Pathobiological signatures of dysbiotic lung injury in pediatric patients undergoing stem cell transplantation.

Hematopoietic cell transplantation (HCT) uses cytotoxic chemotherapy and/or radiation followed by intravenous infusion of stem cells to cure malignancies, bone marrow failure and inborn errors of immunity, hemoglobin and metabolism. Lung injury is a known complication of the process, due in part to disruption in the pulmonary microenvironment by insults such as infection, alloreactive inflammation and cellular toxicity. How microorganisms, immunity and the respiratory epithelium interact to contribute to lung injury is uncertain, limiting the development of prevention and treatment strategies. Here we used 278 bronchoalveolar lavage (BAL) fluid samples to study the lung microenvironment in 229 pediatric patients who have undergone HCT treated at 32 children's hospitals between 2014 and 2022. By leveraging paired microbiome and human gene expression data, we identified high-risk BAL compositions associated with in-hospital mortality (P = 0.007). Disadvantageous profiles included bacterial overgrowth with neutrophilic inflammation, microbiome contraction with epithelial fibroproliferation and profound commensal depletion with viral and staphylococcal enrichment, lymphocytic activation and cellular injury, and were replicated in an independent cohort from the Netherlands (P = 0.022). In addition, a broad array of previously occult pathogens was identified, as well as a strong link between antibiotic exposure, commensal bacterial depletion and enrichment of viruses and fungi. Together these lung-immune system-microorganism interactions clarify the important drivers of fatal lung injury in pediatric patients who have undergone HCT. Further investigation is needed to determine how personalized interpretation of heterogeneous pulmonary microenvironments may be used to improve pediatric HCT outcomes.

Tubular Injury Biomarkers to Predict Chronic Kidney Disease and Hypertension at 3-Months Post-Cisplatin in Children.

BACKGROUND: Urine kidney injury biomarkers measured during cisplatin therapy may identify patients at risk for adverse subsequent kidney outcomes. We examined relationships between tubular injury biomarkers collected early (early visit [EV]: first or second cisplatin cycle) and late (late visit [LV]: last or second-last cisplatin cycle) during cisplatin therapy, with 3-month post-cisplatin chronic kidney disease (CKD) and hypertension. METHODS: We analyzed data from the Applying Biomarkers to Minimize Long-Term Effects of Childhood/Adolescent Cancer Treatment Nephrotoxicity Study: twelve-center prospective cohort study of 159 children receiving cisplatin. We measured urine neutrophil gelatinase-associated lipocalin (NGAL)/creatinine, kidney injury molecule-1 (KIM-1)/creatinine, tissue inhibitor of metalloproteinase-2 (TIMP-2), and insulin-like growth factor-binding protein 7 (IGFBP-7) (TIMP-2 and IGFBP-7 expressed as their product, ng/ml^2/1000) at an EV and LV during cisplatin therapy with pre-infusion, post-infusion, and hospital discharge sampling. Area under the curve (AUC) was calculated for biomarkers to detect 3-month post-cisplatin CKD (KDIGO guidelines: low estimated glomerular filtration rate (eGFR) or elevated uACR for age) and hypertension (three blood pressures; per American Academy of Pediatrics guidelines). RESULTS: At median follow-up of 90 days, 52/118 (44%) and 17/125 (14%) developed CKD and hypertension, respectively. Biomarker prediction for 3-month CKD was low to modest; NGAL combined with KIM-1 at EV discharge yielded the highest AUC (0.67, 95% CI 0.57-0.77). Biomarker prediction of 3-month hypertension was stronger, but modest; the highest AUC was from combining EV pre-infusion NGAL and TIMP-2*IGFBP-7 (0.71, 95% CI 0.62-0.80). When EV pre-infusion NGAL and TIMP-2*IGFBP-7 were added to the 3-month hypertension clinical predictive model, AUCs increased from 0.81 (0.72-0.91) to 0.89 (0.83-0.95) (p<0.05). CONCLUSIONS: Tubular injury biomarkers we studied were individually not strong predictors of 3-month post-cisplatin kidney outcomes. Adding biomarkers to existing clinical prediction models may help predict post-therapy hypertension and identify higher kidney-risk patients.

Relevance of lymphocyte proliferation to PHA in severe combined immunodeficiency (SCID) and T cell lymphopenia.

