Publications by authors named "Stuart Adams"

55 Publications

Autologous Ex Vivo Lentiviral Gene Therapy for Adenosine Deaminase Deficiency.

N Engl J Med 2021 May 11. Epub 2021 May 11.

From the Departments of Microbiology, Immunology, and Molecular Genetics (D.B.K., K.L.S., D.A.C.-S., D.T., A.D., A. Icreverzi, P.B., B.C.F., R.P.H., M.C., A.Y., K.M.C., C.E.C., R.Z.), Pediatrics (D.B.K., T.B.M., S.D.O., S.S.), and Pathology and Laboratory Medicine (G.M.C.) and the Eli and Edythe Broad Center of Regenerative Medicine and Stem Cell Research (D.B.K., G.M.C.), University of California, Los Angeles (UCLA), and the Department of Pharmaceutical Services, Ronald Reagan UCLA Medical Center (J.T.), Los Angeles, and Stanford School of Medicine, Palo Alto (A.J.S.) - all in California; University College London Great Ormond Street Institute of Child Health and Great Ormond Street Hospital for Children NHS Foundation Trust (C.B., J.X.-B., V.T., K. Soni, K. Snell, D.L.-R., K.F.B., K.C.G., C.R., N.I., S.A., H.R., C.U., A.J.T., H.B.G.), and Orchard Therapeutics (Europe) (D.A.C.-S., S.A., F.L., M.K., A.S., H.B.G.) - all in London; the National Institute of Allergy and Infectious Diseases (H.L.M.) and the National Human Genome Research Institute (E.G., R.S., F.C.), National Institutes of Health, Bethesda, MD; Cure 4 The Kids Foundation, Las Vegas (A. Ikeda); Cincinnati Children's Hospital Medical Center, Cincinnati (L.R.); Indiana University School of Medicine, Indianapolis (K.C.); Duke University, Durham, NC (R.P., R.H.B., M.H.); Division of Immunology and Allergy, Lausanne University Hospital and University of Lausanne, Lausanne, Switzerland (F.C.); and GeneWerk (M.Z., R.H., I.L., M.S.) and the German Cancer Research Center and the National Center for Tumor Diseases (M.Z., M.S.) - all in Heidelberg, Germany.

Background: Severe combined immunodeficiency due to adenosine deaminase (ADA) deficiency (ADA-SCID) is a rare and life-threatening primary immunodeficiency.

Methods: We treated 50 patients with ADA-SCID (30 in the United States and 20 in the United Kingdom) with an investigational gene therapy composed of autologous CD34+ hematopoietic stem and progenitor cells (HSPCs) transduced ex vivo with a self-inactivating lentiviral vector encoding human . Data from the two U.S. studies (in which fresh and cryopreserved formulations were used) at 24 months of follow-up were analyzed alongside data from the U.K. study (in which a fresh formulation was used) at 36 months of follow-up.

Results: Overall survival was 100% in all studies up to 24 and 36 months. Event-free survival (in the absence of reinitiation of enzyme-replacement therapy or rescue allogeneic hematopoietic stem-cell transplantation) was 97% (U.S. studies) and 100% (U.K. study) at 12 months; 97% and 95%, respectively, at 24 months; and 95% (U.K. study) at 36 months. Engraftment of genetically modified HSPCs persisted in 29 of 30 patients in the U.S. studies and in 19 of 20 patients in the U.K. study. Patients had sustained metabolic detoxification and normalization of ADA activity levels. Immune reconstitution was robust, with 90% of the patients in the U.S. studies and 100% of those in the U.K. study discontinuing immunoglobulin-replacement therapy by 24 months and 36 months, respectively. No evidence of monoclonal expansion, leukoproliferative complications, or emergence of replication-competent lentivirus was noted, and no events of autoimmunity or graft-versus-host disease occurred. Most adverse events were of low grade.

Conclusions: Treatment of ADA-SCID with ex vivo lentiviral HSPC gene therapy resulted in high overall and event-free survival with sustained expression, metabolic correction, and functional immune reconstitution. (Funded by the National Institutes of Health and others; ClinicalTrials.gov numbers, NCT01852071, NCT02999984, and NCT01380990.).
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http://dx.doi.org/10.1056/NEJMoa2027675DOI Listing
May 2021

Spontaneous bleeding from an unusual renal mass: A case of gestational choriocarcinoma related to previous pregnancy over a decade earlier.

Urol Case Rep 2021 Jul 17;37:101614. Epub 2021 Mar 17.

Nepean Urology Research Group, Nepean Hospital, Kingswood, NSW, Australia.

Gestational choriocarcinoma is an uncommon trophoblastic malignancy, occurring in females after pregnancy, which is rarely encountered by urologists. It can be rapidly progressive, however metastases to other organs can occur after a prolonged latency period. We describe a rare case of solitary metastatic gestational choriocarcinoma presenting with spontaneous bleeding from a renal mass, over a decade after the associated pregnancy with a presumed sub-clinical primary tumour. This case demonstrates the importance of recognising gestational choriocarcinoma as a potential differential diagnosis of spontaneous bleeding renal mass in females of child-bearing age as a urologist given the often-aggressive nature of the disease.
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http://dx.doi.org/10.1016/j.eucr.2021.101614DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8020421PMC
July 2021

Long-term lymphoid progenitors independently sustain naïve T and NK cell production in humans.

Nat Commun 2021 03 12;12(1):1622. Epub 2021 Mar 12.

