Conferring alloantigen specificity to regulatory T cells: A comparative analysis of cell preparations undergoing clinical development in transplantation.

Conferring alloantigen-specificity to ex vivo expanded CD4+CD25+FOXP3+ regulatory T cells (Tregs) increases their capacity to counteract effector alloimmune responses following adoptive transfer into transplant recipients. Three strategies are currently undergoing clinical development, which involve the following: (1) expanding Tregs in the presence of donor B cells (donor alloantigen-reactive [DAR] Tregs); (2) culturing Tregs with donor cells in the presence of costimulation blockade (CSB-Tregs); and (3) transducing Tregs with an human leukocyte antigen A2-specific chimeric antigen receptor (CAR-Tregs). Our goal in this study was to assess the relative potency of each of these manufactured Treg products both in vitro and in vivo. When compared with polyclonal Tregs, all 3 manufacturing strategies increased the precursor frequency of alloreactive Tregs, and this was proportional to the overall in vitro immunosuppressive properties of the cell products. Accordingly, CAR-Tregs, which contained the highest frequency of donor-reactive Tregs, exhibited the strongest suppressive effects on a cell-per-cell basis. Similarly, in an in vivo mouse model of graft-vs-host disease, infusion of CAR-Tregs conferred a significantly longer recipient survival than any other Treg product. Our results highlighting the alloantigen-reactivity and associated immunosuppressive properties of different manufactured Treg products have implications for the mechanistic interpretation of currently ongoing clinical trials in transplantation.

Lactic acidosis and hyperlactatemia associated with lamivudine accumulation and sepsis in a kidney transplant recipient-a case report and review of the literature.

BACKGROUND: We report a case of sudden, lethal metabolic acidosis in a 70-year-old man on long-term nucleoside reverse transcriptase inhibitor (NRTI) -based antiretroviral therapy (ART) who had developed atypical necrotizing fasciitis 1 month after kidney transplantation. CASE PRESENTATION: The HIV infection of the patient was treated for the last four months with an integrase strand inhibitor (dolutegravir 50 mg/d) plus a NRTI backbone including lamivudine (150 mg/d) and abacavir (600 mg/d). In this renal transplant patient we hypothesize that the co-existence of sepsis, renal failure and an accumulation of lamivudine led to the development of fatal metabolic acidosis and hyperlactatemia. Although lamivudine is only rarely associated with hyperlactatemia, there is evidence that overdose may be a risk factor for developing it. In our patient the lamivudine concentration two days after stopping and during hemodiafiltration was more than 50 times higher than therapeutic target trough concentrations. Likely reasons for this were renal impairment and concurrent treatment with trimethoprim, known to inhibit the renal elimination of lamivudine. CONCLUSIONS: NRTIs could trigger the development of hyperlactatemia in septic patients. The use of NRTI sparing regimens might be considered in the presence of this critical condition.

Dysregulated anti-oxidant signalling and compromised mitochondrial integrity negatively influence regulatory T cell function and viability in liver disease.

BACKGROUND: Dysregulated inflammatory responses and oxidative stress are key pathogenic drivers of chronic inflammatory diseases such as liver cirrhosis (LC). Regulatory T cells (Tregs) are essential to prevent excessive immune activation and maintain tissue homeostasis. While inflammatory cues are well known to modulate the function and stability of Tregs, the extent to which Tregs are influenced by oxidative stress has not been fully explored. METHODS: The phenotypic and functional properties of CD4+CD25+CD127lo/- Tregs isolated from patients with LC were compared to healthy controls (HC). Treg redox state was investigated by characterizing intracellular reactive oxygen species (ROS), NADPH oxidase-2 (Nox2) activity, mitochondrial function, morphology, and nuclear factor-erythroid 2-related factor (Nrf2) antioxidant signalling. The relevance of Nrf2 and its downstream target, Heme-oxygenase-1 (HO-1), in Treg function, stability, and survival, was further assessed using mouse models and CRISPR/Cas9-mediated HO-1 knock-out. FINDINGS: Circulating Tregs from LC patients displayed a reduced suppressive function, correlating with liver disease severity, associated with phenotypic abnormalities and increased apoptosis. Mechanistically, this was linked to a dysregulated Nrf2 signalling with resultant lower levels of HO-1, enhanced Nox2 activation, and impaired mitochondrial respiration and integrity. The functional deficit in LC Tregs could be partially recapitulated by culturing control Tregs in patient sera. INTERPRETATION: Our findings reveal that Tregs rely on functional redox homeostasis for their function, stability, and survival. Targeting Treg specific anti-oxidant pathways may have therapeutic potential to reverse the Treg impairment in conditions of oxidative damage such as advanced liver disease. FUNDING: This study was funded by the Wellcome Trust (211113/A/18/Z).

