4-year follow-up, we showed that variations of the virus-specific immune responses but not of the viral load correlated well with the disease course, characterized by 2 remission and subsequent relapse phases, following changes of immunosuppressive therapy. CONCLUSIONS: We have provided evidence of a clear-cut correlation between changes in immunologic markers of HHV-8 infection and the disease course of this viral associated tumor, concomitant with variations of immunosuppressive treatment. Thus, ex vivo enzyme-linked immunospot for HHV-8-specific T-cell responses represents a new tool for the clinical management of rheumatic patients with IKS.
Abstract: BACKGROUND: Transplantation of hematopoietic stem cells from partially matched family donors is a promising therapy for patients who have a hematologic cancer and are at high risk for relapse. The donor T-cell infusions associated with such transplantation can promote post-transplantation immune reconstitution and control residual disease. METHODS: We identified 43 patients who underwent haploidentical transplantation and infusion of donor T cells for acute myeloid leukemia or myelodysplastic syndrome and conducted post-transplantation studies that included morphologic examination of bone marrow, assessment of hematopoietic chimerism with the use of short-tandem-repeat amplification, and HLA typing. The genomic rearrangements in mutant variants of leukemia were studied with the use of genomic HLA typing, microsatellite mapping, and single-nucleotide-polymorphism arrays. The post-transplantation immune responses against the original cells and the mutated leukemic cells were analyzed with the use of mixed lymphocyte cultures. RESULTS: In 5 of 17 patients with leukemia relapse after haploidentical transplantation and infusion of donor T cells, we identified mutant variants of the original leukemic cells. In the mutant leukemic cells, the HLA haplotype that differed from the donor's haplotype had been lost because of acquired uniparental disomy of chromosome 6p. T cells from the donor and the patient after transplantation did not recognize the mutant leukemic cells, whereas the original leukemic cells taken at the time of diagnosis were efficiently recognized and killed. CONCLUSIONS: After transplantation of haploidentical hematopoietic stem cells and infusion of donor T cells, leukemic cells can escape from the donor's antileukemic T cells through the loss of the mismatched HLA haplotype. This event leads to relapse.
Abstract: The importance of HLA donor-recipient matching in unrelated haematopoietic SCT (HSCT) is the subject of debate. In this retrospective study, we analyzed 805 adult patients from the Italian Registry receiving HSCT for a haematological malignancy from January 1999 to June 2006 and correlated the degree of HLA matching with transplant outcome. All patient-donor pairs had high-resolution typing at HLA-A, -B, -C, -DRB1 and -DQB1. There was a significantly higher risk of overall mortality, non-relapse mortality, graft failure and acute GVHD (aGVHD) for patients receiving HSCT from an unrelated donor with one or more low- or high-resolution mismatch/es (Mm/s). When only a single HLA Mm is present (9/10 matched pairs), mortality risk is higher than among 10/10 matched pairs in patients transplanted with acute leukaemia in the first CR ('early' patients) but not in the other patients (advanced patients): HR=1.69, 95% CI=0.94-3.02, P=0.08; HR=1.03, 95% CI=0.80-1.32, P=0.82, for early and advanced patients, respectively. These results confirm that the advantage of a 10/10 match has a greater effect in early patients, thus suggesting that a 9/10 matched donor can be chosen in patients with advanced disease lacking a rapidly available 10/10 matched one.
Abstract: Long-term clinical remissions of leukemia, after allogeneic hematopoietic stem cell transplantation, depend on alloreactive memory T cells able to self-renew and differentiate into antileukemia effectors. This is counterbalanced by detrimental graft-versus-host disease (GVHD). Induction of a selective suicide in donor T cells is a current gene therapy approach to abrogate GVHD. Unfortunately, genetic modification reduces alloreactivity of lymphocytes. This associates with an effector memory (T(EM)) phenotype of gene-modified lymphocytes and may limit antileukemia effect. We hypothesized that alloreactivity of gene-modified lymphocytes segregates with the central memory (T(CM)) phenotype. To this, we generated suicide gene-modified T(CM) lymphocytes with a retroviral vector after CD28 costimulation and culture with IL-2, IL-7, or a combination of IL-7 and IL-15. In vitro, suicide gene-modified T(CM) cells self-renewed upon alloantigen stimulation and resisted activation-induced cell death. In a humanized mouse model, only suicide gene-modified T cells cultured with IL-7 and IL-15 persisted, differentiated in T(EM) cells, and were as potent as unmanipulated lymphocytes in causing GVHD. GVHD was halted through the activation of the suicide gene machinery. These results warrant the use of suicide gene-modified T(CM) cells cultured with IL-7 and IL-15 for the safe exploitation of the alloreactive response against cancer.
Abstract: Dendritic cell (DC) targeting in vivo has recently been shown to confer strong and protective cytotoxic T lymphocyte (CTL)-based immunity in tumor murine models. Our group has recently demonstrated in preclinical models that the infusion of genetically modified lymphocytes (GMLs) expressing the self/tumor antigen TRP-2 is able to elicit functional TRP-2-specific effectors with antitumor activity by targeting DCs in vivo. Here we have analyzed vaccine- and tumor-specific immune responses of 10 melanoma patients treated with autologous GMLs expressing the cancer germline gene MAGE-A3. Three of 10 patients treated with MAGE-A3-GML showed an increase of circulating anti-MAGE-A3 T cells, and developed skin delayed-type hypersensitivity to MAGE-A3. Interestingly, in 2 of these patients, with progressive and measurable tumors at study entry, anti-MAGE-A3 T cells were detected not only in the blood but also within tumors resected after vaccination. These results demonstrate that the infusion of MAGE-A3-GML elicits antitumor T cells, which are capable of trafficking to inflamed tissues and of infiltrating tumors. Clinical studies on a larger group of patients are needed to evaluate the clinical efficacy of such a strategy.
Abstract: The importance of donor-recipient human leukocyte antigen (HLA)-DPB1 matching for the clinical outcome of unrelated hematopoietic stem cell transplantation (HSCT) is controversial. We have previously described an algorithm for nonpermissive HLA-DPB1 disparities involving HLA-DPB1*0901,*1001,*1701,*0301,*1401,*4501, based on T-cell alloreactivity patterns. By revisiting the immunogenicity of HLA-DPB1*02, a modified algorithm was developed and retrospectively tested in 621 unrelated HSCTs facilitated through the Italian Registry for oncohematologic adult patients. The modified algorithm proved to be markedly more predictive of outcome than the original one, with significantly higher Kaplan-Meier probabilities of 2-year survival in permissive compared with nonpermissive transplantations (55% vs 39%, P = .005). This was the result of increased adjusted hazards of nonrelapse mortality (hazard ratio [HR] = 1.74; confidence interval [CI], 1.19-2.53; P = .004) but not of relapse (HR = 1.02; CI, 0.73-1.42; P = .92). The increase in the hazards of overall mortality by nonpermissive HLA-DPB1 disparity was similar in 10 of 10 (HR = 2.12; CI, 1.23-3.64; P = .006) and 9 of 10 allele-matched transplantations (HR = 2.21; CI, 1.28-3.80; P = .004), both in early-stage and in advanced-stage disease. These data call for revisiting current HLA matching strategies for unrelated HSCT, suggesting that searches should be directed up-front toward identification of HLA-DPB1 permissive, 10 of 10 or 9 of 10 matched donors.