Great Ormond Street Institute of Child Health Faculty of Population Health Sciences, London, UK.

Our mathematical model of integration site data in clinical gene therapy supported the existence of long-term lymphoid progenitors capable of surviving independently from hematopoietic stem cells. To date, no experimental setting has been available to validate this prediction. We here report evidence of a population of lymphoid progenitors capable of independently maintaining T and NK cell production for 15 years in humans. The gene therapy patients of this study lack vector-positive myeloid/B cells indicating absence of engineered stem cells but retain gene marking in both T and NK. Decades after treatment, we can still detect and analyse transduced naïve T cells whose production is likely maintained by a population of long-term lymphoid progenitors. By tracking insertional clonal markers overtime, we suggest that these progenitors can support both T and NK cell production. Identification of these long-term lymphoid progenitors could be utilised for the development of next generation gene- and cancer-immunotherapies.
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http://dx.doi.org/10.1038/s41467-021-21834-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7954865PMC
March 2021

T cell phenotype in paediatric heart transplant recipients.

Pediatr Transplant 2020 Dec 16:e13930. Epub 2020 Dec 16.

UCL Great Ormond Street Institute of Child Health, London, UK.

Paediatric heart transplantation recipients suffer an increased incidence of infectious, autoimmune and allergic problems. The relative roles of thymus excision and immunosuppressive treatments in contributing to these sequelae are not clear. We compared the immunological phenotypes of 25 heart transplant recipients (Tx), 10 children who underwent thymus excision during non-transplantation cardiac surgery (TE) and 25 age range-matched controls, in two age bands: 1-9 and 10-16 years. Significant differences from controls were seen mainly in the younger age band with Tx showing lower CD3 and CD4 cell counts whilst TE showed lower CD8 cell counts. Naïve T cell and recent thymic emigrant proportions and counts were significantly lower than controls in both groups in the lower age band. T cell recombination excision circle (TREC) levels were lower than controls in both groups in both age bands. There were no differences in regulatory T cells, but in those undergoing thymus excision in infancy, their proportions were higher in TE than Tx, a possible direct effect of immunosuppression. T cell receptor V beta spectratyping showed fewer peaks in both groups than in controls (predominantly in the older age band). Thymus excision in infancy was associated with lower CD8 cell counts and higher proportions of Tregs in TE compared to Tx. These data are consistent with thymus excision, particularly in infancy, being the most important influence on immunological phenotype after heart transplantation.
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http://dx.doi.org/10.1111/petr.13930DOI Listing
December 2020

Cord blood CD8+ T-cell expansion following granulocyte transfusions eradicates refractory leukemia.

Blood Adv 2020 09;4(17):4165-4174

Department of Bone Marrow Transplantation, Royal Manchester Children's Hospital, Manchester, United Kingdom.

The action of hematopoietic cell transplantation in controlling leukemia is principally mediated by donor T cells directed against residual recipient malignant cells. However, its utility is limited by graft-versus-host disease (GVHD), where alloreactivity is extended beyond leukemic and marrow cells. In a human/murine chimeric model, we previously showed that the preferential infiltration of cord blood (CB) CD8+ T cells eradicates an Epstein-Barr virus-driven lymphoblastoid tumor without causing xenogeneic GVHD. In the clinic, however, cord blood CD8+ T-cell reconstitution is significantly delayed, and the observation of such a robust antileukemia effect mediated by cord blood CD8+ T cells has not been reported. We describe an observation of very early T-cell expansion in 4 high-risk pediatric leukemia patients receiving third-party, pooled granulocytes after T cell-replete CB transplantation (CBT). The T-cell expansion was transient but robust, including expansion of CD8+ T cells, in contrast to the delayed CD8+ T-cell expansion ordinarily observed after T cell-replete CBT. The CD8+ T cells were polyclonal, rapidly switched to memory phenotype, and had the ability to mediate cytotoxicity. This phenomenon is reproducible, and each patient remains in long-term remission without GVHD. The results suggest that fetal-derived CB CD8+ T cells can be exploited to generate robust antileukemia effects without GVHD.
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http://dx.doi.org/10.1182/bloodadvances.2020001737DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7479944PMC
September 2020

Thirty-seven jump-landing biomechanical variables are associated with asymptomatic patellar tendon abnormality and patellar tendinopathy: A systematic review.

Phys Ther Sport 2020 Sep 18;45:38-55. Epub 2020 Apr 18.

University of Newcastle, School of Environmental and Life Sciences, Ourimbah, NSW, Australia; University of Newcastle, Priority Research Centre for Physical Activity and Nutrition, Callaghan, NSW, Australia. Electronic address:

Objective: To identify studies that report three-dimensional (3D) biomechanical analysis of jump-landing tasks in relation to athletes with current patellar tendinopathy (PT), and/or asymptomatic with history of PT or patellar tendon abnormality (PTA) on diagnostic imaging.

Methods: Five electronic databases were searched. Included articles were required to: (1) investigate the 3D biomechanics of a jump-landing task; (2) be cross-sectional or longitudinal in design; and (3) include participants that had symptomatic PT, were asymptomatic with a history of PT, asymptomatic with PTA on diagnostic imaging and/or asymptomatic with an unknown pathology or PT history.

Results: Thirty-seven statistically significant jump-landing variables were associated with PT, history of PT and/or PTA. The only consistent variable that could be replicated between studies was knee flexion angle at initial foot-ground contact (IC) and an altered hip flexion/extension strategy during a horizontal land phase of a vertical stop-jump.

