Ruxolitinib Pharmacokinetics and Pharmacodynamics in Children with Acute and Chronic Graft-versus-Host Disease.

We evaluated the pharmacokinetics (PK) of oral ruxolitinib in children with steroid-refractory acute graft-versus-host disease (aGVHD) (age <12 years) and chronic GVHD (cGVHD) (age ≤18 years) using our published pediatric dosing. PK sampling was performed before and 2 hours after ruxolitinib administration in patients with established cGVHD. More extensive PK analyses were performed in patients with newly diagnosed aGVHD or cGVHD before and .5, 1, 2, 4, and 6 hours after ruxolitinib administration in patients weighing >10 kg and before, 3+, and 6+ hours in children weighing <10 kg. pSTAT1, pSTAT3, and pSTAT5 expression levels were measured on CD4+ and CD8+ T cells before and 2 hours after ruxolitinib administration as a pharmacodynamic marker of JAK/STAT inhibition. Thirteen patients were prospectively enrolled, including 8 with existing cGVHD (age 0 to ≤18 years), 4 with new-onset steroid-refractory aGVHD (age 0 to <12 years) and 1 with newly diagnosed steroid-refractory cGVHD. Great variability in PK was seen. Mean oral clearance (CL/F) was 7.76 ± 4.09 L/h (range, 3.1 to 15.3 L/h). The average elimination half-life was 2.32 ± 1.0 hours. Mean ruxolitinib clearance was higher in children age <2 years versus those age >2 years (12.1 ± 3.0 L/h versus 5.7 ± 2.8 L/h; P = .005) and was reduced with concurrent treatment with azoles and azithromycin. We saw a variable reduction in pSTAT1/3/5 expression on T cells at time of peak ruxolitinib absorption (2 hours after dosing). Children <10 kg had lower ruxolitinib exposure, possibly due to inherent increased drug clearance or variability in dosing methods, leading to decreased drug absorption.

A randomized phase 2 trial of oral vitamin A for graft-versus-host disease in children and young adults.

ABSTRACT: Vitamin A plays a key role in the maintenance of gastrointestinal homeostasis and promotes a tolerogenic phenotype in tissue resident macrophages. We conducted a prospective randomized double-blinded placebo-controlled clinical trial in which 80 recipients of hematopoietic stem cell transplantation (HSCT) were randomized 1:1 to receive pretransplant high-dose vitamin A or placebo. A single oral dose of vitamin A of 4000 IU/kg, maximum 250 000 IU was given before conditioning. The primary end point was incidence of acute graft-versus-host disease (GVHD) at day +100. In an intent-to-treat analysis, incidence of acute GVHD was 12.5% in the vitamin A arm and 20% in the placebo arm (P = .5). Incidence of acute gastrointestinal (GI) GVHD was 2.5% in the vitamin A arm (P = .09) and 12.5% in the placebo arm at day +180. Incidence of chronic GVHD was 5% in the vitamin A arm and 15% in the placebo arm (P = .02) at 1 year. In an "as treated" analysis, cumulative incidence of acute GI GVHD at day +180 was 0% and 12.5% in recipients of vitamin A and placebo, respectively (P = .02), and cumulative incidence of chronic GVHD was 2.7% and 15% in recipients of vitamin A and placebo, respectively (P = .01). The only possibly attributable toxicity was asymptomatic grade 3 hyperbilirubinemia in 1 recipient of vitamin A at day +30, which self-resolved. Absolute CCR9+ CD8+ effector memory T cells, reflecting gut T-cell trafficking, were lower in the vitamin A arm at day +30 after HSCT (P = .01). Levels of serum amyloid A-1, a vitamin A transport protein with proinflammatory effects, were lower in the vitamin A arm. The vitamin A arm had lower interleukin-6 (IL-6), IL-8, and suppressor of tumorigenicity 2 levels and likely a more favorable gut microbiome and short chain fatty acids. Pre-HSCT oral vitamin A is inexpensive, has low toxicity, and reduces GVHD. This trial was registered at www.ClinicalTrials.gov as NCT03202849.