Abstract: The role of 18FDG-PET/CT during follow-up of patients affected by Hodgkin's lymphoma (HL) in complete remission after treatment is not fully elucidated, since a wide use of 18F fluorodeoxyglucose positron emission tomography/computed tomography (18FDG-PET/CT) in this setting could be limited by a relative high rate of false-positive results. Herein, we summarize a retrospective analysis of 27 patients with Hodgkin's lymphoma in complete remission after the first-line (n = 20) or salvage (n = 7) therapy receiving serial 18FDG-PET/CT scans during follow-up. Out of 165 scans, 13 were suspected for relapse, which was confirmed in seven patients. All relapses were correctly identified by 18FDG-PET/CT positivity, with a 100% sensitivity; false-positive rate was 46% and negative predictive value was 100%. True-positive findings were mostly associated with multiple sites, subdiaphragmatic involvement, and/or previous sites of disease. According to our results, we conclude that performing routine PET/CT scan during follow-up of those patients who are at high risk of relapse would be advisable, although caution must be adopted when interpreting PET/CT results due to the relatively high rate of false-positive findings. If FDG abnormal uptake is present at multiple nodal sites, subdiaphragmatic lymph nodes, or previous sites of disease, histological verification of PET abnormal findings is warranted.
Abstract: Clinical risk classification is inaccurate in predicting relapse in adult patients with acute lymphoblastic leukemia, sometimes resulting in patients receiving inappropriate chemotherapy or stem cell transplantation (SCT). We studied minimal residual disease (MRD) as a predictive factor for recurrence and as a decisional tool for postconsolidation maintenance (in MRD(neg)) or SCT (in MRD(pos)). MRD was tested at weeks 10, 16, and 22 using real-time quantitative polymerase chain reaction with 1 or more sensitive probes. Only patients with t(9;22) or t(4;11) were immediately eligible for allogeneic SCT. Of 280 registered patients (236 in remission), 34 underwent an early SCT, 60 suffered from relapse or severe toxicity, and 142 were evaluable for MRD at the end of consolidation. Of these, 58 were MRD(neg), 54 MRD(pos), and 30 were not assessable. Five-year overall survival/disease-free survival rates were 0.75/0.72 in the MRD(neg) group compared with 0.33/0.14 in MRD(pos) (P = .001), regardless of the clinical risk class. MRD was the most significant risk factor for relapse (hazard ratio, 5.22). MRD results at weeks 16 to 22 correlated strongly with the earlier time point (P = .001) using a level of 10(-4) or higher to define persistent disease. MRD analysis during early postremission therapy improves risk definitions and bolsters risk-oriented strategies. ClinicalTrials.gov identifier: NCT00358072.
Abstract: Hepatic veno-occlusive disease (VOD) is a common complication of haematopoietic stem cell transplantation (HSCT), with reported incidences of 5-40% in children. Recently, defibrotide (DF) has been successfully used as prophylaxis and treatment of VOD. This study reports data on 63 human leucocyte antigen-matched HSCT performed in 57 children affected by beta thalassemia at very high risk for developing VOD (liver fibrosis, iron overload, hepatitis C virus infections, busulphan-based conditioning, methotraexate + ciclosporine). All patients received a busulphan-based conditioning regimen, either orally (four HSCT) or intravenously (59 HSCT). All patients received oral DF (40 mg/kg per day, final dose) as VOD prophylaxis from median day -9 to median day +29. In order to overcome the lack of oral paediatric formulations, a galenic formulation was administered. DF was well tolerated. Only one patient fulfilled Seattle Criteria for VOD diagnosis. This patient had discontinued DF 6 d prior to VOD onset, due to high risk of haemorrhage. We concluded that oral defibrotide prophylaxis and i.v. busulphan safely abated VOD incidence in high-risk patients who had undergone HSCT. A galenic preparation of oral DF also permits this treatment in low-weight patients. Costs of DF prophylaxis are acceptable considering the reduced incidence of VOD.
Abstract: BACKGROUND: Procedures to prevent severe graft-versus-host disease (GVHD) delay immune reconstitution secondary to transplants of haploidentical haemopoietic stem cells for the treatment of leukaemia, leading to high rates of late infectious mortality. We aimed to systematically add back genetically engineered donor lymphocytes to facilitate immune reconstitution and prevent late mortality. METHODS: In a phase I-II, multicentre, non-randomised trial of haploidentical stem-cell transplantation, we infused donor lymphocytes expressing herpes-simplex thymidine kinase suicide gene (TK-cells) after transplantation. The primary study endpoint was immune reconstitution defined as circulating CD3+ count of 100 cells per muL or more for two consecutive observations. Analysis was by intention to treat. This trial is registered with ClinicalTrials.gov, number NCT00423124. FINDINGS: From Aug 13, 2002, to March 26, 2008, 50 patients (median age 51 years, range 17-66) received haploidentical stem-cell transplants for high-risk leukaemia. Immune reconstitution was not recorded before infusion of TK-cells. 28 patients received TK-cells starting 28 days after transplantation; 22 patients obtained immune reconstitution at median 75 days (range 34-127) from transplantation and 23 days (13-42) from infusion. Ten patients developed acute GVHD (grade I-IV) and one developed chronic GVHD, which were controlled by induction of the suicide gene. Overall survival at 3 years was 49% (95% CI 25-73) for 19 patients who were in remission from primary leukaemia at the time of stem-cell transplantation. After TK-cell infusion, the last death due to infection was at 166 days, this was the only infectious death at more than 100 days. No acute or chronic adverse events were related to the gene-transfer procedure. INTERPRETATION: Infusion of TK-cells might be effective in accelerating immune reconstitution, while controlling GVHD and protecting patients from late mortality in those who are candidates for haploidentical stem-cell transplantation. FUNDING: MolMed SpA, Italian Association for Cancer Research.
Abstract: PURPOSE: To investigate the impact of adding rituximab to intensive chemotherapy with peripheral-blood progenitor cell (PBPC) autograft for high-risk diffuse large B-cell lymphoma (DLB-CL) and follicular lymphoma (FL). PATIENTS AND METHODS: Data were collected from 10 centers associated with Gruppo Italiano Terapie Innnovative nei Linfomi for 522 patients with DLB-CL and 223 patients with FL (median age, 47 years) who received the original or a modified high-dose sequential (HDS) chemotherapy regimen. HDS was delivered to 396 patients without (R-) and to 349 patients with (R+) rituximab; 154 (39%) and 178 patients (51%) in the R- and R+ subsets, respectively, underwent HDS for relapsed/refractory disease. RESULTS: A total of 355 R- (90%) and 309 R+ patients (88%) completed the final PBPC autograft. Early treatment-related mortality was 3.3% for R- and 2.8% for R+ (P = not significant). Two parameters significantly influenced the outcome: disease status at HDS, with 5-year overall survival (OS) projections of 69% versus 57% for diagnosis versus refractory/relapsed status, respectively, and rituximab addition, with 5-year OS of 69% versus 60% in the R+ versus R- groups, respectively. In the multivariate analysis, these two variables maintained an independent prognostic value. The marked benefit of rituximab was evident in patients receiving HDS as salvage treatment: the 5-year OS projections for R+ versus R- were, respectively, 64% versus 38%, for patients with refractory disease or early relapse and 71% versus 57%, for patients with late relapse, partial response, or second/third relapse. CONCLUSION: The results of this large series indicate that rituximab should be included in the current practice of PBPC autograft for DLB-CL and FL.