Conclusion: Isolated vertical landings or take-offs alone may not be sensitive enough to identify key jump-landing variables associated with PT, thus clinicians and researchers should incorporate a whole jump-landing task with a horizontal landing component. Sagital plane hip and knee kinematics in a horizontal landing phase appear to provide the most valuable information for evaluating those with PT.
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http://dx.doi.org/10.1016/j.ptsp.2020.03.011DOI Listing
September 2020

Metachronous ureteral metastasis of clear cell renal cell carcinoma in a duplex collecting system 1 year after nephrectomy.

Urol Case Rep 2020 Sep 16;32:101214. Epub 2020 Apr 16.

Nepean Urology Research Group, Nepean Hospital, PO Box 63, Penrith, NSW, 2751, Australia.

While renal cell carcinoma is known to metastasise in an unpredictable pattern, even after resection of a primary tumour, delayed ureteric metastasis is a very rarely reported phenomenon. In this case report, we describe a case of ipsilateral metachronous ureteric metastasis in a patient with a complete duplex collecting system. This case demonstrates some useful strategies in the diagnosis and treatment of renal cell carcinoma in this context. To our knowledge, this is the first case report of delayed ureteric metastasis of a renal clear cell carcinoma in a patient with a duplex collecting system.
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http://dx.doi.org/10.1016/j.eucr.2020.101214DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7184155PMC
September 2020

Urogenital Abnormalities in Adenosine Deaminase Deficiency.

J Clin Immunol 2020 05 19;40(4):610-618. Epub 2020 Apr 19.

Pediatric Immunohematology and Stem Cell Program, IRCCS San Raffaele Scientific Institute, Via Olgettina, 60, 20132, Milan, Italy.

Background: Improved survival in ADA-SCID patients is revealing new aspects of the systemic disorder. Although increasing numbers of reports describe the systemic manifestations of adenosine deaminase deficiency, currently there are no studies in the literature evaluating genital development and pubertal progress in these patients.

Methods: We collected retrospective data on urogenital system and pubertal development of 86 ADA-SCID patients followed in the period 2000-2017 at the Great Ormond Street Hospital (UK) and 5 centers in Italy. In particular, we recorded clinical history and visits, and routine blood tests and ultrasound scans were performed as part of patients' follow-up.

Results And Discussion: We found a higher frequency of congenital and acquired undescended testes compared with healthy children (congenital, 22% in our sample, 0.5-4% described in healthy children; acquired, 16% in our sample, 1-3% in healthy children), mostly requiring orchidopexy. No urogenital abnormalities were noted in females. Spontaneous pubertal development occurred in the majority of female and male patients with a few cases of precocious or delayed puberty; no patient presented high FSH values. Neither ADA-SCID nor treatment performed (PEG-ADA, BMT, or GT) affected pubertal development or gonadic function.

Conclusion: In summary, this report describes a high prevalence of cryptorchidism in a cohort of male ADA-SCID patients which could represent an additional systemic manifestation of ADA-SCID. Considering the impact urogenital and pubertal abnormalities can have on patients' quality of life, we feel it is essential to include urogenital evaluation in ADA-SCID patients to detect any abnormalities, initiate early treatment, and prevent long-term complications.
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http://dx.doi.org/10.1007/s10875-020-00777-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7253380PMC
May 2020

Population Pharmacokinetics of Alemtuzumab (Campath) in Pediatric Hematopoietic Cell Transplantation: Towards Individualized Dosing to Improve Outcome.

Clin Pharmacokinet 2019 12;58(12):1609-1620

Division of Stem Cell Transplantation, Department of Pediatrics, Leiden University Medical Center, Leiden, The Netherlands.

Background And Objective: Alemtuzumab (Campath) is used to prevent graft-versus-host disease and graft failure following pediatric allogeneic hematopoietic cell transplantation. The main toxicity includes delayed immune reconstitution, subsequent viral reactivations, and leukemia relapse. Exposure to alemtuzumab is highly variable upon empirical milligram/kilogram dosing.

Methods: A population pharmacokinetic (PK) model for alemtuzumab was developed based on a total of 1146 concentration samples from 206 patients, aged 0.2-19 years, receiving a cumulative intravenous dose of 0.2-1.5 mg/kg, and treated between 2003 and 2015 in two centers.

Results: Alemtuzumab PK were best described using a two-compartment model with a parallel saturable and linear elimination pathway. The linear clearance pathway, central volume of distribution, and intercompartmental distribution increased with body weight. Blood lymphocyte counts, a potential substrate for alemtuzumab, did not impact clearance.

Conclusion: The current practice with uniform milligram/kilogram doses leads to highly variable exposures in children due to the non-linear relationship between body weight and alemtuzumab PK. This model may be used for individualized dosing of alemtuzumab.
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http://dx.doi.org/10.1007/s40262-019-00782-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6885503PMC
December 2019

Clinical benefit of a high-throughput sequencing approach for minimal residual disease in acute lymphoblastic leukemia.

Pediatr Blood Cancer 2019 08 29;66(8):e27787. Epub 2019 Apr 29.

SIHMDS-Haematology, Great Ormond Street Hospital for Children NHS Foundation Trust, London, UK.

