David S. Allan

Differential genomic targeting of the transcription factor TAL1 in alternate haematopoietic lineages

TAL1/SCL is a master regulator of haematopoiesis whose expression promotes opposite outcomes depending on the cell type: differentiation in the erythroid lineage or oncogenesis in the T‐cell lineage. Here, we used a combination of ChIP sequencing and gene expression profiling to compare the function of TAL1 in normal erythroid and leukaemic T cells. Analysis of the genome‐wide binding properties of TAL1 in these two haematopoietic lineages revealed new insight into the mechanism by which transcription factors select their binding sites in alternate lineages. Our study shows limited overlap in the TAL1‐binding profile between the two cell types with an unexpected preference for ETS and RUNX motifs adjacent to E‐boxes in the T‐cell lineage. Furthermore, we show that TAL1 interacts with RUNX1 and ETS1, and that these transcription factors are critically required for TAL1 binding to genes that modulate T‐cell differentiation. Thus, our findings highlight a critical role of the cellular environment in modulating transcription factor binding, and provide insight into the mechanism by which TAL1 inhibits differentiation leading to oncogenesis in the T‐cell lineage.

The transcription factor E4bp4/Nfil3 controls commitment to the NK lineage and directly regulates Eomes and Id2 expression

E4bp4 is required for commitment to the NK lineage and promotes NK development by directly regulating the expression of Eomes and Id2.

Immunoablation and autologous haemopoietic stem-cell transplantation for aggressive multiple sclerosis: a multicentre single-group phase 2 trial.

BACKGROUND Strong immunosuppression, including chemotherapy and immune-depleting antibodies followed by autologous haemopoietic stem-cell transplantation (aHSCT), has been used to treat patients with multiple sclerosis, improving control of relapsing disease. We addressed whether near-complete immunoablation followed by immune cell depleted aHSCT would result in long-term control of multiple sclerosis. METHODS We did this phase 2 single-arm trial at three hospitals in Canada. We enrolled patients with multiple sclerosis, aged 18-50 years with poor prognosis, ongoing disease activity, and an Expanded Disability Status Scale of 3.0-6.0. Autologous CD34 selected haemopoietic stem-cell grafts were collected after mobilisation with cyclophosphamide and filgrastim. Immunoablation with busulfan, cyclophosphamide, and rabbit anti-thymocyte globulin was followed by aHSCT. The primary outcome was multiple sclerosis activity-free survival (events were clinical relapse, appearance of a new or Gd-enhancing lesion on MRI, and sustained progression of Expanded Disability Status Scale score). This study was registered at ClinicalTrials.gov, NCT01099930. FINDINGS Between diagnosis and aHSCT, 24 patients had 167 clinical relapses over 140 patient-years with 188 Gd-enhancing lesions on 48 pre-aHSCT MRI scans. Median follow-up was 6.7 years (range 3.9-12.7). The primary outcome, multiple sclerosis activity-free survival at 3 years after transplantation was 69.6% (95% CI 46.6-84.2). With up to 13 years of follow-up after aHSCT, no relapses occurred and no Gd enhancing lesions or new T2 lesions were seen on 314 MRI sequential scans. The rate of brain atrophy decreased to that expected for healthy controls. One of 24 patients died of transplantation-related complications. 35% of patients had a sustained improvement in their Expanded Disability Status Scale score. INTERPRETATION We describe the first treatment to fully halt all detectable CNS inflammatory activity in patients with multiple sclerosis for a prolonged period in the absence of any ongoing disease-modifying drugs. Furthermore, many of the patients had substantial recovery of neurological function despite their diseases aggressive nature. FUNDING Multiple Sclerosis Scientific Research Foundation.