Abstract: In order to control graft-versus-host disease after donor lymphocyte infusion, T cells can be retrovirally transduced with a suicide gene. However, the immune competence of activated T cells appears compromised, responsible for reduced alloreactivity. The present study compared different activation protocols using soluble or bead-coupled antibodies regarding T-cell subtype expansion capacity and functionality. T cells were purified on a laboratory and clinical scale using both CD3 and CD4/CD8 antibodies for selection, leading to a mean purity of 96%. Transductions were performed with a GMP-grade CD34/HSV-TK vector. Activation with soluble CD3/CD28-antibodies +1000 U/ml IL-2 induced a 50-fold expansion of T cells over 14 days, whereas T cells activated with bead-coupled antibodies only expanded 2-4-fold restricted to the first week. Apart from using soluble antibodies, proliferation was highly IL-2 dependent. Expansion of CMV-specific T cells coincided with the expansion of whole CD3(+) cells. Soluble antibodies and higher IL-2 concentrations preferentially stimulated CD8(+) T cells, while bead-coupled antibodies +20 U/ml IL-2 preserved the CD4/CD8 ratio. Irrespective of the activation protocol, there was a shift from a naive to memory phenotype. When activated with soluble antibodies, mainly CD8(+) T cells were transduced. Furthermore, Th1/Th2 cytokine secretion was reduced. In contrast, CD4(+)/CD8(+) T cells activated with bead-coupled antibodies were rather homogenously transduced and cytokine secretion did not appear to be affected.
Abstract: We evaluated the prognostic role of 18FDG-PET performed before ASCT in patients affected by lymphoma who underwent high-dose chemotherapy followed by ASCT as first-line treatment for high-risk disease or as second-line or more for relapsed or refractory disease. We retrospectively analyzed 53 consecutive patients, 14 with Hodgkin Lymphoma (HL) and 39 with non-Hodgkin Lymphoma (NHL), treated between February 1999 and October 2006 at our institution, who had a pre-ASCT FDG-PET (pPET) evaluation. Median age was 45 years (range: 18-69). After a median follow-up of 31 months (range: 8-91), 7 out of 16 pPET+ patients and 10 out of 37 pPET- patients experienced lymphoma relapse. The 5-year OS is 90% and 55% (p = 0.01) in patients with negative and positive pPET, respectively. In conclusion, a positive pPET indicates a poorer outcome after ASCT with respect to a negative pPET; this subset of patients should be considered candidate to more intensive or investigational approaches.
Abstract: Allogeneic BMT represents the only chance of cure for beta-thalassemia. Occasionally, two affected individuals from the same family share a matched healthy sibling. Moreover, a high incidence of transplant rejection is still observed in Pesaro class III patients, requiring a second BMT procedure. In these settings, one option is to perform a second BM harvest from the same donor. Although BM harvest is a safe procedure in children, ethical issues concerning this invasive practice still arise. Here, we describe our series of seven pediatric, healthy donors, who donated BM more than once in favor of their beta-thalassemic HLA-identical siblings between June 2005 and January 2008. Three donors donated BM twice to two affected siblings and four donors donated twice for the same sibling following graft rejection of the first BMT. All donors tolerated the procedures well and no relevant side effects occurred. There was no significant difference between the two harvests concerning cell yield and time to engraftment. Our experience shows that for pediatric donors, a second BM donation is safe and feasible and good cellularity can be obtained. We suggest that a second harvest of a pediatric donor can be performed when a strong indication for BMT exists.
Abstract: Haploidentical hematopoietic stem cell transplantation (haplo-HSCT) is an alternative treatment to patients with high-risk acute leukemia lacking a human leukocyte antigen-matched donor. We analyzed 173 adults with acute myeloid leukemia (AML) and 93 with acute lymphoblastic leukemia (ALL) who received a haplo-HSCT in Europe. All grafts were T cell-depleted peripheral blood progenitor cells from a direct family or other related donor. At transplantation, there were 25 patients with AML in CR1 (complete remission 1), 61 in more than or equal to CR2, and 87 in nonremission, and 24 with ALL in CR1, 37 in more than or equal to CR2, and 32 in nonremission. Median follow-up was 47 months in AML and 29 months in the ALL groups. Engraftment was observed in 91% of the patients. Leukemia-free survival at 2 years was 48% plus or minus 10%, 21% plus or minus 5%, and 1% for patients with AML undergoing transplantation in CR1, more than or equal to CR2, and nonremission, and 13% plus or minus 7%, 30% plus or minus 8%, and 7% plus or minus 5% in ALL patients, respectively. In conclusion, haplo-HSCT can be an alternative option for the treatment of high-risk acute leukemia patients in remission, lacking a human leukocyte antigen-matched donor.
Abstract: In nine patients with posttransplant Kaposi sarcoma (KS) T-cell responses to human herpesvirus (HHV)-8 latent and lytic antigens, as detected by enzyme-linked-immunospot (Elispot) assay, were absent at disease onset. Virus-specific T-cell responses were detected in six renal recipients at remission after a reduction of calcineurin inhibitors (CIs), and in two HHV-8 seropositive renal recipients without KS. In two liver recipients undergoing switch from CIs to sirolimus (SRL), normalization of the T-cell repertoire and recovery of both HHV-8-specific effector and memory T lymphocytes were associated with complete KS remission. In a renal recipient undergoing SRL conversion, the early recovery of HHV-8-specific effector but not of memory T lymphocytes, was associated only with partial remission. Neither rejection nor changes in graft function were observed after SRL conversion. HHV-8-specific T-cell responses are required to achieve posttransplant KS remission, and may be restored under SRL, while maintaining effective immunosuppression.
Abstract: There is a substantial incidence of graft failure in patients with thalassemia after myeloablative conditioning regimens especially in class 3 patients in whom its incidence could be as high as 8-38.5%. Most patients with graft failure have recurrence of thalassemic marrow. Historically, results of second transplants for thalassemia were poor because of a high rejection rate and/or increased TRM. Sixteen patients with thalassemia recurrence following rejection of the first graft and with a median age of 9 years (range, 4-20) were given second transplants using BM (n=7) or PBSC (n=9) after preparation with a new treatment protocol. All but two patients received stem cells from the same donor. The median interval between two transplants was 28 months (range, 8-204). The sustained engraftment rate was high (94%) with only one patient having primary graft failure. The probability of overall survival, event-free survival, TRM and graft failure were 79, 79, 16 and 6%, respectively. There were three transplant-related deaths. Thirteen patients are alive with Lansky/Karnofsky score of 100. This intensified treatment protocol was well tolerated with no significant increase in toxicity. The excellent results obtained with this new preparative regimen allow us to recommend it for second transplantation for patients with thalassemia recurrence.