The molecular detection of minimal residual disease (MRD) is standard of care in acute lymphoblastic leukemia to personalize the stratification of patients to appropriate intensity chemotherapy regimens. High-throughput sequencing (HTS) techniques are driving changes to MRD methodologies. Our study demonstrates HTS can identify suitable diagnostic markers, even in cases where traditional screening has been unsuccessful. Markers identified by HTS were used to track MRD using standard real-time quantitative PCR. We show, with six patient examples, clinical benefits of utilizing HTS to screen diagnostic samples and its necessity when traditional screening techniques fail. This is practical evidence that current MRD diagnostic marker screening should be replaced by an HTS approach.
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http://dx.doi.org/10.1002/pbc.27787DOI Listing
August 2019

Clinical and immunological features in a cohort of patients with partial DiGeorge syndrome followed at a single center.

Blood 2019 06 23;133(24):2586-2596. Epub 2019 Apr 23.

Department of Paediatric Immunology.

DiGeorge syndrome (DGS) is a primary immunodeficiency characterized by various degrees of T-cell deficiency. In partial DGS (pDGS), other risk factors could predispose to recurrent infections, autoimmunity, and allergy. The aim of this study was to assess the effect of different factors in the development of infections, autoimmunity, and/or allergy in patients with pDGS. We studied 467 pDGS patients in follow-up at Great Ormond Street Hospital. Using a multivariate approach, we observed that palatal anomalies represent a risk factor for the development of recurrent otitis media with effusion. Gastroesophageal reflux/dysphagia and asthma/rhinitis represent a risk factor for the development of recurrent upper respiratory tract infections. Allergy and autoimmunity were associated with persistently low immunoglobulin M levels and lymphopenia, respectively. Patients with autoimmunity showed lower levels of CD3, CD3CD4, and naïve CD4CD45RACD27 T lymphocytes compared with pDGS patients without autoimmunity. We also observed that the physiological age-related decline of the T-cell number was slower in pDGS patients compared with age-matched controls. The age-related recovery of the T-cell number depended on a homeostatic peripheral proliferation of T cells, as suggested by an accelerated decline of the naïve T lymphocytes in pDGS as well as a more skewed T-cell repertoire in older pDGS patients. These evidences suggest that premature CD4 T-cell aging and lymphopenia induced spontaneous peripheral T-cell proliferation might contribute to the pathogenesis of autoimmunity in patients with pDGS. Infections in these patients represent, in most of the cases, a complication of anatomical or gastroenterological anomalies rather than a feature of the underlying immunodeficiency.
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http://dx.doi.org/10.1182/blood.2018885244DOI Listing
June 2019

New graft manipulation strategies improve the outcome of mismatched stem cell transplantation in children with primary immunodeficiencies.

J Allergy Clin Immunol 2019 07 4;144(1):280-293. Epub 2019 Feb 4.

Molecular and Cellular Immunology Unit, University College London (UCL) Great Ormond Street Institute of Child Health, London, United Kingdom; Blood and Bone Marrow Transplant Unit, Great Ormond Street Hospital, London, United Kingdom.

Background: Mismatched stem cell transplantation is associated with a high risk of graft loss, graft-versus-host disease (GvHD), and transplant-related mortality. Alternative graft manipulation strategies have been used over the last 11 years to reduce these risks.

Objective: We investigated the outcome of using different graft manipulation strategies among children with primary immunodeficiencies.

Methods: Between 2006 and 2017, 147 patients with primary immunodeficiencies received 155 mismatched grafts: 30 T-cell receptor (TCR) αβ/CD19-depleted grafts, 43 cord blood (CB) grafts (72% with no serotherapy), 17 CD34 selection with T-cell add-back grafts, and 65 unmanipulated grafts.

Results: The estimated 8-year survival of the entire cohort was 79%, transplant-related mortality was 21.7%, and the graft failure rate was 6.7%. Posttransplantation viral reactivation, grade II to IV acute graft-versus-host disease (aGvHD), and chronic graft-versus-host disease (cGvHD) complicated 49.6%, 35%, and 15% of transplantations, respectively. Use of TCRαβ/CD19 depletion was associated with a significantly lower incidence of grade II to IV aGvHD (11.5%) and cGvHD (0%), although with a greater incidence of viral reactivation (70%) in comparison with other grafts. T-cell immune reconstitution was robust among CB transplants, although with a high incidence (56.7%) of grade II to IV aGvHD. Stable full donor engraftment was significantly greater at 80% among TCRαβ/CD19-depleted and CB transplants versus 40% to 60% among the other groups.

Conclusions: Rapidly accessible CB and haploidentical grafts are suitable alternatives for patients with no HLA-matched donor. Cord transplantation without serotherapy and TCRαβ/CD19-depleted grafts produced comparable survival rates of around 80%, although with a high rate of aGvHD with the former and a high risk of viral reactivation with the latter that need to be addressed.
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http://dx.doi.org/10.1016/j.jaci.2019.01.030DOI Listing
July 2019

Clinical T Cell Receptor Repertoire Deep Sequencing and Analysis: An Application to Monitor Immune Reconstitution Following Cord Blood Transplantation.

Front Immunol 2018 5;9:2547. Epub 2018 Nov 5.

Infection, Immunity and Inflammation Section, Great Ormond Street Institute of Child Health, University College London, London, United Kingdom.

