Larissa Verda

Autologous nonmyeloablative hematopoietic stem cell transplantation in patients with severe anti-TNF refractory Crohn disease: long-term follow-up

We evaluated the safety and clinical outcome of autologous nonmyeloablative hematopoietic stem cell transplantation (HSCT) in patients with severe Crohn disease (CD) defined as a Crohn Disease Activity Index (CDAI) greater than 250, and/or Crohn Severity Index greater than 16 despite anti-tumor necrosis factor therapy. Stem cells were mobilized from the peripheral blood using cyclophosphamide (2.0 g/m(2)) and G-CSF (10 μg/kg/day), enriched ex vivo by CD34(+) selection, and reinfused after immune suppressive conditioning with cyclophosphamide (200 mg/kg) and either equine antithymocyte globulin (ATG, 90 mg/kg) or rabbit ATG (6 mg/kg). Eighteen of 24 patients are 5 or more years after transplantation. All patients went into remission with a CDAI less than 150. The percentage of clinical relapse-free survival defined as the percent free of restarting CD medical therapy after transplantation is 91% at 1 year, 63% at 2 years, 57% at 3 years, 39% at 4 years, and 19% at 5 years. The percentage of patients in remission (CDAI < 150), steroid-free, or medication-free at any posttransplantation evaluation interval more than 5 years after transplantation has remained at or greater than 70%, 80%, and 60%, respectively. This trial was registered at www.clinicaltrials.gov as NCT0027853.

Embryonic stem cells as an alternate marrow donor source : engraftment without graft-versus-host disease

A single embryonic stem cell (ESC) line can be repetitively cryopreserved, thawed, expanded, and differentiated into various cellular components serving as a potentially renewable and well-characterized stem cell source. Therefore, we determined whether ESCs could be used to reconstitute marrow and blood in major histocompatibility complex (MHC)-mismatched mice. To induce differentiation toward hematopoietic stem cells (HSCs) in vitro, ESCs were cultured in methylcellulose with stem cell factor, interleukin (IL)-3, and IL-6. ESC-derived, cytokine-induced HSCs (c-kit+/CD45+) were isolated by flow cytometry and injected either intra bone marrow or intravenously into lethally irradiated MHC-mismatched recipient mice. From 2 wk to 6 mo after injection, the peripheral blood demonstrated increasing ESC-derived mononuclear cells that included donor-derived T and B lymphocytes, monocytes, and granulocytes without clinical or histologic evidence of graft-versus-host disease (GVHD). Mixed lymphocyte culture assays demonstrated T cell tolerance to both recipient and donor but intact third party proliferative responses and interferon γ production. ESCs might be used as a renewable alternate marrow donor source that reconstitutes hematopoiesis with intact immune responsiveness without GVHD despite crossing MHC barriers.

Mobilization, harvesting and selection of peripheral blood stem cells in patients with autoimmune diseases undergoing autologous hematopoietic stem cell transplantation

Peripheral blood stem cells (PBSC) were mobilized in 130 patients with autoimmune diseases undergoing autologous hematopoietic stem cell transplantation using cyclophosphamide 2 g/m2 and either granulocyte colony-stimulating factor (G-CSF) 5 mcg/kg/day (for systemic lupus erythematosus (SLE) and secondary progressive multiple sclerosis, SPMS) or G-CSF 10 mcg/kg/day (for relapsing remitting multiple sclerosis (RRMS), Crohns disease (CD), systemic sclerosis (SSc), and other immune-mediated disorders). Mobilization-related mortality was 0.8% (one of 130) secondary to infection. Circulating peripheral blood (PB) CD34+ cells/μl differed significantly by disease. Collected CD34+ cells/kg/apheresis and overall collection efficiency was significantly better using Spectra apheresis device compared to the Fenwall CS3000 instrument. Patients with SLE and RRMS achieved the lowest and the highest CD34+ cell yields, respectively. Ex vivo CD34+ cell selection employing Isolex 300iv2.5 apparatus was significantly more efficient compared to CEPRATE CS device. Circulating PB CD34+ cells/μl correlated positively with initial CD34+ cells/kg/apheresis and enriched product CD34+ cells/kg. Mean WBC and platelet engraftment (ANC>0.5 × 109/l and platelet count >20 × 109/l) occurred on days 9 and 11, respectively. Infused CD34+ cell/kg dose showed significant direct correlation with faster white blood cell (WBC) and platelet engraftment. When adjusted for CD34+ cell/kg dose, patients treated with a myeloablative regimen had significantly slower WBC and platelet recovery compared to non-myeloablative regimens.