Abstract: In this study, we have characterized reconstitution of the natural killer (NK) cell repertoire after haploidentical CD34(+) selected hematopoietic stem cell transplantation (HSCT) for high-risk hematologic malignancies. Analysis focused on alloreactive single-KIR(+) NK cells, which reportedly are potent antileukemic effectors. One month after HSCT, CD56(bright)/CD56(dim) NK-cell subsets showed inverted ratio and phenotypic features. CD25 and CD117 down-regulation on CD56(bright), and NKG2A and CD62L up-regulation on CD56(dim), suggest sequential CD56(bright)-to-CD56(dim) NK-cell maturation in vivo. Consistently, the functional potential of these maturation intermediates against leukemic blasts was impaired. Mature receptor repertoire reconstitution took at least 3 months. Importantly, at this time point, supposedly alloreactive, single-KIR(+) NK cells were not yet fully functional. Frequency of these cells was highly variable, independently from predicted NK alloreactivity, and below 1% of NK cells in 3 of 6 alloreactive patients studied. In line with these observations, no clinical benefit of predicted NK alloreactivity was observed in the total cohort of 56 patients. Our findings unravel the kinetics, and limits, of NK-cell differentiation from purified haploidentical hematopoietic stem cells in vivo, and suggest that NK-cell antileukemic potential could be best exploited by infusion of mature single-KIR(+) NK cells selected from an alloreactive donor.
Abstract: Gene therapy is a promising therapeutic strategy for genetic and acquired hematologic diseases. With the improvements in gene transfer and expression, factors affecting safety and efficacy of gene therapy can now be evaluated to establish the best clinical benefit-to-risk ratio. The induction of immune responses against gene therapy components is one of the potential limitations. We studied the occurrence of such event in 23 patients treated with donor lymphocyte infusions (DLIs), with lymphocytes transduced to express the HSV-TK suicide gene for relapse of hematologic malignancies occurring after allogeneic hematopoietic stem cell transplantation (HSCT). The suicide gene was used to selectively control graft-versus-host disease (GvHD). Seven patients given infusions late after HSCT developed an immune response against the transgene. Immunization involved appearance of thymidine kinase (TK)-specific CD8(+) effectors and required a level of immunocompetence at the time of TK-DLI that can be achieved only several months after transplantation. This did not prevent graft-versus-leukemia (GvL) effect of the TK-DLI, since 5 of 7 immunized patients maintained the complete remission achieved prior to immunization. We suggest that appropriate study designs taking into account the immune suppression of the patient and time-kinetics of GvL mediated by TK-transduced donor lymphocytes may allow the full exploitation of TK-DLI.
Abstract: The majority of unrelated donor (UD) hematopoietic stem cell (HSC) transplants are performed across HLA-DP mismatches, which, if involving disparity in a host-versus-graft (HVG) direction for an alloreactive T cell epitope (TCE), have been shown by our group to be associated with poor clinical outcome in 2 cohorts of patients transplanted for hematopoietic malignancies and beta-thalassemia, respectively. Using site-directed mutagenesis of DPB1*0901, we show here that the TCE is abrogated by the presence of amino acids LFQG in positions 8-11 of the DP beta-chain. Based on this and on alloreactive T cell responsiveness, we have determined the presence or absence of the TCE for 72 DPB1 alleles reported in the ethnic groups representative of the worldwide UD registries, and predict that 67%-87% (mean 77%) of UD recipient pairs will not present a DPB1 TCE disparity in the HVG direction. We developed and validated in 112 healthy Italian blood donors an innovative approach of DPB1 epitope-specific typing (EST), based on 2 PCR reactions. Our data show that DPB1 TCE disparities may hamper the clinical success of a considerable number of transplants when DPB1 matching is not included into the donor selection criteria, and that a donor without DPB1 TCE disparities in the HVG direction can be found for the majority of patients. Moreover, the study describes the first protocol of targeted epitope-specific DPB1 donor-recipient matching for unrelated HSC transplantation. This protocol will facilitate large-scale retrospective clinical studies warranted to more precisely determine the clinical relevance of DPB1 TCE disparities in different transplant conditions.
Abstract: The transfer of a suicide gene into donor lymphocytes to control alloreactivity in the context of allogeneic hematopoietic stem cell transplantation (allo-HSCT) represents the widest clinical application of T-cell based gene transfer, as shown by more than 100 patients treated worldwide to date, several phase I-II studies completed, and a registrative phase III study, sponsored by a biotech firm, about to begin. In this mini-review, we will summarize the clinical results obtained to date, and attempt to identify the steps envisaged to optimize the suicide gene therapy approach.
Abstract: The exploitation of the physiologic processing and presenting machinery of DCs by in vivo loading of tumor-associated antigens may improve the immunogenic potential and clinical efficacy of DC-based cancer vaccines. Here we show that lymphocytes genetically modified to express self/tumor antigens, acting as antigen carriers, efficiently target DCs in vivo in tumor-bearing mice. The infusion of tyrosinase-related protein 2-transduced (TRP-2-transduced) lymphocytes induced the establishment of protective immunity and long-term memory in tumor-bearing mice. Analysis of the mechanism responsible for the induction of such an immune response allowed us to demonstrate that cross-presentation of the antigen mediated by the CD11c(+)CD8alpha(+) DC subset had occurred. Furthermore, we demonstrated in vivo and in vitro that DCs had undergone activation upon phagocytosis of genetically modified lymphocytes, a process mediated by a cell-to-cell contact mechanism independent of CD40 triggering. Targeting and activation of secondary lymphoid organ-resident DCs endowed antigen-specific T cells with full effector functions, which ultimately increased tumor growth control and animal survival in a therapeutic tumor setting. We conclude that the use of transduced lymphocytes represents an efficient method for the in vivo loading of tumor-associated antigens on DCs.
Abstract: The extensive exploitation of the antitumor effect of donor lymphocytes infused after allogeneic hematopoietic stem-cell transplantation (allo-HSCT) is limited by the risk of graft-versus-host disease (GvHD). To overcome this limitation, we investigated the therapeutic potential of donor lymphocytes engineered with the suicide gene thymidine kinase of herpes simplex virus (TK) in 23 patients experiencing recurrence of hematologic malignancies after allo-HSCT. Long-term follow-up of infused patients included analysis of engraftment of genetically engineered lymphocytes, in vivo assessment of antitumor effect, and control of GvHD by ganciclovir. All 17 patients evaluable for engraftment and graft-versus-leukemia (GvL) had circulating TK(+) cells detectable beginning at a median time of 18 days. Eleven patients (65%) experienced a substantial clinical benefit resulting in 6 (35%) complete remissions and 5 (29%) partial responses. The antitumor effect tightly correlated with the in vivo expansion of TK(+) cells. Seven patients received ganciclovir, resulting in elimination of TK(+) cells and effective and selective treatment of GvHD. Immunization against HSV-TK was observed in 7 patients but did not preclude an effective GvL. These data validate the feasibility, safety, and efficacy of TK(+) cells in the context of allografting and represent the basis for a broader application of this technology.
