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Gemtuzumab-ozogamicin (GO) is a humanized anti-CD33 antibody, which is conjugated to a cytotoxic calicheamicin. It is used to treat acute myeloid leukemia (AML) in combination with chemotherapy. We describe here two GO-treated acute myeloid leukemia (AML) cases: both patients suffered from a toxic syndrome, which manifested as impaired hemoglobin-haptoglobin scavenging and accumulation of hemolysis-related products. Our observations and earlier reports indicated that the reaction was caused by GO-targeted destruction of CD33 + CD163+ monocytes/macrophages, which are responsible for the clearance of hemoglobin-haptoglobin complexes. The rise of plasma lactate dehydrogenase was an early sign of the reaction, and both patients had high levels of free plasma hemoglobin, but plasma haptoglobin and bilirubin levels were paradoxically normal. Symptoms included septic fever and abnormalities in cardiac tests and in the case of the first patient, severe neurological symptoms which required intensive care unit admittance. Therapeutic plasma exchanges supported the patients until the recovery of normal hematopoiesis. The symptoms may be easily confounded with infectious complications-related organ damage. Regarding the increasing use of gemtuzumab-ozogamicin and other emerging CD33-targeted cell therapies, we want to highlight this mostly unknown and probably underdiagnosed toxicity.

Common variable immunodeficiency and other late-onset immunodeficiencies often co-manifest with autoimmunity and lymphoproliferation. The pathogenesis of most cases is elusive, as only a minor subset harbors known monogenic germline causes. The involvement of both B and T cells is however implicated. To study whether somatic mutations in CD4+ and CD8+ T cells associate with immunodeficiency, we recruited 17 patients and 21 healthy controls. Eight patients had late-onset common variable immunodeficiency and nine patients other immunodeficiency and/or severe autoimmunity. In total, autoimmunity occurred in 94% and lymphoproliferation in 65%. We performed deep sequencing of 2533 immune-associated genes from CD4+ and CD8+ cells. Deep T-cell receptor beta sequencing was used to characterize CD4+ and CD8+ T-cell receptor repertoires. The prevalence of somatic mutations was 65% in all immunodeficiency patients, 75% in common variable immunodeficiency and 48% in controls. Clonal hematopoiesis-associated variants in both CD4+ and CD8+ cells occurred in 24% of immunodeficiency patients. Results demonstrated mutations in known tumor suppressors, oncogenes, and genes that are critical for immune- and proliferative functions, such as STAT5B (two patients), C5AR1 (two patients), KRAS (one patient), and NOD2 (one patient). Additionally, as a marker of T-cell receptor repertoire perturbation, common variable immunodeficiency patients harbored increased frequencies of clones with identical complementarity determining region 3 sequences despite unique nucleotide sequences when compared to controls. In conclusion, somatic mutations in genes implicated for autoimmunity and lymphoproliferation are common in CD4+ and CD8+ cells of patients with immunodeficiency. They may contribute to immune dysregulation in a subset of immunodeficiency patients.

Graft-versus-host-disease (GvHD) is the main complication of allogeneic hematopoietic stem cell transplantation. GvHD patients have aberrant T cell expansions, which are thought to drive pathological immune activation. Here we report mechanistic insights that somatic mutations may account for persistent clonal T cell expansions in chronic GvHD (cGvHD). In an index patient suffering from cGVHD, we discovered persisting somatic MTOR, NFKB2, and TLR2 mutations in an expanded CD4+ T clone. In the screening cohort (n=135), the MTOR P2229R kinase domain mutation was detected in two additional cGvHD patients, but not in controls. Functional analysis of the discovered MTOR mutation indicated a gain-of-function alteration in translational regulation yielding in up-regulation of phosphorylated S6K1, S6, and AKT. Paired single-cell RNA and T cell receptor alpha and beta sequencing strongly supported cytotoxicity and abnormal proliferation of the clonally expanded CD4+ T cells. Real-time impedance measurements indicated increased cytotoxicity of mutated CD4 + T cells against the patient’s fibroblasts. High throughput drug-sensitivity testing suggested that mutations induce resistance to mTOR inhibitors but increase sensitivity for HSP90 inhibitors. Our findings suggest a novel explanation for the aberrant, persistent T cell activation in cGvHD, and pave the way for novel targeted therapies.

BACKGROUND: Obliterative bronchiolitis after lung transplantation is characterized by chronic airway inflammation leading to the obliteration of small airways. Hypoxia-inducible factor-1 (HIF-1) is a master regulator of cellular responses to hypoxia and inflammation. The Von Hippel-Lindau protein (pVHL) drives the degradation of oxygen-sensitive subunit HIF-1 alpha that controls the activity of HIF-1. We investigated the effect of myeloid cell targeted gene deletion of HIF-1 alpha or its negative regulator pVHL on the development of obliterative airway disease (OAD) in the recipients of tracheal allografts, a mouse model for obliterative bronchiolitis after lung transplantation. METHODS: Tracheal allografts were heterotopically transplanted from BALB/c donor mice to fully major histocompatibility complex mismatched recipient mice with HIF-1 alpha or VHL gene deletion in myeloid cells. The recipients were left non-immunosuppressed or received tacrolimus daily. Histologic, immunohistochemical, and real-time reverse transcription polymerase chain reaction analyses were performed at 3, 10, and 30 days. RESULTS: In the absence of immunosuppression, myeloid cell-specific VHL deficiency of the recipient mice improved epithelial recovery, decreased inflammatory cell infiltration and expression of pro-inflammatory cytokines, increased regulatory forkhead box P3 messenger RNA expression, and reduced OAD development in tracheal allografts. In the presence of tacrolimus immunosuppression, loss of HIF-1 alpha activity in myeloid cells of the recipient by HIF-1 alpha gene deletion accelerated OAD development in mouse tracheal allografts. CONCLUSIONS: Activity of the HIF-pathway affects the development of allograft rejection, and our results suggest that myeloid cell-specific VHL-deficiency that potentially increases HIF-activity decreases allograft inflammation and the subsequent development of OAD in mouse tracheal allografts. (C) 2016 International Society for Heart and Lung Transplantation. All rights reserved.