Spectratyping assays are well recognized as the clinical gold standard for assessing the T cell receptor (TCR) repertoire in haematopoietic stem cell transplant (HSCT) recipients. These assays use length distributions of the hyper variable complementarity-determining region 3 (CDR3) to characterize a patient's T cell immune reconstitution post-transplant. However, whilst useful, TCR spectratyping is notably limited by its resolution, with the technique unable to provide data on the individual clonotypes present in a sample. High-resolution clonotype data are necessary to provide quantitative clinical TCR assessments and to better understand clonotype dynamics during clinically relevant events such as viral infections or GvHD. In this study we developed and applied a CDR3 Next Generation Sequencing (NGS) methodology to assess the TCR repertoire in cord blood transplant (CBT) recipients. Using this, we obtained comprehensive TCR data from 16 CBT patients and 5 control cord samples at Great Ormond Street Hospital (GOSH). These were analyzed to provide a quantitative measurement of the TCR repertoire and its constituents in patients post-CBT. We were able to both recreate and quantify inferences typically drawn from spectratyping data. Additionally, we demonstrate that an NGS approach to TCR assessment can provide novel insights into the recovery of the immune system in these patients. We show that NGS can be used to accurately quantify TCR repertoire diversity and to provide valuable inference on clonotypes detected in a sample. We serially assessed the progress of T cell immune reconstitution demonstrating that there is dramatic variation in TCR diversity immediately following transplantation and that the dynamics of T cell immune reconstitution is perturbed by the presence of GvHD. These findings provide a proof of concept for the adoption of NGS TCR sequencing in clinical practice.
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http://dx.doi.org/10.3389/fimmu.2018.02547DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6231291PMC
September 2019

Naive B Cell Output in HIV-Infected and HIV-Uninfected Children.

AIDS Res Hum Retroviruses 2019 01 26;35(1):33-39. Epub 2018 Dec 26.

1 UCL Great Ormond Street Institute of Child Health, London, United Kingdom.

In this study, we aimed to quantify KREC (kappa-deleting recombination excision circle) levels and naive B cell output in healthy HIV-uninfected children, compared with HIV-infected South African children, before and after starting ART (antiretroviral therapy). Samples were acquired from a Child Wellness Clinic (n = 288 HIV-uninfected South African children, 2 weeks-12 years) and the Children with HIV Early Antiretroviral Therapy (CHER) trial (n = 153 HIV-infected South African children, 7 weeks-8 years). Naive B cell output was estimated using a mathematical model combining KREC levels to reflect B cell emigration into the circulation, flow cytometry measures of naive unswitched B cells to quantify total body naive B cells, and their rates of proliferation using the intracellular marker Ki67. Naive B cell output increases from birth to 1 year, followed by a decline and plateau into late childhood. HIV-infected children on or off ART had higher naive B cell outputs than their uninfected counterparts (p = .01 and p = .04). This is the first study to present reference ranges for measurements of KRECs and naive B cell output in healthy and HIV-infected children. Comparison between HIV-uninfected healthy children and HIV-infected children suggests that HIV may increase naive B cell output. Further work is required to fully understand the mechanisms involved and clinical value of measuring naive B cell output in children.
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http://dx.doi.org/10.1089/AID.2018.0170DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6863188PMC
January 2019

One hundred percent survival after transplantation of 34 patients with Wiskott-Aldrich syndrome over 20 years.

J Allergy Clin Immunol 2018 11 25;142(5):1654-1656.e7. Epub 2018 Jul 25.

Blood and Marrow Transplant Unit, Great Ormond Street Hospital NHS Trust, University College London, London, United Kingdom.

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http://dx.doi.org/10.1016/j.jaci.2018.06.042DOI Listing
November 2018

CD19CD24CD38 B Cells Are Expanded in Juvenile Dermatomyositis and Exhibit a Pro-Inflammatory Phenotype After Activation Through Toll-Like Receptor 7 and Interferon-α.

Front Immunol 2018 22;9:1372. Epub 2018 Jun 22.

Centre for Adolescent Rheumatology, Arthritis Research UK, University College London Hospital and Great Ormond Street Hospital, London, United Kingdom.

Juvenile dermatomyositis (JDM) is a rare form of childhood autoimmune myositis that presents with proximal muscle weakness and skin rash. B cells are strongly implicated in the pathogenesis of the disease, but the underlying mechanisms are unknown. Therefore, the main objective of our study was to investigate mechanisms driving B cell lymphocytosis and define pathological features of B cells in JDM patients. Patients were recruited through the UK JDM Cohort and Biomarker study. Peripheral blood B cell subpopulations were immunophenotyped by flow cytometry. The results identified that immature transitional B cells were significantly expanded in active JDM, actively dividing, and correlated positively with disease activity. Protein and RNAseq analysis revealed high interferon alpha (IFNα) and TLR7-pathway signatures pre-treatment. Stimulation of B cells through TLR7/8 promoted both IL-10 and IL-6 production in controls but failed to induce IL-10 in JDM patient cells. Interrogation of the CD40-CD40L pathway (known to induce B cell IL-10 and IL-6) revealed similar expression of IL-10 and IL-6 in B cells cultured with CD40L from both JDM patients and controls. In conclusion, JDM patients with active disease have a significantly expanded immature transitional B cell population which correlated with the type I IFN signature. Activation through TLR7 and IFNα may drive the expansion of immature transitional B cells in JDM and skew the cells toward a pro-inflammatory phenotype.
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http://dx.doi.org/10.3389/fimmu.2018.01372DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6024011PMC
June 2018

Chimerism Analysis in the Pediatric Setting: Direct PCR from Bone Marrow, Whole Blood, and Cell Fractions.

J Mol Diagn 2018 05 21;20(3):381-388. Epub 2018 Feb 21.

Specialist Integrated Haematology and Malignancy Diagnostic Service, Department of Haematology, Camelia Botnar Laboratories, Great Ormond Street Hospital for Children NHS Trust, London, United Kingdom.

