Early Enzyme Replacement Therapy Improves Hearing and Immune Defects in Adenosine Deaminase Deficient-Mice

Background: Inherited defects in adenosine deaminase (ADA) cause severe immune deficiency, which can be corrected by ADA enzyme replacement therapy (ERT). Additionally, ADA-deficient patients suffer from hearing impairment. We hypothesized that ADA-deficient (–/–) mice also exhibit hearing abnormalities and that ERT from an early age will improve the hearing and immune defects in these mice.Methods: Auditory brainstem evoked responses, organ weights, thymocytes numbers, and subpopulations, lymphocytes in peripheral blood as well as T lymphocytes in spleen were analyzed in ADA–/– and ADA-proficient littermate post-partum (pp). The cochlea was visualized by scanning electron microscopy (SEM). The effects of polyethylene glycol conjugated ADA (PEG-ADA) ERT or 40% oxygen initiated at 7 days pp on the hearing and immune abnormalities were assessed.Results: Markedly abnormal hearing thresholds responses were found in ADA–/– mice at low and medium tone frequencies. SEM demonstrated extensive damage to the cochlear hair cells of ADA–/– mice, which were splayed, short or missing, correlating with the hearing deficits. The hearing defects were not reversed when hypoxia in ADA–/– mice was corrected. Progressive immune abnormalities were detected in ADA–/– mice from 4 days pp, initially affecting the thymus followed by peripheral lymphocytes and T cells in the spleen. ERT initiated at 7 days pp significantly improved the hearing of ADA–/– mice as well as the number of thymocytes and T lymphocytes, although not all normalized.Conclusions: ADA deficiency is associated with hearing deficits and damage to cochlear hair cells. Early initiation of ERT improves the hearing and immune abnormalities

Hyperbaric Oxygen Therapy as a Sole Agent Is Not Immunosuppressant in a Highly Immunogenic Mouse Model

Background. Hyperbaric oxygen (HBO) therapy, which is used for many conditions, may also have immunosuppressive effects and could be used for prevention or treatment of graft-versus-host disease (GvHD). If HBO is immunosuppressant, then we hypothesize that HBO therapy will delay the T-cell mediated skin graft rejection. Methods. C57/BL6 black-coated (H2B) mice received skin graft from CBA (H2D) white-coated mice. Mice were treated with either 19 session of 240 kpa oxygen or 29 session of 300 kpa oxygen, for 90 minutes. Mice were housed either 4 per cage or separately, to prevent friction and mechanical factors that may affect graft survival. Skin grafts were assessed daily. Results. There was no difference in length of graft survival between mice that received either regimens of HBO therapy and mice that did not receive HBO therapy. Conclusions. HBO therapy, as a sole agent, did not delay skin graft rejection in a highly immunogenic mouse model

The transcriptional repressor bs69 is a conserved target of the e1a proteins from several human adenovirus species

Early region 1A (E1A) is the first viral protein produced upon human adenovirus (HAdV) infection. This multifunctional protein transcriptionally activates other HAdV early genes and reprograms gene expression in host cells to support productive infection. E1A functions by interacting with key cellular regulatory proteins through short linear motifs (SLiMs). In this study, the molecular determinants of interaction between E1A and BS69, a cellular repressor that negatively regulates E1A transactivation, were systematically defined by mutagenesis experiments. We found that a minimal sequence comprised of MPNLVPEV, which contains a conserved PXLXP motif and spans residues 112–119 in HAdV-C5 E1A, was necessary and sufficient in binding to the myeloid, Nervy, and DEAF-1 (MYND) domain of BS69. Our study also identified residues P113 and L115 as critical for this interaction. Furthermore, the HAdV-C5 and-A12 E1A proteins from species C and A bound BS69, but those of HAdV-B3,-E4,-D9,-F40, and-G52 from species B, E, D, F, and G, respectively, did not. In addition, BS69 functioned as a repressor of E1A-mediated transactivation, but only for HAdV-C5 and HAdV-A12 E1A. Thus, the PXLXP motif present in a subset of HAdV E1A proteins confers interaction with BS69, which serves as a negative regulator of E1A mediated transcriptional activation

Gene therapy for primary immune deficiencies: a Canadian perspective

The use of gene therapy (GT) for the treatment of primary immune deficiencies (PID) including severe combined immune deficiency (SCID) has progressed significantly in the recent years. In particular, long-term studies have shown that adenosine deaminase (ADA) gene delivery into ADA-deficient hematopoietic stem cells that are then transplanted into the patients corrects the abnormal function of the ADA enzyme, which leads to immune reconstitution. In contrast, the outcome was disappointing for patients with X-linked SCID, Wiskott–Aldrich syndrome and chronic granulomatous disease who received GT followed by autologous gene corrected transplantations, as many developed hematological malignancies. The malignancies were attributed to the predilection of the viruses used for gene delivery to integrated at oncogenic areas. The availability of safer and more efficient self-inactivating lentiviruses for gene delivery has reignited the interest in GT for many PID that are now in various stages of pre-clinical studies and clinical trials. Moreover, advances in early diagnosis of PID and gene editing technology coupled with enhanced abilities to generate and manipulate stem cells ex vivo are expected to further contribute to the benefit of GT for PID. Here we review the past, the present and the future of GT for PID, with particular emphasis on the Canadian perspective

Outcome of hematopoietic cell transplantation for DNA double-strand break repair disorders