Abstract: Minor histocompatibility antigens (mHags) HA-1 and HA-2 are encoded by biallelic loci, with immunogenic variants, HA-1H and HA-2V, which induce strong HLA-A2-restricted alloreactive T-cell responses, and nonimmunogenic counterparts, HA-1R and HA-2M, which represent functional null alleles that are poorly presented by HLA class I molecules. HA-1 and HA-2 are potential targets of selective graft-versus-leukemia and graft-versus-tumor reactivity after allogeneic hematopoietic stem cell transplantation (HSCT); however, these applications are restricted to a limited number of patients. Here, we show that a far more frequent application of HA-1 and HA-2 disparity relies on their use as markers for the state of host chimerism after allogeneic HSCT. We have determined allelic frequencies of 29.3% and 70.7% for HA-1H and HA-1R, respectively, and of 83.7% and 16.3% for HA-2V and HA-2M, respectively, in >200 healthy individuals from northern Italy. Similar frequencies were observed in nearly 100 patients affected by hematologic malignancies or solid tumors, thus showing that HA-1 and HA-2 variability are not associated with the presence of cancer. On the basis of these data, we predict that HA-1 and HA-2 can be used in 32.8% and 23.5% of Italian transplant patients, respectively, as markers for the state of host chimerism, whereas exploitation of disparity for these mHags for targeted immunotherapy will be possible in 10.7% and 1.1% of Italian patients, respectively. Retrospective HA-2 typing of bone marrow aspirates obtained from a patient during complete remission or recurrence of acute myeloid leukemia after haploidentical HSCT showed the feasibility of using HA-2 as a surrogate marker for disease monitoring. Because of an apparent north-south gradient for HA-1 allelic frequencies, with higher frequencies for the HA-1H variant reported in white populations from Southern Europe as compared with Northern Europe and North America, the diagnostic applicability of HA-1 disparity will be slightly more frequent in transplant patients from the north. Taken together, our data show that determination of HA-1 and HA-2 variability can be an important parameter for the selection of allogeneic stem cell donors, in particular for patients affected by hematologic malignancies without a tumor-specific molecular marker.
Abstract: In allogeneic hematopoietic cell transplantation (allo-HCT), the immune recognition of host antigens by donor T lymphocytes leads to a beneficial graft-versus-leukemia (GvL) effect as well as to life-threatening graft-versus-host disease (GvHD). Genetic modification of T lymphocytes with a retroviral vector (RV) expressing the herpes simplex virus-thymidine kinase (TK) suicide gene confers selective sensitivity to the prodrug ganciclovir (GCV). In patients, the infusion of TK+ lymphocytes and the subsequent administration of GCV resulted in a time-wise modulation of antihost reactivity for a GvL effect, while controlling GvHD. Because activation required for genetic modification with RV may reduce antihost reactivity, we investigated the requirements for maximizing the potency of human TK+ lymphocytes. Whereas T-cell receptor triggering alone led to effector memory (EM) TK+ lymphocytes, the addition of CD28 costimulation through cell-sized beads resulted in the generation of central memory (CM) TK+ lymphocytes. In a quantitative model for GvHD using nonobese diabetic/severely combined immunodeficient mice, CM TK+ lymphocytes were more potent than EM TK+ lymphocytes. GCV administration efficiently controlled GvHD induced by CM TK+ lymphocytes. These results warrant the clinical investigation of CM suicide gene-modified human T lymphocytes for safe and effective allo-HCT.
Abstract: The use of retroviral vectors in gene therapy has raised safety concerns for the genotoxic risk associated with their uncontrolled insertion into the human genome. We have analyzed the consequences of retroviral transduction in T cells from leukemic patients treated with allogeneic stem cell transplantation and donor lymphocytes genetically modified with a suicide gene (HSV-TK). Retroviral vectors integrate preferentially within or near transcribed regions of the genome, with a preference for sequences around promoters and for genes active in T cells at the time of transduction. Quantitative transcript analysis shows that one fifth of these integrations affect the expression of nearby genes. However, transduced T cell populations maintain remarkably stable gene expression profiles, phenotype, biological functions, and immune repertoire in vivo, with no evidence of clonal selection up to 9 yr after administration. Analysis of integrated proviruses in transduced cells before and after transplantation indicates that integrations interfering with normal T cell function are more likely to lead to clonal ablation than expansion in vivo. Despite the potentially dangerous interactions with the T cell genome, retroviral integration has therefore little consequence on the safety and efficacy of T cell transplantation.
Abstract: Classic Hodgkin's Lymphoma is a disease usually characterized by lymph nodal presentation. Rare cases with extra-nodal first clinical manifestation may occur, and primary bone marrow involvement may be part of them: isolated bone marrow forms have been recently described in the context of HIV infection. We herein report a case of isolated bone marrow Hodgkin's Lymphoma occurrence in a HIV-negative, elderly patient. The impacts of such unusual Hodgkin's Lymphoma manifestation on clinical practice as well as the related diagnostic and therapeutic implications are discussed.
Abstract: Tens of thousands of man-made chemicals are in regular use and discharged into the environment. Many of them are known to interfere with the hormonal systems in humans and wildlife. Given the complexity of endocrine systems, there are many ways in which endocrine-disrupting chemicals (EDCs) can affect the body's signaling system, and this makes unraveling the mechanisms of action of these chemicals difficult. A major concern is that some of these EDCs appear to be biologically active at extremely low concentrations. There is growing evidence to indicate that the guiding principle of traditional toxicology that "the dose makes the poison" may not always be the case because some EDCs do not induce the classical dose-response relationships. The European Union project COMPRENDO (Comparative Research on Endocrine Disrupters--Phylogenetic Approach and Common Principles focussing on Androgenic/Antiandrogenic Compounds) therefore aims to develop an understanding of potential health problems posed by androgenic and antiandrogenic compounds (AACs) to wildlife and humans by focusing on the commonalities and differences in responses to AACs across the animal kingdom (from invertebrates to vertebrates) .
Abstract: Human cytomegalovirus (HCMV) infection is the most frequent infectious complication after conventional allogeneic stem cell transplantation (alloSCT). From December 1998 to December 2002, we prospectively monitored HCMV reactivation in 59 patients affected by solid tumors and undergoing nonmyeloablative alloSCT (NST). Patients were allografted from HLA-identical sibling donors after fludarabine/cyclophosphamide-based conditioning regimens. Seventeen (28.8%) of 59 patients presented with HCMV antigenemia, and 14 received ganciclovir, with successful HCMV clearance in all cases. No patient developed HCMV viremia or disease. The median time to HCMV reactivation was 54 days (range, 16-245 days) after NST. These patients were compared with a cohort of hematologic patients who were treated with conventional myeloablative alloSCT. Matching criteria included HCMV risk group, stem cell source, donor type, and age. In the myeloablative group, HCMV active infection was observed in 47 (85.4%) of 55 patients at a median time of 30 days (range, 13-64 days) after alloSCT, and HCMV infection occurred more frequently ( P < .001) and earlier ( P = .001) than in NST patients. Patients affected with solid tumors undergoing NST had a reduced and delayed incidence of HCMV active infection.
Abstract: Allogeneic hematopoietic stem cell transplantation (allo-SCT) is the treatment of choice for many malignant diseases. It is recognized that the curative potential of allo-SCT relates closely to the immune advantage conferred by allogeneic T-lymphocytes. In allo-SCT donor T-lymphocytes favor engraftment, provide early immune reconstitution, and fight the underlying malignancy--the so-called graft-vs-leukemia (GVL) effect. These benefits are counterbalanced by the occurrence of a life-threatening disease: graft-vs-host disease (GVHD). A suicide gene encodes a protein able to convert a nontoxic prodrug into a toxic product. Therefore, cells expressing the suicide gene become selectively sensitive to the prodrug. The transfer of a suicide gene into donor lymphocytes could allow, upon administration of the prodrug, the in vivo selective elimination of transduced lymphocytes, resulting in the switch-off of GVHD, thus allowing full exploitation of the curative potential of donor T-lymphocytes in the context of allo-SCT. In this chapter the rationale, the materials, and the methods of the suicide-gene strategy with human T-lymphocytes are described.