Severe combined immunodeficiency (SCID) is characterized by a severe deficiency in T cell numbers. We analyzed data collected (n = 307) for PHA-based T cell proliferation from the PIDTC SCID protocol 6901, using either a radioactive or flow cytometry method. In comparing the two groups, a smaller number of the patients tested by flow cytometry had <10% of the lower limit of normal proliferation as compared to the radioactive method (p = 0.02). Further, in patients with CD3+ T cell counts between 51 and 300 cells/µL, there was a higher proliferative response with the PHA flow assay compared to the 3H-T assay (p < 0.0001), suggesting that the method of analysis influences the resolution and interpretation of PHA results. Importantly, we observed many SCID patients with profound T cell lymphopenia having normal T cell proliferation when assessed by flow cytometry. We recommend this test be considered only as supportive in the diagnosis of typical SCID.

Pulmonary microbiome and transcriptome signatures reveal distinct pathobiologic states associated with mortality in two cohorts of pediatric stem cell transplant patients.

Lung injury is a major determinant of survival after pediatric hematopoietic cell transplantation (HCT). A deeper understanding of the relationship between pulmonary microbes, immunity, and the lung epithelium is needed to improve outcomes. In this multicenter study, we collected 278 bronchoalveolar lavage (BAL) samples from 229 patients treated at 32 children's hospitals between 2014-2022. Using paired metatranscriptomes and human gene expression data, we identified 4 patient clusters with varying BAL composition. Among those requiring respiratory support prior to sampling, in-hospital mortality varied from 22-60% depending on the cluster (p=0.007). The most common patient subtype, Cluster 1, showed a moderate quantity and high diversity of commensal microbes with robust metabolic activity, low rates of infection, gene expression indicating alveolar macrophage predominance, and low mortality. The second most common cluster showed a very high burden of airway microbes, gene expression enriched for neutrophil signaling, frequent bacterial infections, and moderate mortality. Cluster 3 showed significant depletion of commensal microbes, a loss of biodiversity, gene expression indicative of fibroproliferative pathways, increased viral and fungal pathogens, and high mortality. Finally, Cluster 4 showed profound microbiome depletion with enrichment of Staphylococci and viruses, gene expression driven by lymphocyte activation and cellular injury, and the highest mortality. BAL clusters were modeled with a random forest classifier and reproduced in a geographically distinct validation cohort of 57 patients from The Netherlands, recapitulating similar cluster-based mortality differences (p=0.022). Degree of antibiotic exposure was strongly associated with depletion of BAL microbes and enrichment of fungi. Potential pathogens were parsed from all detected microbes by analyzing each BAL microbe relative to the overall microbiome composition, which yielded increased sensitivity for numerous previously occult pathogens. These findings support personalized interpretation of the pulmonary microenvironment in pediatric HCT, which may facilitate biology-targeted interventions to improve outcomes.

Genotype, oxidase status, and preceding infection or autoinflammation do not affect allogeneic HCT outcomes for CGD.

Chronic granulomatous disease (CGD) is a primary immunodeficiency characterized by life-threatening infections and inflammatory conditions. Hematopoietic cell transplantation (HCT) is the definitive treatment for CGD, but questions remain regarding patient selection and impact of active disease on transplant outcomes. We performed a multi-institutional retrospective and prospective study of 391 patients with CGD treated either conventionally (non-HCT) enrolled from 2004 to 2018 or with HCT from 1996 to 2018. Median follow-up after HCT was 3.7 years with a 3-year overall survival of 82% and event-free survival of 69%. In a multivariate analysis, a Lansky/Karnofsky score <90 and use of HLA-mismatched donors negatively affected survival. Age, genotype, and oxidase status did not affect outcomes. Before HCT, patients had higher infection density, higher frequency of noninfectious lung and liver diseases, and more steroid use than conventionally treated patients; however, these issues did not adversely affect HCT survival. Presence of pre-HCT inflammatory conditions was associated with chronic graft-versus-host disease. Graft failure or receipt of a second HCT occurred in 17.6% of the patients and was associated with melphalan-based conditioning and/or early mixed chimerism. At 3 to 5 years after HCT, patients had improved growth and nutrition, resolved infections and inflammatory disease, and lower rates of antimicrobial prophylaxis or corticosteroid use compared with both their baseline and those of conventionally treated patients. HCT leads to durable resolution of CGD symptoms and lowers the burden of the disease. Patients with active infection or inflammation are candidates for transplants; HCT should be considered before the development of comorbidities that could affect performance status. This trial was registered at www.clinicaltrials.gov as #NCT02082353.