Background: Rare DNA breakage repair disorders predispose to infection and lymphoreticular malignancies. Hematopoietic cell transplantation (HCT) is curative, but coadministered chemotherapy or radiotherapy is damaging because of systemic radiosensitivity. We collected HCT outcome data for Nijmegen breakage syndrome, DNA ligase IV deficiency, Cernunnos-XRCC4-like factor (Cernunnos-XLF) deficiency, and ataxia-telangiectasia (AT). Methods: Data from 38 centers worldwide, including indication, donor, conditioning regimen, graft-versus-host disease, and outcome, were analyzed. Conditioning was classified as myeloablative conditioning (MAC) if it contained radiotherapy or alkylators and reduced-intensity conditioning (RIC) if no alkylators and/or 150 mg/m(2) fludarabine or less and 40 mg/kg cyclophosphamide or less were used. Results: Fifty-five new, 14 updated, and 18 previously published patients were analyzed. Median age at HCT was 48 months (range, 1.5-552 months). Twenty-nine patients underwent transplantation for infection, 21 had malignancy, 13 had bone marrow failure, 13 received pre-emptive transplantation, 5 had multiple indications, and 6 had no information. Twenty-two received MAC, 59 received RIC, and 4 were infused; information was unavailable for 2 patients. Seventy-three of 77 patients with DNA ligase IV deficiency, Cernunnos-XLF deficiency, or Nijmegen breakage syndrome received conditioning. Survival was 53 (69%) of 77 and was worse for those receiving MAC than for those receiving RIC (P=.006). Most deaths occurred early after transplantation, suggesting poor tolerance of conditioning. Survival in patients with AT was 25%. Forty-one (49%) of 83 patients experienced acute GvHD, which was less frequent in those receiving RIC compared with those receiving MAC (26/56 [46%] vs 12/21 [57%], P=.45). Median follow-up was 35 months (range, 2-168 months). No secondary malignancies were reported during 15 years of follow-up. Growth and developmental delay remained after HCT; immune-mediated complications resolved. Conclusion: RIC HCT resolves DNA repair disorder associated immunodeficiency. Long-term follow-up is required for secondary malignancy surveillance. Routine HCT for AT is not recommended.Peer reviewe

TAT-mediated intracellular delivery of purine nucleoside phosphorylase corrects its deficiency in mice

Defects in purine nucleoside phosphorylase (PNP) enzyme activity result in abnormal nucleoside homeostasis, severe T cell immunodeficiency, neurological dysfunction, and early death. Protein transduction domain (PTD) can transfer molecules into cells and may help restore PNP activity in cases of PNP deficiency. However, long-term use of PTD to replace enzymes in animal models or patients has not previously been described. We fused human PNP to the HIV-TAT PTD and found that the fusion with TAT changed the retention and distribution of PNP in PNP-deficient mice. TAT induced rapid intracellular delivery of PNP into tissues, including the brain, prevented urinary excretion of PNP, and protected PNP from neutralizing antibodies, resulting in significant extension of the enzyme’s biological activity in vivo. Frequent TAT-PNP injections in PNP-deficient mice corrected the metabolic disorder and immune defects with no apparent toxicity. TAT-PNP remained effective over 24 weeks of treatment, resulting in continued improvement in immune function and extended survival. Our data demonstrate that TAT changes the properties of PNP in vivo and that long-term intracellular delivery of PNP by TAT corrects PNP deficiency in mice. We provide evidence to promote further use of PTD to treat diseases that require repeated intracellular enzyme or protein delivery

Monitoring patients with uncomplicated common variable immunodeficiency: a systematic review

Abstract Background Non-infectious complications have become a major cause of morbidity and mortality in patients with Common Variable Immunodeficiency (CVID). The monitoring of patients with CVID prior to the development of non-infectious complications is not well defined. Objective Our objectives were to systematically review the current literature on the monitoring of CVID patients without non-infectious complications and to develop recommendations for such monitoring. Methods MEDLINE and EMBASE were searched from January 1st, 2000 to March 25th, 2021. Studies on any aspects of CVID monitoring were included. Studies that included only children, on monitoring CVID patients with existing non-infectious complications, or in the format of case reports were excluded. Results Nine studies on CVID monitoring, including 3 cohort studies, 3 experts’ opinions, 2 consensus statements and a single guideline report were identified. These studies revealed that clinical assessment and bloodwork were preformed every 6 to 12 months in asymptomatic patients. Some centers performed computerized tomography scan of the chest every 2–5 years to identify chronic lung disease, although the majority did chest imaging in accordance with clinical indications. Pulmonary function tests were done annually at most centers. Most studies did not address the role of abdominal imaging to screen for liver diseases or endoscopy to screen for gastric cancer in asymptomatic patients with uncomplicated CVID. Conclusions There is paucity of evidence-based information to guide the routine monitoring of CVID patients without non-infectious complications. Prospective studies are needed to determine the best monitoring practices in this group of patients

Gene therapy for primary immune deficiencies: a Canadian perspective

Abstract The use of gene therapy (GT) for the treatment of primary immune deficiencies (PID) including severe combined immune deficiency (SCID) has progressed significantly in the recent years. In particular, long-term studies have shown that adenosine deaminase (ADA) gene delivery into ADA-deficient hematopoietic stem cells that are then transplanted into the patients corrects the abnormal function of the ADA enzyme, which leads to immune reconstitution. In contrast, the outcome was disappointing for patients with X-linked SCID, Wiskott–Aldrich syndrome and chronic granulomatous disease who received GT followed by autologous gene corrected transplantations, as many developed hematological malignancies. The malignancies were attributed to the predilection of the viruses used for gene delivery to integrated at oncogenic areas. The availability of safer and more efficient self-inactivating lentiviruses for gene delivery has reignited the interest in GT for many PID that are now in various stages of pre-clinical studies and clinical trials. Moreover, advances in early diagnosis of PID and gene editing technology coupled with enhanced abilities to generate and manipulate stem cells ex vivo are expected to further contribute to the benefit of GT for PID. Here we review the past, the present and the future of GT for PID, with particular emphasis on the Canadian perspective