A Pilot Study of Human Milk to Reduce Intestinal Inflammation After Bone Marrow Transplant.

OBJECTIVE: Human milk administration in the early peritransplant period would lower intestinal inflammation after bone marrow transplant (BMT). MATERIALS AND METHODS: Children 0-5 years undergoing BMT received either a ready-to-feed human milk preparation designed for these children (Prolacta Bioscience, Duarte, CA) or standard formula. Babies breastfeeding at the time of BMT were also enrolled on the human milk arm. Human milk was administered from day -3 until day +14 after BMT. Metagenomic shotgun sequencing and metabolomics of stool, plasma cytokines, and regenerating islet-derived 3α (REG3α) levels were measured at enrollment and day +14. Human leukocyte antigen-DR isotype (HLA-DR), CD38, and CD69 expression on T cells were evaluated at day +21. RESULTS: Forty-six children were enrolled, 32 received human milk (donor milk n = 23, breastfeeding babies n = 9), and 14 were controls who received standard feeds supervised by a BMT dietician. Twenty-four patients received at least 60% of goal human milk and were evaluable. Plasma interleukin (IL)-8 (p = 0.04), IL-10 (p = 0.02), and REG3α (p = 0.03) were decreased in the human milk cohort. Peripheral blood CD69+ CD8+ T cells were higher in controls (p = 0.01). Species abundance of Adenovirus (p = 0.00034), Escherichia coli (p = 0.0017), Cryptosporidium parvum (p = 0.0006), Dialister invisus (p = 0.01), and Pseudomonas aeruginosa (p = 0.05) from stool was higher in controls. Stool alanine, tyrosine, methionine, and the ratio of fecal alanine to choline and phosphocholine were higher in controls (p < 0.05). No difference was observed in stool propionate and butyrate levels as measures of short-chain fatty acids between the two cohorts. CONCLUSIONS: Administration of human milk resulted in decreased markers of intestinal inflammation and could be a valuable adjunct for patients after BMT.

A novel strategy for identifying early acute kidney injury in pediatric hematopoietic stem cell transplantation.

Acute kidney injury (AKI) is a common complication in pediatric hematopoietic stem cell transplantation (HSCT). Serum creatinine is an imprecise biomarker of AKI. We hypothesized that combining creatinine with serum cystatin C (cysC) and urinary neutrophil gelatinase-associated lipocalin (NGAL) more effectively characterizes AKI during the first 28 days of HSCT and better identifies patients at risk of adverse outcomes than creatinine alone. We prospectively assessed the type and severity of AKI in 80 consecutive allogeneic HSCT patients using weekly creatinine, cysC, and NGAL. We combined the biomarkers to define 7 Composite Types of AKI, including All Positive AKI (simultaneously detected creatinine, cysC, and NGAL AKI). Outcomes included renal replacement therapy and transplant-related mortality. In all, 75% of patients had AKI by at least one measure; 33% developed >1 type of AKI. Mild AKI often preceded Severe AKI. Patients with creatinine or NGAL AKI that were Severe or Repeated tended to have worse outcomes. The five patients with All Positive AKI had the highest rates of morbidity and mortality. AKI evaluation with creatinine, cysC, and NGAL provides a comprehensive profile of early AKI and narrowly identifies patients at highest risk of adverse outcomes, providing opportunities for early, impactful intervention.

Circulating dsDNA, endothelial injury, and complement activation in thrombotic microangiopathy and GVHD.