Hematopoietic Mixed Chimerism Derived from Allogeneic Embryonic Stem Cells Prevents Autoimmune Diabetes Mellitus in NOD Mice

Embryonic stem cell (ESC)‐derived hematopoietic stem cells (HSC), unlike HSC harvested from the blood or marrow, are not contaminated by lymphocytes. We therefore evaluated whether ESC‐derived HSC could produce islet cell tolerance, a phenomenon termed graft versus autoimmunity (GVA), without causing the usual allogeneic hematopoietic stem cell transplant complication, graft‐versus‐host disease (GVHD). Herein, we demonstrate that ESC‐derived HSC may be used to prevent autoimmune diabetes mellitus in NOD mice without GVHD or other adverse side effects. ESC were cultured in vitro to induce differentiation toward HSC, selected for c‐kit expression, and injected either i.v. or intra‐bone marrow (IBM) into sublethally irradiated NOD/LtJ mice. Nine of 10 mice from the IBM group and 5 of 8 from the i.v. group did not become hyperglycemic, in contrast to the control group, in which 8 of 9 mice developed end‐stage diabetes. All mice with >5% donor chimerism remained free of diabetes and insulitis, which was confirmed by histology. Splenocytes from transplanted mice were unresponsive to glutamic acid decarboxylase isoform 65, a diabetic‐specific autoantigen, but responded normally to third‐party antigens. ESC‐derived HSC can induce an islet cell tolerizing GVA effect without GVHD. This study represents the first instance, to our knowledge, of ESC‐derived HSC cells treating disease in an animal model.

Autologous non-myeloablative haematopoietic stem cell transplantation for refractory systemic vasculitis.

Objective: For patients with systemic vasculitis (SV) refractory to conventional therapy, new treatment strategies aimed at aggressive induction of remission and relapse prevention are being sought. We herein report our single-centre experience in treating four patients with refractory SV employing non-myeloablative autologous haematopoietic stem cell transplantation (HSCT). Methods: Four patients with refractory SV (two with neurovascular Behcet disease, one with neurovascular Sjögren syndrome, and one with Wegener granulomatosis) were involved in an Institutional Review Board (IRB) and US Food and Drug Administration (FDA) approved phase I clinical trial of high dose chemotherapy and autologous HSCT. Peripheral blood stem cells were mobilised with cyclophosphamide (Cy) and granulocyte-colony stimulating factor (G-CSF). Conditioning regimen consisted of Cy 200 mg/kg and rabbit anti-thymocyte globulin 5.5 mg/kg intravenously (iv). Results: All four patients tolerated HSCT well without transplant related mortality or any significant toxicity. At median follow-up of 28 (range 22–36) months all patients were alive. Three patients (one with Behcet disease, one with Sjögren syndrome, and one with Wegener granulomatosis) entered a sustained remission at 6, 6 and 24 months, respectively, after transplant. They had significant decrease in disease activity and disease or treatment related damage, as measured by the Birmingham Vasculitis Activity Score and Vasculitis Damage Index, respectively. All three patients who achieved remission discontinued immunosuppressive therapy at the time of transplant and have not required treatment since. One patient with Behcet disease and positive for human leukocyte antigen (HLA)-B51 has not improved after HSCT. Conclusion: We suggest non-myeloablative autologous HSCT is an alternative therapy for select patients with SV refractory to conventional immunosuppressive therapies.

Non-myeloablative stem cell transplantation for autoimmune diseases

Treatment of life-threatening autoimmune diseases in animal models with induced or spontaneous autoimmune diseases can be accomplished by a 2-step procedure involving elimination of self-reactive lymphocytes with an immune ablative conditioning regimen followed by infusion of autologous or allogeneic stem cells, respectively. In animal models it was shown that using such a strategy, autoimmunity could be adequately controlled. It is speculated that de-novo development of the T and B cell repertoire from uncommitted progenitor cells in the presence of the autoantigens may be the best recipe for re-induction of self-tolerance, similarly to the normal ontogeny of the immune system during the induction of self tolerance in fetal stage. For both autologous and allogeneic hematopoietic stem cell transplantation, a non-myeloablative stem cell transplantation (NST) regimen may be used for safer lymphoablation rather than myeloablation. In addition, for allogeneic hematopoietic stem cell transplantation engraftment of disease resistant donor stem cells will alter the genetic predisposition towards autoimmune disease susceptibility.