Updates in hematopoietic cell transplant and cellular therapies that enhance the risk for opportunistic infections.

BACKGROUND: Infectious disease physicians may be asked to evaluate and manage a variety of infections in immunocompromised hosts undergoing hematopoietic cell transplant (HCT) and cellular therapies. Over the last decade, several advances in cellular therapy have occurred, with implications for the types of infectious complications that may be seen. AIMS: The purpose of this review is to update the infectious disease physician on newer advances in HCT and cellular therapy, including haploidentical transplant, expanding indications for transplant in older individuals and children, and chimeric antigen receptor T-cells. We will review how these advances might influence infectious disease complications following HCT. We will also provide a perspective that infectious disease physicians can use to evaluate the degree of immune suppression in an individual patient to help determine the type of infections that may be encountered.

Measuring the effect of newborn screening on survival after haematopoietic cell transplantation for severe combined immunodeficiency: a 36-year longitudinal study from the Primary Immune Deficiency Treatment Consortium.

BACKGROUND: Severe combined immunodeficiency (SCID) is fatal unless durable adaptive immunity is established, most commonly through allogeneic haematopoietic cell transplantation (HCT). The Primary Immune Deficiency Treatment Consortium (PIDTC) explored factors affecting the survival of individuals with SCID over almost four decades, focusing on the effects of population-based newborn screening for SCID that was initiated in 2008 and expanded during 2010-18. METHODS: We analysed transplantation-related data from children with SCID treated at 34 PIDTC sites in the USA and Canada, using the calendar time intervals 1982-89, 1990-99, 2000-09, and 2010-18. Categorical variables were compared by χ2 test and continuous outcomes by the Kruskal-Wallis test. Overall survival was estimated by the Kaplan-Meier method. A multivariable analysis using Cox proportional hazards regression models examined risk factors for HCT outcomes, including the variables of time interval of HCT, infection status and age at HCT, trigger for diagnosis, SCID type and genotype, race and ethnicity of the patient, non-HLA-matched sibling donor type, graft type, GVHD prophylaxis, and conditioning intensity. FINDINGS: For 902 children with confirmed SCID, 5-year overall survival remained unchanged at 72%-73% for 28 years until 2010-18, when it increased to 87% (95% CI 82·1-90·6; n=268; p=0·0005). For children identified as having SCID by newborn screening since 2010, 5-year overall survival was 92·5% (95% CI 85·8-96·1), better than that of children identified by clinical illness or family history in the same interval (79·9% [69·5-87·0] and 85·4% [71·8-92·8], respectively [p=0·043]). Multivariable analysis demonstrated that the factors of active infection (hazard ratio [HR] 2·41, 95% CI 1·56-3·72; p<0·0001), age 3·5 months or older at HCT (2·12, 1·38-3·24; p=0·001), Black or African-American race (2·33, 1·56-3·46; p<0·0001), and certain SCID genotypes to be associated with lower overall survival during all time intervals. Moreover, after adjusting for several factors in this multivariable analysis, HCT after 2010 no longer conveyed a survival advantage over earlier time intervals studied (HR 0·73, 95% CI 0·43-1·26; p=0·097). This indicated that younger age and freedom from infections at HCT, both directly driven by newborn screening, were the main drivers for recent improvement in overall survival. INTERPRETATION: Population-based newborn screening has facilitated the identification of infants with SCID early in life, in turn leading to prompt HCT while avoiding infections. Public health programmes worldwide can benefit from this definitive demonstration of the value of newborn screening for SCID. FUNDING: National Institute of Allergy and Infectious Diseases, Office of Rare Diseases Research, and National Center for Advancing Translational Sciences.

Circulating cell-free mitochondrial DNA as a diagnostic and prognostic biomarker in chronic and late acute graft-versus-host disease in children.