Abstract: Malignant forms of multiple sclerosis (MS) represent a limited group of very aggressive demyelinating diseases, which rapidly progress to severe disability leading often to life-threatening conditions. On these clinical entities, currently available therapies for MS are not very effective. Recently, it has been demonstrated that intense immunosuppression followed by autologous stem cell transplantation (ASCT) can affect the clinical course of individuals with severe MS and completely abrogate the inflammatory activity detected by magnetic resonance imaging (MRI). We report on the treatment with intense immune ablation followed by ASCT of three patients with malignant MS whose clinical course indicated a dramatically poor prognosis. This procedure succeeded in halting the rapidly worsening course of disease. The effect was long lasting, as demonstrated by a sustained efficacy over a two-year period in two subjects and 12 months in the third case. In addition, a striking effect on inflammation-related MRI findings was obtained. These results support a role for intense immunosuppression followed by ASCT as treatment in rapidly evolving malignant MS cases unresponsive to conventional therapies.
Abstract: In allogeneic hematopoietic cell transplantation (allo-HCT), donor lymphocytes play a central therapeutic role in both GvL and immune reconstitution. However, the full exploitation of these therapeutic properties is limited by the occurrence of GvHD. Different strategies have been investigated to obtain all the benefits derived from donor lymphocytes while avoiding the risk of GvHD. The genetic engineering of donor lymphocytes with the herpes simplex virus-thymidine kinase (HSV-TK) suicide gene confers the ability to modulate GvHD by invivo ganciclovir-induced elimination of the transduced cells. The suicide-gene strategy has applications in both donor lymphocyte infusion (DLI) for disease relapse and in add-back infusions after T-cell depleted allo-HCT. TK cell DLI resulted in anti-tumor activity in a relevant proportion of treated patients. Haplo-identical stem cell transplantation (haplo-HCT) is a promising therapeutic option for patients with high risk hematologic malignancies lacking an HLA-matched donor. However, the profound T-cell depletion required to overcome the risk of lethal GvHD has been associated with a marked delayed T-cell recovery with a prolonged risk of post-transplant viral, fungal and other opportunistic infections. TK cell add-backs efficiently promote early immune reconstitution after haplo-HCT and prevent disease relapse, providing a unique tool for the control of GvHD. The genetic manipulation of donor lymphocytes with a suicide gene is a promising strategy to increase feasibility and safety of allo-HCT.
Abstract: BACKGROUND: The objective of this study was to identify prognostic factors for predicting survival in patients with advanced renal cell carcinoma (RCC) who had undergone an allogeneic stem cell transplantation after failure on immunotherapy. METHODS: The authors studied 70 patients with advanced RCC who underwent allogeneic transplantation with a fludarabine-based, reduced-intensity regimen. Ten parameters were analyzed at the time of transplantation for their power to predict survival. Clinical features were examined first univariately; then, variables that were correlated significantly with survival in the univariate analysis were included in a multivariate Cox regression model. RESULTS: Factors that were found to be associated significantly with limited survival were performance status, the number of metastatic sites, the presence of mediastinal metastasis, hemoglobin level, C-reactive protein (CRP) level, lactate dehydrogenase (LDH) level, and neutrophil counts. All these variables were included in a multivariate Cox regression model, and three were retained in the final model. Patients were classified according to the score estimated by the final Cox model in two groups (above or below the median value): The median survival was 3.5 months for patients who had a poor prognosis patients versus 23 months for patients who had a good prognosis. CONCLUSIONS: The current findings suggested that three easily available parameters (performance status, CRP level, and LDH level) could be used to stratify patients with advanced RCC who are candidates for allografting and to assist clinicians in decision-making and selection of an appropriate treatment program.
Abstract: PURPOSE: Older age and a previously failed autologous stem-cell transplantation (SCT) are poor prognostic factors for patients receiving myeloablative conditioning and allogeneic SCT. Reduced-intensity conditioning (RIC) regimens achieved a significant reduction of treatment-related mortality, but the influence of previously described risk factors on the outcome of this novel transplantation strategy have not been fully analyzed yet. PATIENTS AND METHODS: One hundred fifty patients with advanced hematologic malignancies received a RIC regimen containing thiotepa (10 mg/kg), fludarabine (60 mg/m2), and cyclophosphamide (60 mg/kg), followed by an allogeneic transplantation from an HLA-identical sibling donor. Patients were divided into two cohorts according to age; 90 patients were younger than 55 years, and 60 patients were 55 years old or older. The other pretransplantation characteristics were fairly balanced. RESULTS: Actuarial 5-year nonrelapse mortality (NRM) rate was not statistically different between the groups (13% in the younger group and 19% in the older group). By univariate and multivariate analysis, NRM was significantly higher in older patients who previously experienced failure with an autograft. The occurrence of grade 3 to 4 acute graft-versus-host disease (GVHD) or extensive chronic GVHD was associated with a higher NRM in both age cohorts. Overall survival (OS) was not statistically different between the younger (66%) and older groups (61%). By multivariate analysis, refractory disease was associated with a worse OS irrespective of age group. CONCLUSION: RIC transplantations show a rather low NRM, and age > or = 55 years per se cannot be considered a risk factor anymore. The timing of transplantation and novel strategies for the prevention of severe GVHD could further improve patient outcome.
Abstract: The importance of HLA-DPB1 matching for the outcome of allogeneic hematologic stem cell (HSC) transplantation is controversial. We have previously identified HLA-DPB1*0901 as a target of cytotoxic T cells mediating in vivo rejection of an HSC allograft. Here we show that HLA-DPB1*0901 encodes a T-cell epitope shared by a subset of DPB1 alleles that determines nonpermissive mismatches for HSC transplantation. Several T-cell clones obtained from the patient at the time of rejection showed HLA-DP restricted recognition of allogeneic targets expressing HLA-DPB1*0901, *1001, *1701, *0301, *1401, and *4501, but not other alleles. Based on these findings, we developed an algorithm for prediction of nonpermissive HLA-DPB1 mismatches. Retrospective evaluation of 118 transplantations showed that the presence of nonpermissive HLA-DPB1 mismatches was correlated with significantly increased hazards of acute grade II to IV graft-versus-host disease (HR = 1.87, P =.046) and transplantation-related mortality (HR = 2.69, P =.027) but not relapse (HR = 0.98, P =.939), as compared with the permissive group. There was also a marked but statistically not significant increase in the hazards of overall mortality (HR = 1.64, P =.1). These data suggest that biologic characterization of in vivo alloreactivity can be a tool for definition of clinically relevant nonpermissive HLA mismatches for unrelated HSC transplantation.
Abstract: OBJECTIVE: To analyse the durability of the responses after haematopoietic stem cell transplantation (HSCT) for severe systemic sclerosis (SSc) and determine whether the high transplant related mortality (TRM) improved with experience. This EBMT/EULAR report describes the longer outcome of patients originally described in addition to newly recruited cases. METHODS: Only patients with SSc, treated by HSCT in European phase I-II studies from 1996 up to 2002, with more than 6 months of follow up were included. Transplant regimens were according to the international consensus statements. Repeated evaluations analysed complete, partial, or non-response and the probability of disease progression and survival after HSCT (Kaplan-Meier). RESULTS: Given as median (range). Among 57 patients aged 40 (9.1-68.7) years the skin scores improved at 6 (n = 37 patients), 12 (n = 30), 24 (n = 19), and 36 (n = 10) months after HSCT (p<0.005). After 22.9 (4.5-81.1) months, partial (n = 32) or complete response (n = 14) was seen in 92% and non-response in 8% (n = 4) of 50 observed cases. 35% of the patients with initial partial (n = 13/32) or complete response (n = 3/14) relapsed within 10 (2.2-48.7) months after HSCT. The TRM was 8.7% (n = 5/57). Deaths related to progression accounted for 14% (n = 8/57) of the 23% (n = 13/57) total mortality rate. At 5 years, progression probability was 48% (95% CI 28 to 68) and the projected survival was 72% (95% CI 59 to 75). CONCLUSION: This EBMT/EULAR report showed that response in two thirds of the patients after HSCT was durable with an acceptable TRM. Based on these results prospective, randomised trials are proceeding.