Human Endothelial Colony-Forming Cells Protect against Acute Kidney Injury: Role of Exosomes

The administration of certain progenitor cells is protective in experimental acute kidney injury (AKI), and mechanisms may involve the release of paracrine factors. Endothelial colony-forming cells (ECFCs) are endothelial precursor cells with a high proliferative capacity and pro-angiogenic potential. We examined the effects of human umbilical cord blood-derived ECFCs and their extracellular vesicles in a mouse model of ischemic AKI and in cultured human umbilical vein endothelial cells subjected to hypoxia/reoxygenation. In mice with ischemic AKI, administration of ECFCs (i.v.) at the time of reperfusion significantly attenuated increases in plasma creatinine, tubular necrosis, macrophage infiltration, oxidative stress, and apoptosis, without cell persistence in the kidneys. In cultured human umbilical vein endothelial cells, hypoxia/reoxygenation stimulated apoptosis. This effect was inhibited by incubation with conditioned medium or exosomes (40- to 100-nm diameter) derived from ECFCs, but not by microparticles (100- to 1000-nm diameter) or vesicle-depleted conditioned medium. Administration of exosomes (i.v.) directly to mice with ischemic AKI attenuated renal injury, as assessed by plasma creatinine, tubular necrosis, and apoptosis. Taken together, these studies indicate protective effects of human cord blood-derived ECFCs in experimental AKI and suggest that ECFC-derived exosomes may mediate the protective response via inhibition of endothelial cell apoptosis.

Incidence of symptomatic venous thromboembolism following hematopoietic stem cell transplantation

Summary.  Background: The incidence of symptomatic venous thromboembolism (VTE) following hematopoietic stem cell transplantation (HSCT) is not well described, particularly with increased use of ambulatory care in the transplant setting. Methods: A retrospective analysis involving 589 patients (382 autologous HSCT, 207 allogeneic HSCT) undergoing transplantation between 2000 and 2005 in a single Canadian institution was undertaken to identify the incidence of proximal deep vein thrombosis (DVT) or pulmonary embolism (PE) in HSCT patients. Results: The total 1‐year incidence of symptomatic VTE was 3.7% [95% confidence interval (CI) 2.5–5.6]. Among the HSCT patients, 7/589 (1.2%, 95% CI 0.6–2.4) developed symptomatic non‐catheter‐related VTE following HSCT (four PE and three DVT). All VTE events occurred after hematopoietic engraftment. Patients undergoing autologous HSCT did not receive thromboprophylaxis, whereas most patients undergoing allogeneic HSCT (79.7%) received enoxaparin 20 mg daily for the prevention of veno‐occlusive disease of the liver, starting 6 ± 3 days before transplantation for a mean of 22 ± 14 days. Conclusion: HSCT patients have a high incidence of VTE. Thromboprophylaxis should potentially be considered in these patients. However, future studies assessing the risk and benefits of thromboprophylaxis are needed in this specific population.

Utility of Comorbidity Assessment in Predicting Transplantation-Related Toxicity Following Autologous Hematopoietic Stem Cell Transplantation for Multiple Myeloma

Patients with coexisting medical problems may suffer increased toxicity and reduced quality of life after autologous hematopoietic stem cell transplantation (HSCT). The benefit of high-dose therapy for some patients with multiple myeloma (MM) is debatable. Decision tools that aid in identifying those patients with MM most suited for autologous HSCT may avoid the risk of excess toxicity. An objective assessment of comorbidities was performed in 126 patients with MM undergoing autologous HSCT using the Charlson comorbidity index (CCI), the hematopoietic cell transplantation comorbidity index (HCT-CI), and a modified pretransplantation assessment of mortality (mPAM) to determine the strength of association with increased transplantation-related toxicity and increased length of hospital stay (LOS). Any comorbidity scored using the CCI or HCT-CI (score > 0) was associated with an increased number of organ systems with serious toxicity (at least grade 2 toxicity using the Seattle criteria), an increased total sum of toxicity grades for all organs, and prolonged LOS. An mPAM score > or = 24 was associated with increased LOS. When considering autologous HSCT for a patient with MM, assessment of comorbidities using the CCI or HCT-CI may assist in predicting the risk of transplantation-related toxicity as an adjunct to physician judgment and patient preference.

Mesenchymal stromal cells from patients with acute myeloid leukemia have altered capacity to expand differentiated hematopoietic progenitors.