Certain blood components and anticoagulants interfere with the PCR process and subsequent analysis. Here we demonstrate that reliable test results can be obtained for chimerism analysis despite omitting a DNA-extraction step and performing PCR and fragment analysis directly on bone marrow, whole blood, and individual cell fractions. For chimerism analysis, direct-tissue PCR is possible with the use of a robust, commercially available PCR mix containing a DNA polymerase capable of DNA amplification directly from the sample without the need for pretreatment. A total of 178 chimerism samples were processed directly, and results were compared to those obtained from the corresponding DNA sample. No differences were observed between the two sets of results. For the cell fraction-purity assessment, commercially available PCR kits were used directly on T and B cells without the use of any additional lysing agent. A total of 53 purity samples and their corresponding DNA samples were analyzed and showed a correlation similar to that obtained for the chimerism samples. The results show that chimerism testing and associated cell fraction-purity assessment can be performed reliably without the need for prior DNA extraction and that this method can easily be integrated into existing routine laboratory procedures.
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http://dx.doi.org/10.1016/j.jmoldx.2018.02.003DOI Listing
May 2018

Thymus transplantation for complete DiGeorge syndrome: European experience.

J Allergy Clin Immunol 2017 Dec 8;140(6):1660-1670.e16. Epub 2017 Apr 8.

The Children's Clinic, Tartu University Hospital, Tartu, Estonia.

Background: Thymus transplantation is a promising strategy for the treatment of athymic complete DiGeorge syndrome (cDGS).

Methods: Twelve patients with cDGS underwent transplantation with allogeneic cultured thymus.

Objective: We sought to confirm and extend the results previously obtained in a single center.

Results: Two patients died of pre-existing viral infections without having thymopoiesis, and 1 late death occurred from autoimmune thrombocytopenia. One infant had septic shock shortly after transplantation, resulting in graft loss and the need for a second transplant. Evidence of thymopoiesis developed from 5 to 6 months after transplantation in 10 patients. Median circulating naive CD4 counts were 44 × 10/L (range, 11-440 × 10/L) and 200 × 10/L (range, 5-310 × 10/L) at 12 and 24 months after transplantation and T-cell receptor excision circles were 2,238/10 T cells (range, 320-8,807/10 T cells) and 4,184/10 T cells (range, 1,582-24,596/10 T cells). Counts did not usually reach normal levels for age, but patients were able to clear pre-existing infections and those acquired later. At a median of 49 months (range, 22-80 months), 8 have ceased prophylactic antimicrobials, and 5 have ceased immunoglobulin replacement. Histologic confirmation of thymopoiesis was seen in 7 of 11 patients undergoing biopsy of transplanted tissue, including 5 showing full maturation through to the terminal stage of Hassall body formation. Autoimmune regulator expression was also demonstrated. Autoimmune complications were seen in 7 of 12 patients. In 2 patients early transient autoimmune hemolysis settled after treatment and did not recur. The other 5 experienced ongoing autoimmune problems, including thyroiditis (3), hemolysis (1), thrombocytopenia (4), and neutropenia (1).

Conclusions: This study confirms the previous reports that thymus transplantation can reconstitute T cells in patients with cDGS but with frequent autoimmune complications in survivors.
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http://dx.doi.org/10.1016/j.jaci.2017.03.020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5716670PMC
December 2017

Development and Validation of Ion Chromatography-Tandem Mass Spectrometry-Based Method for the Multiresidue Determination of Polar Ionic Pesticides in Food.

J Agric Food Chem 2017 Aug 21;65(34):7294-7304. Epub 2017 Apr 21.

Special Solutions Center, Thermo Fisher Scientific , Dreieich, Germany.

An extraction method using acidified methanol based on the quick polar pesticide (QuPPe) method using suppressed ion chromatography coupled to mass spectrometry was developed and validated for the direct analysis of polar pesticides, without the need for derivatization or ion pairing, in cereals and grapes. The method was robust, and results for glyphosate, aminomethyl phosphonic acid (AMPA), N-acetyl-AMPA, glufosinate, 3-methylphosphinicopropionic acid (3-MPPA), N-acetyl glufosinate, ethephon, chlorate, perchlorate, fosetyl aluminum, and phosphonic acid at three concentration levels (typically 0.01, 0.05, and 0.1 mg/kg) were compliant with SANTE/11945/2015 guideline method performance criteria. Cereal-based infant food proved to be a more challenging matrix and validated only for glyphosate, chlorate, and perchlorate at 0.005, 0.01, and 0.05 mg/kg. The developed method enables the multiresidue analysis of 12 ionic pesticides and relevant metabolites in a single analysis. Until now, the analysis of these compounds required several different single-residue methods using different chromatographic conditions. This multiresidue approach offers the possibility of more cost-effective and more efficient monitoring of polar ionic pesticides and contaminants that are of concern to food regulation bodies and consumers.
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http://dx.doi.org/10.1021/acs.jafc.7b00476DOI Listing
August 2017

Evaluation of direct analysis in real time for the determination of highly polar pesticides in lettuce and celery using modified Quick Polar Pesticides Extraction method.

J Chromatogr A 2017 May 15;1496:37-44. Epub 2017 Mar 15.

Fera Science Ltd, Sand Hutton, York, YO41 1LZ, United Kingdom.