In an earlier study, we found that mitochondrial DNA (mtDNA) concentration is elevated in adults with chronic graft-versus-host disease (cGvHD), acting as an endogenous source of TLR9 agonists to augment B-cell responses. To validate this in children, we evaluated mtDNA plasma expression in a large pediatric cohort (ABLE/PBMTC 1202 study). Plasma cell-free mtDNA (cf-mtDNA) copy numbers were measured in 202 pediatric patients using quantitative Droplet Digital polymerase chain reaction (ddPCR). Two evaluations were performed: 1) before the onset of cGvHD or late acute graft-versus-host disease (aGvHD) at day 100 ± 14 days and 2) at the time of cGvHD onset compared with time-matched non-cGvHD controls. We found that cf-mtDNA copy numbers were not affected by immune reconstitution post-hematopoietic stem cell transplantation but were higher on day 100 before the onset of late aGvHD and at the onset of cGvHD. We found that cf-mtDNA was not impacted by previous aGvHD, but correlated with the early onset, NIH moderate/severe cGvHD, and did not correlate with other immune cell populations, cytokines, or chemokines but did with the metabolites spermine and taurine. Similar to adults, children have elevated plasma cf-mtDNA concentrations at the early onset of cGvHD, especially in NIH moderate/severe cGvHD, elevation with late aGvHD, and associated with metabolites involved in mitochondrial function.

Updated Management Guidelines for Adenosine Deaminase Deficiency.

Inherited defects in the adenosine deaminase (ADA) gene typically cause severe combined immunodeficiency. In addition to infections, ADA-deficient patients can present with neurodevelopmental, behavioral, hearing, skeletal, lung, heart, skin, kidney, urogenital, and liver abnormalities. Some patients also suffer from autoimmunity and malignancies. In recent years, there have been remarkable advances in the management of ADA deficiency. Most ADA-deficient patients can be identified by newborn screening for severe combined immunodeficiency, which facilitates early diagnosis and treatment of asymptomatic infants. Most patients benefit from enzyme replacement therapy (ERT). Allogeneic hematopoietic cell transplantation from an HLA-matched sibling donor or HLA-matched family member donor with no conditioning is currently the preferable treatment. When matched sibling donor or matched family member donor is not available, autologous ADA gene therapy with nonmyeloablative conditioning and ERT withdrawal, which is reported in recent studies to result in 100% overall survival and 90% to 95% engraftment, should be pursued. If gene therapy is not immediately available, ERT can be continued for a few years, although its excessive cost might be prohibitive. The recent improved outcome of hematopoietic cell transplantation using HLA-mismatched family-related donors or HLA-matched unrelated donors, after reduced-intensity conditioning, suggests that such procedures might also be considered rather than continuing ERT for prolonged periods. Long-term follow-up will further assist in determining the optimal treatment approach for ADA-deficient patients.

Elevated ADA2 Enzyme Activity at the Onset of Chronic Graft-versus-Host Disease in Children.

Adenosinergic signaling has potent, context-specific effects on immune cells, particularly on the dysregulation of lymphocytes. This in turn may have a role in immune activation and loss of tolerance in such diseases as chronic graft-versus-host disease (chronic GVHD). We assessed whether changes in the enzymatic activity of adenosine deaminase 2 (ADA2), an enzyme that depletes adenosine in the extracellular space via conversion to inosine, may be associated with the onset of chronic GVHD. ADA2-specific enzyme activity was measured in plasma samples from 230 pediatric hematopoietic stem cell transplantation (HSCT) recipients enrolled on the Applied Biomarkers of Late Effects of Childhood Cancer (ABLE)/Pediatric Blood and Marrow Transplant Consortium (PBMTC) 1202 study and compared between patients developing chronic GVHD and those not developing chronic GVHD within 12 months of transplantation. ADA2 and its relationships with 219 previously measured plasma-soluble proteins, metabolites, and immune cell populations were evaluated as well. Plasma ADA2 enzyme activity was significantly elevated in pediatric HSCT recipients at the onset of chronic GVHD compared to patients without chronic GVHD and was not associated with prior history of acute GVHD or generalized inflammation as measured by C-reactive protein concentration. ADA2-specific enzyme activity met our criteria as a potential diagnostic biomarker of chronic GVHD (effect ratio ≥1.30 or ≤.75; area under the receiver operating characteristic curve ≥.60; P < .05) and was positively associated with markers of immune activation previously identified in pediatric chronic GVHD patients. These results support the potential of ADA2 enzyme activity, in combination with other biomarkers and subject to future validation, to aid the diagnosis of chronic GVHD in children post-HSCT.

A diagnostic classifier for pediatric chronic graft-versus-host disease: results of the ABLE/PBMTC 1202 study.