Abstract: Allogeneic stem cell transplantation has emerged as a potentially curative treatment modality for patients with hematological malignancies. The graft-versus-leukemia effect, an immune mechanism mediated by the donor immune system, is an important component of the therapeutic effect of allogeneic transplantation. Recent data from experimental animal models and from preliminary clinical experience suggest that a graft-versus-tumor effect, analogous to the graft-versus-leukemia effect, may be generated against solid tumors such as renal cell cancer, breast cancer, and other malignancies. The use of non-myeloablative, immunosuppressive conditioning regimens offers the opportunity to achieve a full-donor engraftment with reduced transplant-related complications and mortality, enabling also patients of advanced age and with co-morbidities to receive an allografting. Advanced renal cell cancer, an essentially incurable disease, has emerged from pilot studies as a disease susceptible to the graft-versus-tumor effect. Future studies will demonstrate if the tumor responses observed after allografting will translate into a clinically meaningful survival advantage. Other tumors in which tumor responses have been observed are: breast cancer, ovarian cancer, colorectal cancer, soft-tissue sarcoma, and others. Advanced melanoma may not be amenable to graft-versus-tumor effect. Future studies will point to the identification, isolation and cloning of target antigen(s) of graft-versus-tumor effect, to further reduce toxicities and to achieve a selective cell-mediated immunotherapy.
Abstract: A 27-year-old man with the autoimmune lymphoproliferative syndrome and a large-B-cell lymphoma had heterozygous mutations in the Fas and perforin (Prf1) genes. The Fas mutation was inherited from his healthy father and was also carried by his healthy brother, whereas the Prf1 mutation was inherited from his healthy mother. The combined effect of the two mutant genes may have contributed to the development of the autoimmune lymphoproliferative syndrome and lymphoma in this patient.
Abstract: BACKGROUND AND OBJECTIVES: Tumor necrosis factor a is one of the principal cytokines involved in the pathogenesis of acute graft-versus-host-disease (GVHD). Infliximab is an antibody to this cytokine. DESIGN AND METHODS: We performed a retrospective analysis to evaluate the activity of infliximab in 32 patients with severe steroid-refractory acute GVHD. The patients received a median of 3 weekly courses of infliximab. The main organs involved in the patients were skin (n=2) liver (n=1), bowel (n=19), liver and bowel at the same stage (n=10). RESULTS: Nineteen out 32 patients (59%) responded to infliximab with 6 (19%) complete and 13 (40%) partial responses. Age younger than 35 years, intestinal involvement and a longer time between hematopoietic stem cell transplantation and infliximab administration were factors predicting a favorable response. Infective episodes developed in 23/32 (72%) patients. All the 13 unresponsive patients died of GVHD shortly after infliximab. Thirteen of 19 responsive patients were alive at a median follow-up of 449 days (range 155-842) after infliximab, with no signs of chronic GVHD (n=5), limited (n=5) or extensive involvement (n=3). Six patients who responded subsequently died, one of chronic lung GVHD, the others of vascular complications or infections (2 fungal diseases). INTERPRETATION AND CONCLUSIONS: We conclude that infliximab is active in the treatment of severe steroid-refractory acute GVHD, particularly when the intestine is involved. Infections commonly followed its administration. The clinical activity of infliximab and the possibility that it increases the risk of infections are worth investigating in prospective trials.
Abstract: We have previously shown that the infusion of donor lymphocytes expressing the herpes simplex virus thymidine kinase (HSV-tk) gene is an efficient tool for controlling graft-versus-host disease (GVHD) while preserving the graft-versus-leukemia (GVL) effect. In addition to the GVL effect, the administration of donor HSV-tk(+) cells could have a clinical impact in promoting immune reconstitution after T-cell-depleted stem cell transplantation (SCT). To explore this hypothesis, we have investigated whether in vitro polyclonal activation, retroviral transduction, immunoselection, and expansion affect the immune competence of donor T cells. We have observed that, after appropriate in vitro manipulation, T cells specific for antigens relevant in the context of SCT are preserved in terms of frequency, expression of T-cell receptor, proliferation, cytokine secretion, and lytic activity. A reduction in the frequency of allospecific T-cell precursors is observed after prolonged T-cell culture, suggesting that cell manipulation protocols involving a short culture time and high transduction efficiency are needed. Finally, the long-term persistence of HSV-tk(+) cells was observed in a patient treated in the GVL clinical trial, and a reversion of the phenotype of HSV-tk(+) cells from CD45RO(+) to CD45RA(+) was documented more than 2 years after the infusion. Based on all this evidence, we propose a clinical study of preemptive infusions of donor HSV-tk(+) T cells after SCT from haploidentical donors to provide early immune reconstitution against infection and potential immune protection against disease recurrence.
Abstract: A reduced-intensity conditioning regimen was investigated in 45 patients with hematologic malignancies who were considered poor candidates for conventional myeloablative regimens. Median patient age was 49 years. Twenty-six patients previously failed autologous transplantation, and 18 patients had a refractory disease at the time of transplantation. In order to decrease nonrelapse mortality, and enhance the graft-versus-tumor effect, a program was designed in which a reduced conditioning with thiotepa, fludarabine, and cyclophosphamide was associated with programmed reinfusions of donor lymphocytes for patients without graft-versus-host disease (GVHD), not achieving clinical and molecular remission after transplantation. GVHD prophylaxis consisted of cyclosporine A and methotrexate. Seventeen patients received marrow cells and 28 received mobilized hematopoietic cells. All patients engrafted. The probability of grades II-IV and III-IV acute GVHD were 47% and 13%, respectively. The probability of nonrelapse mortality, progression-free survival, and overall survival were 13%, 57%, and 53%, respectively. Thirteen patients in complete remission had a polymerase chain reaction marker for minimal disease monitoring; 10 achieved molecular remission after transplantation. Nine patients received donor lymphocytes: one patient with mantle cell lymphoma had a minimal response, one patient with refractory anemia with excess of blasts in transformation achieved complete remission, and 7 patients did not respond. At a median follow-up of 385 days (range, 24 to 820 days), 25 patients (55%) were alive in complete remission. Although longer follow-up is needed to evaluate the long-term outcome, the study shows that this regimen is associated with a durable engraftment, has a low nonrelapse mortality rate, and can induce clinical and molecular remissions.