Epitope-engineered human hematopoietic stem cells are shielded from CD123-targeted immunotherapy.

Targeted eradication of transformed or otherwise dysregulated cells using monoclonal antibodies (mAb), antibody-drug conjugates (ADC), T cell engagers (TCE), or chimeric antigen receptor (CAR) cells is very effective for hematologic diseases. Unlike the breakthrough progress achieved for B cell malignancies, there is a pressing need to find suitable antigens for myeloid malignancies. CD123, the interleukin-3 (IL-3) receptor alpha-chain, is highly expressed in various hematological malignancies, including acute myeloid leukemia (AML). However, shared CD123 expression on healthy hematopoietic stem and progenitor cells (HSPCs) bears the risk for myelotoxicity. We demonstrate that epitope-engineered HSPCs were shielded from CD123-targeted immunotherapy but remained functional, while CD123-deficient HSPCs displayed a competitive disadvantage. Transplantation of genome-edited HSPCs could enable tumor-selective targeted immunotherapy while rebuilding a fully functional hematopoietic system. We envision that this approach is broadly applicable to other targets and cells, could render hitherto undruggable targets accessible to immunotherapy, and will allow continued posttransplant therapy, for instance, to treat minimal residual disease (MRD).

CRISPR/Cas-based Human T cell Engineering: Basic Research and Clinical Application.

Engineering human T cells for the treatment of cancer, viral infections and autoimmunity has been a long-standing dream of many immunologists and hematologists. Although primary human T cells have been genetically engineered for decades, this process was challenging, time consuming and mostly limited to transgene insertions mediated by viral transduction. The absence of widely accessible tools to efficiently and precisely engineer T cells genetically in a targeted manner limited their applicability as a living drug. This fundamentally changed with the discovery of CRISPR/Cas9 and its adaptation to human T cells. CRISPR/Cas9 has made T cell engineering widely accessible and accelerated the development of engineered adoptive T cell therapies. Only 6 years after the discovery of CRISPR/Cas9 as a biotechnological tool the first CRISPR engineered T cells have been administered to patients with refractory cancers in a phase I clinical trial. Novel Cas proteins - natural and engineered ones - are rapidly emerging. These offer for instance increased flexibility, activity and/or specificity. Moreover, sophisticated protein engineering and fusions of Cas with deaminases or reverse transcriptases enable genomic DNA editing without the need for a double strand cut. Thus, the "CRISPR tool box" for experimental use as well as for novel therapeutic approaches is rapidly expanding. In this review, we will summarize the current state of CRISPR/Cas-based engineering in human T cells for basic research and its clinical applications.

Acute kidney injury in patients with COVID-19: a retrospective cohort study from Switzerland.