The National Institutes of Health Consensus criteria for chronic graft-versus-host disease (cGVHD) diagnosis can be challenging to apply in children, making pediatric cGVHD diagnosis difficult. We aimed to identify diagnostic pediatric cGVHD biomarkers that would complement the current clinical criteria and help differentiate cGVHD from non-cGVHD. The Applied Biomarkers of Late Effects of Childhood Cancer (ABLE) study, open at 27 transplant centers, prospectively evaluated 302 pediatric patients after hematopoietic cell transplant (234 evaluable). Forty-four patients developed cGVHD. Mixed and fixed effect regression analyses were performed on diagnostic cGVHD onset blood samples for cellular and plasma biomarkers, with individual markers declared relevant if they met 3 criteria: an effect ratio ≥1.3 or ≤0.75; an area under the curve (AUC) of ≥0.60; and a P value <5.814 × 10-4 (Bonferroni correction) (mixed effect) or <.05 (fixed effect). To address the complexity of cGVHD diagnosis in children, we built a machine learning-based classifier that combined multiple cellular and plasma biomarkers with clinical factors. Decreases in regulatory natural killer cells, naïve CD4 T helper cells, and naïve regulatory T cells, and elevated levels of CXCL9, CXCL10, CXCL11, ST2, ICAM-1, and soluble CD13 (sCD13) characterize the onset of cGVHD. Evaluation of the time dependence revealed that sCD13, ST2, and ICAM-1 levels varied with the timing of cGVHD onset. The cGVHD diagnostic classifier achieved an AUC of 0.89, with a positive predictive value of 82% and a negative predictive value of 80% for diagnosing cGVHD. Our polyomic approach to building a diagnostic classifier could help improve the diagnosis of cGVHD in children but requires validation in future prospective studies. This trial was registered at www.clinicaltrials.gov as #NCT02067832.

Acute kidney injury during cisplatin therapy and associations with kidney outcomes 2 to 6 months post-cisplatin in children: a multi-centre, prospective observational study.

BACKGROUND: Few studies describe acute kidney injury (AKI) burden during paediatric cisplatin therapy and post-cisplatin kidney outcomes. We determined risk factors for and rate of (1) AKI during cisplatin therapy, (2) chronic kidney disease (CKD) and hypertension 2-6 months post-cisplatin, and (3) whether AKI is associated with 2-6-month outcomes. METHODS: This prospective cohort study enrolled children (aged < 18 years at cancer diagnosis) treated with cisplatin from twelve Canadian hospitals. AKI during cisplatin therapy (primary exposure) was defined based on Kidney Disease: Improving Global Outcomes (KDIGO) serum creatinine criteria (≥ stage one). Severe electrolyte abnormalities (secondary exposure) included ≥ grade three hypophosphatemia, hypokalemia, or hypomagnesemia (National Cancer Institute Common Terminology Criteria for Adverse Events v4.0). CKD was albuminuria or decreased kidney function for age (KDIGO guidelines). Hypertension was defined based on the 2017 American Academy of Pediatrics guidelines. RESULTS: Of 159 children (median [interquartile range [IQR]] age: 6 [2-12] years), 73/159 (46%) participants developed AKI and 55/159 (35%) experienced severe electrolyte abnormalities during cisplatin therapy. At median [IQR] 90 [76-110] days post-cisplatin, 53/119 (45%) had CKD and 18/128 (14%) developed hypertension. In multivariable analyses, AKI was not associated with 2-6-month CKD or hypertension. Severe electrolyte abnormalities during cisplatin were associated with having 2-6-month CKD or hypertension (adjusted odds ratio (AdjOR) [95% CI]: 2.65 [1.04-6.74]). Having both AKI and severe electrolyte abnormalities was associated with 2-6-month hypertension (AdjOR [95% CI]: 3.64 [1.05-12.62]). CONCLUSIONS: Severe electrolyte abnormalities were associated with kidney outcomes. Cisplatin dose optimization to reduce toxicity and clear post-cisplatin kidney follow-up guidelines are needed. A higher resolution version of the Graphical abstract is available as Supplementary information.

The diagnosis of severe combined immunodeficiency: Implementation of the PIDTC 2022 Definitions.