Direct analysis in real time (DART) was evaluated for the determination of a number of highly polar pesticides using the Quick Polar Pesticides Extraction (QuPPe) method. DART was hyphenated to high resolution mass spectrometry (HRMS) in order to get the required selectivity that allows the determination of these compounds in complex samples such as lettuce and celery. Experimental parameters such as desorption temperature, scanning speed, and distances between the DART ion source and MS inlet were optimized. Two different mass analyzers (Orbitrap and QTOF) and two accessories for sample introduction (Dip-it tips and QuickStrip™ sample cards) were evaluated. An extra clean-up step using primary-secondary amine (PSA) was included in the QuPPe method to improve sensitivity. The main limitation found was in-source fragmentation, nevertheless QuPPe-DART-HRMS proved to be a fast and reliable tool with quantitative capabilities for at least seven compounds: amitrole, cyromazine, propamocarb, melamine, diethanolamine, triethanolamine and 1,2,4-triazole. The limits of detection ranged from 20 to 60μg/kg. Recoveries for fortified samples ranged from 71 to 115%, with relative standard deviations <18%.
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http://dx.doi.org/10.1016/j.chroma.2017.03.020DOI Listing
May 2017

Molecular remission of infant B-ALL after infusion of universal TALEN gene-edited CAR T cells.

Sci Transl Med 2017 01;9(374)

Great Ormond Street Institute of Child Health, University College London, London WC1N 1EH, UK.

Autologous T cells engineered to express chimeric antigen receptor against the B cell antigen CD19 (CAR19) are achieving marked leukemic remissions in early-phase trials but can be difficult to manufacture, especially in infants or heavily treated patients. We generated universal CAR19 (UCART19) T cells by lentiviral transduction of non-human leukocyte antigen-matched donor cells and simultaneous transcription activator-like effector nuclease (TALEN)-mediated gene editing of T cell receptor α chain and CD52 gene loci. Two infants with relapsed refractory CD19 B cell acute lymphoblastic leukemia received lymphodepleting chemotherapy and anti-CD52 serotherapy, followed by a single-dose infusion of UCART19 cells. Molecular remissions were achieved within 28 days in both infants, and UCART19 cells persisted until conditioning ahead of successful allogeneic stem cell transplantation. This bridge-to-transplantation strategy demonstrates the therapeutic potential of gene-editing technology.
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http://dx.doi.org/10.1126/scitranslmed.aaj2013DOI Listing
January 2017

Mutations in linker for activation of T cells (LAT) lead to a novel form of severe combined immunodeficiency.

J Allergy Clin Immunol 2017 02 15;139(2):634-642.e5. Epub 2016 Jul 15.

Infection, Immunity, Inflammation and Physiological Medicine, UCL Institute of Child Health, London, United Kingdom; Department of Clinical Immunology, Great Ormond Street Hospital NHS Trust, London, United Kingdom. Electronic address:

Background: Signaling through the T-cell receptor (TCR) is critical for T-cell development and function. Linker for activation of T cells (LAT) is a transmembrane adaptor signaling molecule that is part of the TCR complex and essential for T-cell development, as demonstrated by LAT-deficient mice, which show a complete lack of peripheral T cells.

Objective: We describe a pedigree affected by a severe combined immunodeficiency phenotype with absent T cells and normal B-cell and natural killer cell numbers. A novel homozygous frameshift mutation in the gene encoding for LAT was identified in this kindred.

Methods: Genetic, molecular, and functional analyses were used to identify and characterize the LAT defect. Clinical and immunologic analysis of patients was also performed and reported.

Results: Homozygosity mapping was used to identify potential defective genes. Sanger sequencing of the LAT gene showed a mutation that resulted in a premature stop codon and protein truncation leading to complete loss of function and loss of expression of LAT in the affected family members. We also demonstrate loss of LAT expression and lack of TCR signaling restoration in LAT-deficient cell lines reconstituted with a synthetic LAT gene bearing this severe combined immunodeficiency mutation.

Conclusion: For the first time, the results of this study show that inherited LAT deficiency should be considered in patients with combined immunodeficiency with T-cell abnormalities.
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http://dx.doi.org/10.1016/j.jaci.2016.05.036DOI Listing
February 2017

Effect of stem cell source on long-term chimerism and event-free survival in children with primary immunodeficiency disorders after fludarabine and melphalan conditioning regimen.

J Allergy Clin Immunol 2016 10 20;138(4):1152-1160. Epub 2016 Apr 20.

Department of Blood and Marrow Transplantation, Great Ormond Street Hospital for Children, London, United Kingdom; Molecular Immunology Unit, Institute of Child Health, University College London, London, United Kingdom.

Background: Reduced-intensity conditioning (RIC) regimens are increasingly being used in the transplantation of patients with primary immunodeficiency disorders (PIDs), but there are no large studies looking at long-term lineage-specific chimerism.

Objectives: We sought to analyze long-term chimerism and event-free survival in children undergoing transplantation for PIDs using RIC with fludarabine and melphalan (Flu/Melph) and to study the effect of donor type and stem cell source.

Methods: One hundred forty-two children underwent transplantation with RIC by using Flu/Melph and for PIDs by using bone marrow (n = 93) or peripheral blood stem cells (PBSCs; n = 49). Donors were matched unrelated donors (n = 72), mismatched unrelated donors (n = 37), matched sibling donors (n = 14), matched family donors (n = 12), and mismatched family donors (n = 7).