Abstract: In allogeneic hematopoietic cell transplantation, donor lymphocytes play a central therapeutic role in both graft-versus-leukemia and immune reconstitution. However, their use is limited by the risk of severe graft-versus-host disease (GVHD). Different strategies have been investigated to obtain all the benefits derived from donor lymphocytes while avoiding the risk of GVHD. Infusions of donor lymphocytes transduced with the herpes simplex virus thymidine kinase (HSV-tk) suicide gene resulted in anti-tumor activity in a substantial number of patients. Acute GVHD could be effectively controlled by ganciclovir-induced elimination of the transduced cells. Haplo-identical stem cell transplantation (haplo-SCT) is a promising therapeutic option for patients with high-risk hematologic malignancies lacking an HLA-matched donor. However, the intensive T-cell depletion required to overcome the risk of lethal GVHD has been associated with a delayed immune recovery with a prolonged risk of posttransplantation viral, fungal, and other opportunistic infections. Donor lymphocyte infusions of HSV-tk represent a promising tool for preventing disease relapse and promoting immune reconstitution after haplo-SCT, and a unique tool for the control of GVHD. The genetic manipulation of donor lymphocytes with a suicide gene is a promising strategy to increase feasibility and safety of allogeneic bone marrow transplantation.
Abstract: BACKGROUND: Primary central nervous system lymphomas (PCNSLs) are rare tumors, mostly represented by diffuse large B cells. PCNSLs with a T phenotype are less frequently reported; even rarer are anaplastic large cell lymphomas (ALCLs). PCNSL ALCLs are commonly represented, like their systemic counterpart, by a variably prevalent amount of large pleomorphic tumor cells ('hallmark cells'), and this feature enhances their recognition. Patient and methods We report the first case of primary brain CD30+ ALK-1+ ALCL with a T-cell phenotype, showing the combination of both the 'lymphohistiocytic' and the 'small cell' variants of the disease. A few elements consistent with 'hallmark cells' were recognizable. However, these cells were never prominent, increasing diagnostic difficulties. Immunohistochemistry results were critical for the correct interpretation. Our findings also differ from the majority of PCNSL ALCLs for the absence of tumor necrosis and the lack of prominent mitotic activity. The neuroimaging picture was not specific. A comparison with literature data concerning the clinical/instrumental features shows a very frequent meningeal involvement in PCNSL ALCLs, in contrast to the majority of PCNSLs. CONCLUSION: The occurrence of such a rare form of ALCL may widen the spectrum of differential diagnoses in PCNSL and their recognition may allow a rapid diagnosis, thus encouraging adequate treatment, which should take into account the high rate of meningeal involvement observed in these cases.
Abstract: OBJECTIVES: To assess the feasibility and the activity, as well as the efficacy to treat meninges, of chemotherapy (CHT) containing high-dose methotrexate (HD-MTX) followed by radiation therapy (RT), without intrathecal CHT, in patients with primary central nervous system lymphoma. Methods: Eligibility criteria were histologically proven diagnosis, disease limited to the CNS, age < or = 70, ECOG performance status < or = 3, HIV-negative and no prior treatment. Thirteen patients (1996-1999; median age 54 years) received two courses of vincristine 1.4 mg/m2 day 1, MTX 3 g/m2 days 3 and 10 and procarbazine 100 mg/m2 days 1-14 every 4 weeks. Patients who achieved a complete remission were referred to RT, those with progressive disease were excluded from further study; all the remaining patients received a third course of CHT followed by RT. RESULTS: Twelve patients responded to CHT (overall response rate = 92%, complete response rate = 77%): 9 underwent consolidation RT, 3 did not. Two patients experienced severe acute toxicity; lethal pulmonary thromboembolism and transient renal failure. Five patients relapsed: 2 after CHT and 3 after RT. Relapse was local in all cases, with a case of concomitant hepatic involvement. No cases of ocular or meningeal relapse were observed. In contrast to high-dose cytarabine-containing CHT, salvage therapy with temozolomide produced good results. Two patients died of treatment-related neurotoxicity. Six patients are alive with a median follow-up of 17 months, and a 2-year overall survival (OS) of 61%. The median survival of the 9 patients who completed the planned treatment is 25+ months with a 2-year OS of 80%. CONCLUSIONS: HD-MTX, procarbazine and vincristine followed by RT, without intrathecal therapy, produce similar results with respect to other HD-MTX-containing regimens. These results seem to suggest that adequate meningeal treatment is possible without intrathecal drug delivery, even in CSF-positive patients. Corroborating data from a larger series are, however, necessary. Temozolomide should be tested in relapsed patients in a phase II prospective trial.
Abstract: A 79-year-old patient with post-polycythaemic myelofibrosis presented with severe hypersplenism. After splenic artery catheterization, cytosine arabinoside was given intrasplenically from November 1999 to March 2000 for 5 d/month at 10 mg/m2 and increased each month by 10 mg/m2. It was then administered by continuous infusion until June 2000, starting at 20 mg/m2/d and tapering by 5 mg/m2 every 2 weeks to a final daily dose of 5 mg/m2/d. The drug was then stopped. The spleen had decreased to one third of the initial volume. Clinical conditions and haematological indices improved substantially. Intrasplenic therapy could be a new therapeutic tool for hypersplenism in chronic idiopathic and post-myeloproliferative myelofibrosis.
Abstract: In allogeneic marrow transplantation (BMT), donor lymphocytes play a central therapeutic role in both graft-versus-leukemia and immune reconstitution. However, their use is limited by the risk of severe graft-vs-host disease (GvHD). Different strategies have been investigated to obtain all the benefits derived from donor lymphocytes while avoiding the risk of GvHD. In the first pilot study, infusions of donor lymphocytes transduced with the Herpes Simplex virus thymidine kinase (HSV-tk) suicide resulted in anti-tumor activity in 50% of patients. Acute GvHD could be effectively controlled by ganciclovir-induced elimination of the transduced cells. If these results will be confirmed in larger studies, genetic manipulation of donor lymphocytes will increase efficacy and safety of allogeneic marrow transplantation.
Abstract: We previously demonstrated that severe graft-versus-host disease (GvHD), associated with the therapeutic infusion of donor lymphocytes after allogeneic marrow transplantation (BMT), can be efficiently controlled by the SFCMM-2-mediated expression of the herpes simplex virus thymidine kinase (HSV-tk) suicide gene into the allogeneic lymphocytes. This was achieved by selective elimination of transduced lymphocytes by ganciclovir (GCV) infusion. Despite the positive results of the pilot clinical trial, two vector-related limitations were observed. The induction of a strong immune response against genetically modified cells was observed in two patients. In addition, the only patient who developed chronic GvHD showed only partial response to ganciclovir treatment. In an attempt to overcome these limitations, we developed a new generation of vectors. The neomycin resistance (neoR) gene was removed from the SFCMM-3 and SFCMM-4 retroviral vectors. These vectors are less immunogenic and able to confer higher ganciclovir sensitivity to transduced human lymphocytes. All the vectors carry a modified form of the low-affinity nerve growth factor receptor cDNA, as cell surface selectable marker (deltaLNGFR). The vectors were compared in terms of gene transfer efficiency, and ability to confer high and specific sensitivity to ganciclovir. The SFCMM-3 vector, carrying the entire HSV-tk gene driven by the LTR promoter, allows efficient transduction of human T lymphocytes and confers the highest GCV sensitivity to transduced lymphocytes with a high and a low proliferation index. The expression of the deltaLNGFR marker allows an easier in vitro manipulation and a faster immune selection of transduced cells compared with neoR selection. Finally, the elimination of the neoR gene removes a potent immunogen from transduced lymphocytes.