BACKGROUND: Shearer et al in 2014 articulated well-defined criteria for the diagnosis and classification of severe combined immunodeficiency (SCID) as part of the Primary Immune Deficiency Treatment Consortium's (PIDTC's) prospective and retrospective studies of SCID. OBJECTIVE: Because of the advent of newborn screening for SCID and expanded availability of genetic sequencing, revision of the PIDTC 2014 Criteria was needed. METHODS: We developed and tested updated PIDTC 2022 SCID Definitions by analyzing 379 patients proposed for prospective enrollment into Protocol 6901, focusing on the ability to distinguish patients with various SCID subtypes. RESULTS: According to PIDTC 2022 Definitions, 18 of 353 patients eligible per 2014 Criteria were considered not to have SCID, whereas 11 of 26 patients ineligible per 2014 Criteria were determined to have SCID. Of note, very low numbers of autologous T cells (<0.05 × 109/L) characterized typical SCID under the 2022 Definitions. Pathogenic variant(s) in SCID-associated genes was identified in 93% of patients, with 7 genes (IL2RG, RAG1, ADA, IL7R, DCLRE1C, JAK3, and RAG2) accounting for 89% of typical SCID. Three genotypes (RAG1, ADA, and RMRP) accounted for 57% of cases of leaky/atypical SCID; there were 13 other rare genotypes. Patients with leaky/atypical SCID were more likely to be diagnosed at more than age 1 year than those with typical SCID lacking maternal T cells: 20% versus 1% (P < .001). Although repeat testing proved important, an initial CD3 T-cell count of less than 0.05 × 109/L differentiated cases of typical SCID lacking maternal cells from leaky/atypical SCID: 97% versus 7% (P < .001). CONCLUSIONS: The PIDTC 2022 Definitions describe SCID and its subtypes more precisely than before, facilitating analyses of SCID characteristics and outcomes.

The diagnosis of severe combined immunodeficiency (SCID): The Primary Immune Deficiency Treatment Consortium (PIDTC) 2022 Definitions.

Severe combined immunodeficiency (SCID) results from defects in the differentiation of hematopoietic stem cells into mature T lymphocytes, with additional lymphoid lineages affected in particular genotypes. In 2014, the Primary Immune Deficiency Treatment Consortium published criteria for diagnosing SCID, which are now revised to incorporate contemporary approaches. Patients with typical SCID must have less than 0.05 × 109 autologous T cells/L on repetitive testing, with either pathogenic variant(s) in a SCID-associated gene, very low/undetectable T-cell receptor excision circles or less than 20% of CD4 T cells expressing naive markers, and/or transplacental maternally engrafted T cells. Patients with less profoundly impaired autologous T-cell differentiation are designated as having leaky/atypical SCID, with 2 or more of these: low T-cell numbers, oligoclonal T cells, low T-cell receptor excision circles, and less than 20% of CD4 T cells expressing naive markers. These patients must also have either pathogenic variant(s) in a SCID-associated gene or reduced T-cell proliferation to certain mitogens. Omenn syndrome requires a generalized erythematous rash, absent transplacentally acquired maternal engraftment, and 2 or more of these: eosinophilia, elevated IgE, lymphadenopathy, hepatosplenomegaly. Thymic stromal defects and other causes of secondary T-cell deficiency are excluded from the definition of SCID. Application of these revised Primary Immune Deficiency Treatment Consortium 2022 Definitions permits precise categorization of patients with T-cell defects but does not imply a preferred treatment strategy.

Aberrant T-cell exhaustion in severe combined immunodeficiency survivors with poor T-cell reconstitution after transplantation.

BACKGROUND: Severe combined immunodeficiency (SCID) comprises rare inherited disorders of immunity that require definitive treatment through hematopoietic cell transplantation (HCT) or gene therapy for survival. Despite successes of allogeneic HCT, many SCID patients experience incomplete immune reconstitution, persistent T-cell lymphopenia, and poor long-term outcomes. OBJECTIVE: We hypothesized that CD4+ T-cell lymphopenia could be associated with a state of T-cell exhaustion in previously transplanted SCID patients. METHODS: We analyzed markers of exhaustion in blood samples from 61 SCID patients at a median of 10.4 years after HCT. RESULTS: Compared to post-HCT SCID patients with normal CD4+ T-cell counts, those with poor T-cell reconstitution showed lower frequency of naive CD45RA+/CCR7+ T cells, recent thymic emigrants, and TCR excision circles. They also had a restricted TCR repertoire, increased expression of inhibitory receptors (PD-1, 2B4, CD160, BTLA, CTLA-4), and increased activation markers (HLA-DR, perforin) on their total and naive CD8+ T cells, suggesting T-cell exhaustion and aberrant activation, respectively. The exhaustion score of CD8+ T cells was inversely correlated with CD4+ T-cell count, recent thymic emigrants, TCR excision circles, and TCR diversity. Exhaustion scores were higher among recipients of unconditioned HCT, especially when further in time from HCT. Patients with fewer CD4+ T cells showed a transcriptional signature of exhaustion. CONCLUSIONS: Recipients of unconditioned HCT for SCID may develop late post-HCT T-cell exhaustion as a result of diminished production of T-lineage cells. Elevated expression of inhibitory receptors on their T cells may be a biomarker of poor long-term T-cell reconstitution.