The bone marrow microenvironment may be permissive to the emergence and progression of acute myeloid leukemia (AML). Studying interactions between the microenvironment and leukemia cells should provide new insight for therapeutic advances. Mesenchymal stromal cells (MSCs) are central to the maintenance of the hematopoietic niche. Here we compared the functions and gene expression patterns of MSCs derived from bone marrow aspirates of healthy donors and patients with AML. MSCs expanded from AML patients had heterogeneous morphology and displayed a wide range of proliferation capacity compared to MSCs from healthy controls. The ability of AML-MSCs to support the expansion of committed hematopoietic progenitors from umbilical cord blood-derived CD34+ cells may be impaired while the expression of genes associated with maintaining hematopoietic quiescence appeared to be increased in AML-MSCs compared to healthy donors. These results highlight important potential differences in the biologic profile of MSCs from AML patients compared to healthy donors that may contribute to the emergence or progression of leukemia.

Transplantation of Umbilical Cord Blood–Derived Cells for Novel Indications in Regenerative Therapy or Immune Modulation: A Scoping Review of Clinical Studies

Although used mainly for transplantation of hematopoietic stem cells in the treatment of blood disorders, umbilical cord blood (UCB)-based therapies are now being used increasingly for novel applications in nonhematopoietic diseases and as a form of cellular regenerative therapy or immune modulation. We performed a systematic scoping review by searching Medline, EMBASE, and the Cochrane Library for published articles, and we searched www.clinicaltrials.com and the World Health Organization International Clinical Trials Registry Platform to describe the breadth of published studies and ongoing clinical activity in umbilical cord-based cellular therapy for regenerative therapy and immune modulation. The most commonly published area of expertise in the use of UCB-derived cellular transplantation for novel indications is for neurological disorders and this remains the most active area of study in ongoing registered trials. An increasingly broad range of disorders, however, are reflected in ongoing registered trials, which suggests greater activity, interest, and investment in UCB-derived cellular therapy. Interestingly, adult patients compose the majority of patients reported in published reports and registered ongoing clinical studies continue to enroll predominantly adult subjects. Geographically, Asian countries appear most active in UCB-derived cellular therapy and our analysis of ongoing studies suggests this trend will likely continue. Regular assessment of published and ongoing activity in UCB transplantation for emerging novel indications will be critical for informing UCB banking establishments and funding agencies to guide changes in banking practices related to emerging trends in cell therapy.

C-Linked Antifreeze Glycoprotein (C-AFGP) Analogues as Novel Cryoprotectants

Significant cell damage occurs during cryopreservation resulting in a decreased number of viable and functional cells post-thawing. Recent studies have correlated the unsuccessful outcome of regenerative therapies with poor cell viability after cryopreservation. Cell damage from ice recrystallization during freeze-thawing is one cause of decreased viability after cryopreservation. We have assessed the ability of two C-AFGPs that are potent inhibitors of ice recrystallization to increase cell viability after cryopreservation. Our results indicate that a 1-1.5 mg/mL (0.5-0.8 mM) solution of C-AFGP 1 is an excellent alternative to a 2.5% DMSO solution for the cryopreservation of human embryonic liver cells.

Contaminating tumour cells in autologous PBSC grafts do not influence survival or relapse following transplant for multiple myeloma or B-cell non-Hodgkin's lymphoma

Relapsed disease remains a major obstacle following autologous haematopoietic SCT (HSCT) for non-Hodgkins lymphoma (NHL) and multiple myeloma (MM). Studies regarding the importance of detectable tumour cells in PBSC collections have been inconclusive. Patients undergoing autologous HSCT for NHL and MM between 2001 and 2006 were enrolled (n=158). PBSC grafts were assessed for clonal IgH CDR3 gene rearrangements using qualitative semi-nested PCR. In comparison to patients with PCR-positive PBSC grafts, patients negative for detectable disease had no improvement in overall survival (OS) or PFS for MM (P=0.91 and 0.91) or NHL (P=0.82 and 0.85). Further, no significant difference in OS was observed between patients with PCR-positive compared with PCR-negative PBSC grafts with aggressive NHL histology (P=0.74) or indolent disease (P=0.29). Patients with contaminating tumour cells in autologous PBSCs do not have worsened OS or PFS in MM or NHL. Tumour cells detected by sensitive molecular methods in PBSC collections may be distinct from cells contaminating marrow and appear to have limited utility in identifying patients with MM and B-cell NHL who would benefit from purging strategies.