Non-myeloablative allogeneic hematopoietic stem cell transplantation for severe systemic sclerosis: graft-versus-autoimmunity without graft-versus-host disease?

Non-myeloablative allogeneic hematopoietic stem cell transplantation for severe systemic sclerosis: graft-versus-autoimmunity without graft-versus-host disease?

Mitotically inactivated embryonic stem cells can be used as an in vivo feeder layer to nurse damaged myocardium after acute myocardial infarction: a preclinical study.

Rationale: Various types of viable stem cells have been reported to result in modest improvement in cardiac function after acute myocardial infarction. The mechanisms for improvement from different stem cell populations remain unknown. Objective: To determine whether irradiated (nonviable) embryonic stem cells (iESCs) improve postischemic cardiac function without adverse consequences. Methods and Results: After coronary artery ligation-induced cardiac infarction, either conditioned media or male murine or male human iESCs were injected into the penumbra of ischemic myocardial tissue of female mice or female rhesus macaque monkeys, respectively. Murine and human iESCs, despite irradiation doses that prevented proliferation and induced cell death, significantly improved cardiac function and decreased infarct size compared with untreated or media-treated controls. Fluorescent in situ hybridization of the Y chromosome revealed disappearance of iESCs within the myocardium, whereas 5-bromo-2′-deoxyuridine assays revealed de novo in vivo cardiomyocyte DNA synthesis. Microarray gene expression profiling demonstrated an early increase in metabolism, DNA proliferation, and chromatin remodeling pathways, and a decrease in fibrosis and inflammatory gene expression compared with media-treated controls. Conclusions: As a result of irradiation before injection, ex vivo and in vivo iESC existence is transient, yet iESCs provide a significant improvement in cardiac function after acute myocardial infarction. The mechanism(s) of action of iESCs seems to be related to cell-cell exchange, paracrine factors, and a scaffolding effect between iESCs and neighboring host cardiomyocytes.

Effect of hematopoietic growth factors on severity of experimental autoimmune encephalomyelitis

We have investigated the influence of different hematopoietic growth factors, including granulocyte colony-stimulating factor (G-CSF), stem cell factor (SCF), Flt-3 ligand (Flt-3L) and thrombopoietin (TPO), on the course of relapsing experimental autoimmune encephalomyelitis, a mouse model of multiple sclerosis. Disease course and central nervous system histology were evaluated in all groups. When given after immunization but before either disease onset or during remission, Flt-3L, SCF and G-CSF exacerbated disease severity whereas TPO had no effect compared to non-cytokine-treated controls. When compared to controls, TPO did not exacerbate disease. We conclude that autoimmune disease severity may be affected by hematopoietic growth factors currently being employed in hematopoietic stem cell transplantation of patients with autoimmune disease. The mechanism of their effects remains unknown: it may be related to both T helper (Th) 1/Th2 skewing and/or homing of inflammatory cells to the disease-affected organ.

Unique clinical aspects of hematopoietic stem cell transplantation for autoimmune diseases

Purpose of reviewHematopoietic stem cell transplantation is increasingly used for treatment of severe autoimmune diseases. However, multicenter registry data have demonstrated higher than anticipated toxicity (approximately 10%), which, if uncorrected, will hinder development of this field. Recent findingsAt Northwestern University, the authors began performing autologous hematopoietic stem cell transplantation for autoimmune disease 6 years ago and have had no treatment or infectious mortality in any of the 90 patients receiving a stem cell transplant. The article highlights the rationale for conditioning regimen selection and some of the unique circumstances in hematopoietic stem cell transplantation for three autoimmune diseases: systemic lupus erythematosus, multiple sclerosis, and systemic sclerosis (scleroderma). SummaryExperience, a dedicated team approach, and understanding the rationale and selection for the conditioning regimen and unique aspects of each autoimmune disease should substantially improve the morbidity and mortality of this procedure.