CD56bright CD16- natural killer cells as an important regulatory mechanism in chronic graft-versus-host disease.

Chronic graft-versus-host disease (cGvHD) is a major cause of morbidity after hematopoietic stem cell transplantation (HSCT). In large patient populations, we have shown a CD56bright natural killer (NK) population to strongly associate with a lack of cGvHD and we hypothesize that these cells function to suppress cGvHD. We aimed to isolate and define the characteristics of regulatory NK (NKreg) cells associated with suppression of cGvHD. Immunophenotypic evaluation of a large pediatric population found the CD56bright NK population associated with a lack of cGvHD to be perforin-, Granzyme B-, and CD335+. Transcriptome analysis of a small patient cohort of CD56bright compared to CD56dim NK cells found the NKreg cells to also overexpress Granzyme K, IL-7R, GPR183, RANK, GM-CSFR, TCF7, and IL23A. Further analysis of this CD56bright NKreg population found a subpopulation that overexpressed IRF1, and TNF. We also found that viable NKreg cells may be isolated by sorting on CD56+ and CD16- NK cells, and this population can suppress allogeneic CD4+ T cells, but not Treg cells or CD8+ T cells through a non-cytolytic, cell-cell contact dependent mechanism. Suppression was not reliant upon the NKp44, NKp46, or GPR183 receptors. Additionally, NKreg cells do not kill leukemic cells. Moreover, this is the first paper to clearly establish that a CD56brightCD3-CD16-perforin- NKreg population associates with a lack of cGvHD and has several unique characteristics, including the suppression of helper T-cell function in vitro. With further investigation we may decipher the mechanism of NKreg suppression and operationalize expansion of NKreg cells associated with cGvHD suppression.

Toward a Better Understanding of the Atypical Features of Chronic Graft-Versus-Host Disease: A Report from the 2020 National Institutes of Health Consensus Project Task Force.

Alloreactive and autoimmune responses after allogeneic hematopoietic cell transplantation can occur in nonclassical chronic graft-versus-host disease (chronic GVHD) tissues and organ systems or manifest in atypical ways in classical organs commonly affected by chronic GVHD. The National Institutes of Health (NIH) consensus projects were developed to improve understanding and classification of the clinical features and diagnostic criteria for chronic GVHD. Although still speculative whether atypical manifestations are entirely due to chronic GVHD, these manifestations remain poorly captured by the current NIH consensus project criteria. Examples include chronic GVHD impacting the hematopoietic system as immune mediated cytopenias, endothelial dysfunction, or as atypical features in the musculoskeletal system, central and peripheral nervous system, kidneys, and serous membranes. These purported chronic GVHD features may contribute significantly to patient morbidity and mortality. Most of the atypical chronic GVHD features have received little study, particularly within multi-institutional and prospective studies, limiting our understanding of their frequency, pathogenesis, and relation to chronic GVHD. This NIH consensus project task force report provides an update on what is known and not known about the atypical manifestations of chronic GVHD while outlining a research framework for future studies to be undertaken within the next 3 to 7 years. We also provide provisional diagnostic criteria for each atypical manifestation, along with practical investigation strategies for clinicians managing patients with atypical chronic GVHD features.

Outcomes following treatment for ADA-deficient severe combined immunodeficiency: a report from the PIDTC.