Results: Overall survival at a median follow-up of 7.5 years was 78%, irrespective of stem cell source or donor type. When bone marrow was used as the stem cell source, 26% of patients ended up with very low levels of donor chimerism (<10% donor), especially in the myeloid lineage. Event-free survival in this group was significantly lower compared with that in the rest of the group (25% vs 70%, P < .001). With the use of PBSCs, more than 90% of patients achieved complete donor chimerism or high-level mixed chimerism (>50% donor chimerism) in all lineages.

Conclusions: On the basis of our experience, we would suggest that PBSCs should be the stem cell source of choice in children with PIDs undergoing transplantation with Flu/Melph RIC from a matched donor source. This is most likely to ensure sustained high-level donor chimerism.
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http://dx.doi.org/10.1016/j.jaci.2016.01.053DOI Listing
October 2016

Erratum to: Adenosine Deaminase Deficient Severe Combined Immunodeficiency Presenting as Atypical Haemolytic Uraemic Syndrome.

J Clin Immunol 2016 May;36(4):413

Department of Clinical Immunology and Bone Marrow, Transplantation, Great Ormond Street Hospital National Health Service Trust, London, UK.

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http://dx.doi.org/10.1007/s10875-016-0256-7DOI Listing
May 2016

Spectrum of mutations in a cohort of UK patients with ADA deficient SCID: Segregation of genotypes with specific ethnicities.

Clin Immunol 2015 Dec 5;161(2):174-9. Epub 2015 Aug 5.

Molecular and Cellular Immunology, Institute of Child Health, University College London, UK; Immunology and Bone Marrow Transplant Units, Great Ormond Street Hospital for Children, London NHS Trust, UK.

Severe combined immunodeficiency (SCID) arises from a number of different genetic defects, one of the most common being mutations in the gene encoding adenosine deaminase (ADA). In the UK, ADA deficient SCID compromises approximately 20% of all known cases of SCID. We carried out a retrospective analysis of the ADA gene in 46 known ADA deficient SCID patients on whom DNA had been stored. Here, we report a high frequency of two previously reported mutations and provide a link between the mutations and patient ethnicity within our patient cohort. We also report on 9 novel mutations that have been previously unreported.
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http://dx.doi.org/10.1016/j.clim.2015.08.001DOI Listing
December 2015

Adenosine deaminase deficient severe combined immunodeficiency presenting as atypical haemolytic uraemic syndrome.

J Clin Immunol 2015 May 15;35(4):366-72. Epub 2015 Apr 15.

Department of Clinical Immunology and Bone Marrow Transplantation, Great Ormond Street Hospital National Health Service Trust, London, UK.

Purpose: Adenosine deaminase (ADA) deficiency is a systemic disorder of purine metabolism. Deficiency of the purine salvage enzyme ADA leads to the build-up of the toxic metabolites, deoxyadenosine triphosphate and deoxyadenosine. ADA is ubiquitously expressed in all tissues of the body but most profoundly affects lymphocyte development and function leading to severe combined immunodeficiency (SCID). Unlike most other forms of SCID, ADA deficiency also results in non-immunologic manifestations. Associations between ADA deficiency and sensorineural hearing loss, behavioural abnormalities, non-infectious pulmonary disease and skeletal dysplasia are all recognised, and affect the long term outcome for these patients. Identification of new non-immunological manifestations and clinical presentations of ADA deficiency is essential to allow early optimisation of supportive care.

Methods And Results: Here we report four patients with ADA deficiency whose presenting feature was haemolytic uremic syndrome (HUS). 3 of 4 patients were diagnosed with ADA deficiency only after developing HUS, and one diagnosis was made post mortem, after a sibling was diagnosed with SCID. Shiga-toxigenic organisms were not isolated from any of the patients. 2 patients made a good recovery from their HUS with supportive treatment and initiation of PEG-ADA. Both remain well on enzyme replacement with mild or no residual renal impairment.

Conclusions: Clinicians should be aware of this previously unreported non-immunologic manifestation of ADA deficiency.
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http://dx.doi.org/10.1007/s10875-015-0158-0DOI Listing
May 2015

A trial of alemtuzumab adjunctive therapy in allogeneic hematopoietic cell transplantation with minimal conditioning for severe combined immunodeficiency.

Pediatr Transplant 2014 Sep 30;18(6):609-16. Epub 2014 Jun 30.

Division of Pediatric Allergy, Immunology, and Blood and Marrow Transplant, University of California San Francisco, Benioff Children's Hospital, San Francisco, CA, USA.

For infants with SCID the ideal conditioning regimen before allogeneic HCT would omit cytotoxic chemotherapy to minimize short- and long-term complications. We performed a prospective pilot trial with alemtuzumab monotherapy to overcome NK-cell mediated immunologic barriers to engraftment. We enrolled four patients who received CD34-selected haploidentical cells, two of whom failed to engraft donor T cells. The two patients who engrafted had delayed T-cell reconstitution, despite rapid clearance of circulating alemtuzumab. Although well-tolerated, alemtuzumab failed to overcome immunologic barriers to donor engraftment. Furthermore, alemtuzumab may slow T-cell development in patients with SCID in the setting of a T-cell depleted graft.
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http://dx.doi.org/10.1111/petr.12310DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4134761PMC
September 2014

An uncommon complication of percutaneous endoscopic gastrostomy tube placement.

Gastroenterology 2014 Aug 25;147(2):e3-4. Epub 2014 Jun 25.

Department of Anatomical Pathology, Nepean Hospital, NSW, Australia.

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http://dx.doi.org/10.1053/j.gastro.2014.02.041DOI Listing
August 2014