Transplant-associated thrombotic microangiopathy (TA-TMA) is a common and poorly recognized complication of hematopoietic stem cell transplantation (HSCT) associated with excessive complement activation, likely triggered by endothelial injury. An important missing piece is the link between endothelial injury and complement activation. We hypothesized that neutrophil extracellular traps (NETs) mechanistically link endothelial damage with complement activation and subsequent TA-TMA. Neutrophil activation releases granule proteins together with double-stranded DNA (dsDNA) to form extracellular fibers known as NETs. NETs have been shown to activate complement and can be assessed in humans by quantification of dsDNA in serum. We measured levels of dsDNA, as a surrogate for NETs in 103 consecutive pediatric allogeneic transplant recipients at day 0, +14, +30, +60, and +100. A spike in dsDNA production around day +14 during engraftment was associated with subsequent TA-TMA development. Peak dsDNA production around day +14 was associated with interleukin-8-driven neutrophil recovery. Increased dsDNA levels at days +30, +60, and +100 were also associated with increased mortality and gastrointestinal graft-versus-host disease (GVHD). NETs may serve as a mechanistic link between endothelial injury and complement activation. NET formation may be one mechanism contributing to the clinical overlap between GVHD and TA-TMA.

Lower levels of vitamin A are associated with increased gastrointestinal graft-versus-host disease in children.

Vitamin A promotes development of mucosal tolerance and enhances differentiation of regulatory T cells. Vitamin A deficiency impairs epithelial integrity, increasing intestinal permeability. We hypothesized that higher vitamin A levels would reduce the risk of graft-versus-host disease (GVHD) through reduced gastrointestinal (GI) permeability, reduced mucosal injury, and reduced lymphocyte homing to the gut. We tested this hypothesis in a cohort study of 114 consecutive patients undergoing allogeneic stem cell transplant. Free vitamin A levels were measured in plasma at day 30 posttransplant. GI GVHD was increased in patients with vitamin A levels below the median (38% vs 12.4% at 100 days, P = .0008), as was treatment-related mortality (17.7% vs 7.4% at 1 year, P = .03). Bloodstream infections were increased in patients with vitamin A levels below the median (24% vs 8% at 1 year, P = .03), supporting our hypothesis of increased intestinal permeability. The GI mucosal intestinal fatty acid-binding protein was decreased after transplant, confirming mucosal injury, but was not correlated with vitamin A levels, indicating that vitamin A did not protect against mucosal injury. Expression of the gut homing receptor CCR9 on T-effector memory cells 30 days after transplant was increased in children with vitamin A levels below the median (r = -0.34, P = .03). Taken together, these data support our hypothesis that low levels of vitamin A actively promote GI GVHD and are not simply a marker of poor nutritional status or a sicker patient. Vitamin A supplementation might improve transplant outcomes.

A Genetic Modifier of the Gut Microbiome Influences the Risk of Graft-versus-Host Disease and Bacteremia After Hematopoietic Stem Cell Transplantation.

The human gut microbiome is involved in vital biological functions, such as maintenance of immune homeostasis and modulation of intestinal development and enhanced metabolic capabilities. Disturbances of the intestinal microbiota have been associated with development and progression of inflammatory conditions, including graft-versus-host disease (GVHD). The fucosyltransferase 2 (FUT2) gene produces an enzyme that is responsible for the synthesis of the H antigen in body fluids and on the intestinal mucosa. FUT2 genotype has been shown to modify the gut microbiome. We hypothesized that FUT2 genotype influences risk of GVHD and bacterial translocation after allogeneic hematopoietic stem cell transplantation (HSCT). FUT2 genotype was determined in 150 consecutive patients receiving allogeneic HSCT at our center. We abstracted clinical characteristics and outcomes from the transplantation database. Cumulative risk of any acute GVHD varied by FUT2 genotype, with decreased risk in those with A/A genotype and increased risk in those with G/G genotype. In contrast, the cumulative incidence of bacteremia was increased in those with A/A genotype. We conclude that the FUT2 genotype influences risk of acute GVHD and bacteremia after HSCT. We hypothesize that the mechanisms involve altered intestinal surface glycosylation and microbial composition but this requires additional study.