Adenosine deaminase (ADA) deficiency causes ∼13% of cases of severe combined immune deficiency (SCID). Treatments include enzyme replacement therapy (ERT), hematopoietic cell transplant (HCT), and gene therapy (GT). We evaluated 131 patients with ADA-SCID diagnosed between 1982 and 2017 who were enrolled in the Primary Immune Deficiency Treatment Consortium SCID studies. Baseline clinical, immunologic, genetic characteristics, and treatment outcomes were analyzed. First definitive cellular therapy (FDCT) included 56 receiving HCT without preceding ERT (HCT); 31 HCT preceded by ERT (ERT-HCT); and 33 GT preceded by ERT (ERT-GT). Five-year event-free survival (EFS, alive, no need for further ERT or cellular therapy) was 49.5% (HCT), 73% (ERT-HCT), and 75.3% (ERT-GT; P < .01). Overall survival (OS) at 5 years after FDCT was 72.5% (HCT), 79.6% (ERT-HCT), and 100% (ERT-GT; P = .01). Five-year OS was superior for patients undergoing HCT at <3.5 months of age (91.6% vs 68% if ≥3.5 months, P = .02). Active infection at the time of HCT (regardless of ERT) decreased 5-year EFS (33.1% vs 68.2%, P < .01) and OS (64.7% vs 82.3%, P = .02). Five-year EFS (90.5%) and OS (100%) were best for matched sibling and matched family donors (MSD/MFD). For patients treated after the year 2000 and without active infection at the time of FDCT, no difference in 5-year EFS or OS was found between HCT using a variety of transplant approaches and ERT-GT. This suggests alternative donor HCT may be considered when MSD/MFD HCT and GT are not available, particularly when newborn screening identifies patients with ADA-SCID soon after birth and before the onset of infections. This trial was registered at www.clinicaltrials.gov as #NCT01186913 and #NCT01346150.

Urine Neutrophil Gelatinase-Associated Lipocalin and Kidney Injury Molecule-1 to Detect Pediatric Cisplatin-Associated Acute Kidney Injury.

Background: Few studies have described associations between the AKI biomarkers urinary neutrophil gelatinase-associated lipocalin (NGAL) and kidney injury molecule-1 (KIM-1) with AKI in cisplatin-treated children. We aimed to describe excretion patterns of urine NGAL and KIM-1 and associations with AKI in children receiving cisplatin. Methods: Participants (n=159) were enrolled between 2013 and 2017 in a prospective cohort study conducted in 12 Canadian pediatric hospitals. Participants were evaluated at early cisplatin infusions (at first or second cisplatin cycle) and late cisplatin infusions (last or second-to-last cycle). Urine NGAL and KIM-1 were measured (1) pre-cisplatin infusion, (2) post-infusion (morning after), and (3) at hospital discharge at early and late cisplatin infusions. Primary outcome: AKI defined by serum creatinine rise within 10 days post-cisplatin, on the basis of Kidney Disease Improving Global Outcomes guidelines criteria (stage 1 or higher). Results: Of 159 children, 156 (median [interquartile range (IQR)] age: 5.8 [2.4-12.0] years; 78 [50%] female) had biomarker data available at early cisplatin infusions and 127 had data at late infusions. Forty six of the 156 (29%) and 22 of the 127 (17%) children developed AKI within 10 days of cisplatin administration after early and late infusions, respectively. Urine NGAL and KIM-1 concentrations were significantly higher in patients with versus without AKI (near hospital discharge of late cisplatin infusion, median [IQR] NGAL levels were 76.1 [10.0-232.7] versus 14.9 [5.4-29.7] ng/mg creatinine; KIM-1 levels were 4415 [2083-9077] versus 1049 [358-3326] pg/mg creatinine; P<0.01). These markers modestly discriminated for AKI (area under receiver operating characteristic curve [AUC-ROC] range: NGAL, 0.56-0.72; KIM-1, 0.48-0.75). Biomarker concentrations were higher and better discriminated for AKI at late cisplatin infusions (AUC-ROC range, 0.54-0.75) versus early infusions (AUC-ROC range, 0.48-0.65). Conclusions: Urine NGAL and KIM-1 were modest at discriminating for cisplatin-associated AKI. Further research is needed to determine clinical utility and applicability of these markers and associations with late kidney outcomes.

Is It Possible to Separate the Graft-Versus-Leukemia (GVL) Effect Against B Cell Acute Lymphoblastic Leukemia From Graft-Versus-Host Disease (GVHD) After Hematopoietic Cell Transplant?

Hematopoietic cell transplant is a curative therapy for many pediatric patients with high risk acute lymphoblastic leukemia. Its therapeutic mechanism is primarily based on the generation of an alloreactive graft-versus-leukemia effect that can eliminate residual leukemia cells thus preventing relapse. However its efficacy is diminished by the concurrent emergence of harmful graft-versus-host disease disease which affects healthly tissue leading to significant morbidity and mortality. The purpose of this review is to describe the interventions that have been trialed in order to augment the beneficial graft-versus leukemia effect post-hematopoietic cell transplant while limiting the harmful consequences of graft-versus-host disease. This includes many emerging and promising strategies such as ex vivo and in vivo graft manipulation, targeted cell therapies, T-cell engagers and multiple pharmacologic interventions that stimulate specific donor effector cells.