BACKGROUND: Data about patients in Europe with corona virus disease-2019 (COVID-19) and acute kidney injury (AKI) are scarce. We examined characteristics, presentation and risk factors of AKI in patients hospitalised with COVID-19 in a tertiary hospital in Switzerland. METHODS: We reviewed health records of patients hospitalised with a positive nasopharyngeal polymerase chain reaction test for SARS-CoV2 between 1 February and 30 June 2020, at the University Hospital of Basel. The nadir creatinine of the hospitalisation was used as baseline. AKI was defined according the KDIGO guidelines as a 1.5× increase of baseline creatinine and in-hospital renal recovery as a discharge creatinine <1.25× baseline creatinine. Least absolute shrinkage and selection operator (LASSO) regression was performed to select predictive variables of AKI. Based on this a final model was chosen. RESULTS: Of 188 patients with COVID-19, 41 (22%) developed AKI, and 11 (6%) required renal replacement therapy. AKI developed after a median of 9 days (interquartile range [IQR] 5-12) after the first symptoms and a median of 1 day (IQR 0-5) after hospital admission. The peak AKI stages were stage 1 in 39%, stage 2 in 24% and stage 3 in 37%. A total of 29 (15%) patients were admitted to the intensive care unit and of these 23 (79%) developed AKI. In-hospital renal recovery at discharge was observed in 61% of all AKI episodes. In-hospital mortality was 27% in patients with AKI and 10% in patients without AKI. Age (adjusted odds ratio [aOR] 1.04, 95% confidence interval [CI] 1.01­1.08; p = 0.024), history of chronic kidney disease (aOR 3.47, 95% CI 1.16­10.49;p = 0.026), C-reactive protein levels (aOR 1.09, 95% CI 1.03­1.06; p = 0.002) and creatinine kinase (aOR 1.03, 95% CI 1.01­1.06; p = 0.002) were associated with development of AKI. CONCLUSIONS: AKI is common in hospitalised patients with COVID-19 and more often seen in patients with severe COVID-19 illness. AKI is associated with a high in-hospital mortality.

Venovenous extracorporeal membrane oxygenation to treat hypercapnia in a morbidly obese patient.

Morbid obesity plays an increasingly important role in healthcare. Patients who are severely obese often suffer from a range of medical problems. One problem is obesity-related hypoventilation syndrome with its resulting hypercapnia. We report a case of a 33-year-old female patient who was in an extraordinarily bad medical state, with severe hypercapnia (pCO2 15.1 kPa), sepsis, acute anuric kidney failure and resulting acidosis (pH 6.96). Her body mass index was 84 kg/m2. Her chances of survival were considered very low after failed attempts at noninvasive ventilation. Based on prior research, we refrained from intubation and chose venovenous extracorporeal membrane oxygenation to treat the hypercapnia. In the entire medical literature, we are not aware of a similarly extraordinary case of obesity-related hypoventilation syndrome that could finally be treated successfully. The idea behind this case report is to consider venovenous extracorporeal membrane oxygenation as an alternative to intubation in this patient collective.

Pharmacological treatment options for cryopyrin-associated periodic syndromes.

INTRODUCTION: Cryopyrin-associated periodic syndromes (CAPS) are rare monogenic autoinflammatory diseases, comprising a spectrum of phenotypes of varying severity. CAPS are associated with gain-of-function mutations in the NLRP3 inflammasome, a multiprotein complex critical for the activation of IL-1ß, and are characterized by episodes of fever, urticaria-like rash, musculoskeletal, ocular, and neurological symptoms. Areas covered: Accounting for the pivotal role of IL-1ß in the pathogenesis of CAPS, three therapeutic options, all blocking the action of IL-1ß, are currently approved: anakinra, a recombinant IL-1 receptor antagonist, the IL-1 trap rilonacept and canakinumab, a monoclonal anti-IL-1ß antibody. All agents reduce or even resolve clinical symptoms, biochemical activity markers and improve quality of life in CAPS. This review also covers pharmacokinetic, pharmacodynamic and safety aspects of the approved drugs and the potential utility of IL-1ß blockers in a wide range of other conditions with an autoinflammatory component. Expert commentary: Due to the success story of current pharmaceutics, the therapeutic options in CAPS are not expected to expand in the near future. Prospective observational studies are needed to confirm long-term efficacy and sustained benefit. New IL-1ß blockers will likely address unmet clinical needs in other autoinflammatory conditions.