Cryopreservation of mesenchymal stem/stromal cells using a DMSO-free solution is comparable to DMSO-containing cryoprotectants: results of an international multicenter PACT/BEST collaborative study.

BACKGROUND AND AIM: An essential aspect of ensuring availability and stability of mesenchymal stem/stromal cells (MSCs) products for clinical use is that these cells are cryopreserved before individual infusion into patients. Currently, cryopreservation of MSCs involves use of a cryoprotectant solution containing dimethyl sulfoxide (DMSO). However, it is recognized that DMSO may be toxic for both the patient and the MSC product. In this Production Assistance for Cellular Therapies (PACT) and Biomedical Excellence for Safer Transfusion (BEST) Collaborative study, we compared a novel DMSO-free solution with DMSO containing cryoprotectant solutions for freezing MSCs. METHODS: A DMSO-free cryoprotectant solution containing sucrose, glycerol, and isoleucine (SGI) in a base of Plasmalyte A was prepared at the University of Minnesota. Cryoprotectant solutions containing 5-10% DMSO (in-house) were prepared at seven participating centers (five from USA, one each from Australia and Germany). The MSCs were isolated from bone marrow or adipose tissue and cultured ex vivo per local protocols at each center. The cells in suspension were frozen by aliquoting into vials/bags. For six out of the seven centers, the vials/bags were placed in a controlled rate freezer (one center placed them at -80°C freezer overnight) before transferring to liquid nitrogen. The cells were kept frozen for at least one week before thawing and testing. Pre- and post-thaw assessment included cell viability and recovery, immunophenotype as well as transcriptional and gene expression profiles. Linear regression, mixed effects models and two-sided t-tests were applied for statistical analysis. RESULTS: MSCs had an average viability of 94.3% (95% CI: 87.2-100%) before cryopreservation, decreasing by 4.5% (95% CI: 0.03-9.0%; P: 0.049) and 11.4% (95% CI: 6.9-15.8%; P< 0.001), for MSCs cryopreserved in the in-house and SGI solutions, respectively. The average recovery of viable MSCs cryopreserved in the SGI was 92.9% (95% CI: 85.7-100.0%), and it was lower by 5.6% (95% CI: 1.3-9.8%, P < 0.013) for the in-house solution. Additionally, MSCs cryopreserved in the two solutions had expected level of expressions for CD45, CD73, CD90, and CD105 with no significant difference in global gene expression profiles. CONCLUSION: MSCs cryopreserved in a DMSO-free solution containing sucrose, glycerol, and isoleucine in a base of Plasmalyte A had slightly lower cell viability, better recovery, and comparable immunophenotype and global gene expression profiles compared to MSCs cryopreserved in DMSO containing solutions. The average viability of MSCs in the novel solution was above 80% and, thus, likely clinically acceptable. Future studies are suggested to test the post-thaw functions of MSCs cryopreserved in the novel DMSO-free solution.

The Prevalence of Pretransplant Frailty and Mental Distress in Hematopoietic Cell Transplantation and Association with Clinical Outcomes.

Frailty is a phenotype of decreased physiologic reserve associated with increased risk of toxicities and nonrelapse mortality (NRM) in hematopoietic cell transplant (HCT) recipients. The incidence, predictors, and adverse effects of pre-HCT frailty are not well known. We evaluated the association of pre-HCT frailty, defined using Fried's criteria, with age and baseline characteristics in patients ≥18 years undergoing autologous (auto) or allogeneic (allo) HCT for hematological malignancies. Assessments were performed as part of routine pre-HCT evaluations and then retrospectively analyzed. We additionally investigated the association of mental health distress indicators with frailty and the association between frailty and transplant outcomes including NRM and overall survival (OS) plus healthcare utilization. Patients undergoing HCT for hematological malignancies were analyzed (total n = 300; 162 auto, 138 allo). The overall prevalence of frailty was 18%, 21.7% among alloHCT, and 14.8% among autoHCT recipients, with similar distributions of frailty domains. Logistic regression analysis of the overall cohort revealed that older age was associated with an increased risk of frailty (odds ratio [OR] 1.37, 95% confidence interval [CI] [1.02-1.82]; P = 0.04). AlloHCT (OR 2.03, CI [1.07-3.84]; P = .03), and patient health questionnaire-9 (PHQ-9) (health depression) score ≥10 (OR 6.28, CI 1.93-20.43; P < .01) were each independently associated with pre-HCT frailty. In alloHCT patients, older age (OR 1.44, CI [1.00-2.06]; P = .05) was the only significant risk factor for pre-HCT frailty, while for autoHCT patients, only a higher PHQ-9 score was associated with frailty (OR 6.43, CI [1.34-30.82]; P = .02). For the whole cohort OS at 1 year was lower in frail recipients at 83% (95% CI, 70-91%) versus 92% (95% CI, 88-95%) in nonfrail (P = .04); with multivariate analysis showing higher risk of death in the frail group (hazard ratio [HR] 2.31, CI 0.97-5.46; P = .06). In the alloHCT cohort, multivariate analysis showed greater 1-year mortality in frail recipients (HR 2.55, CI [0.99-6.56]; P = .053). In the alloHCT recipients, we observed a 1-year NRM of 20% in frail patients versus 9% in nonfrail, and multivariate analysis showed a marginally higher risk of NRM in the frail group (HR 2.70, CI 0.90-8.10; P = .08). Frailty was not associated with higher risk of relapse in alloHCT or autoHCT recipients. Frail alloHCT patients experienced a longer initial hospital stay following HCT compared to nonfrail recipients (P < .01). We observed a high prevalence of pre-HCT frailty across all age groups, and identify older age is a risk factor for frailty, particularly in alloHCT recipients. Frailty is associated with a greater risk of NRM and lower survival which needs investigation in a larger cohort. Frailty associates with greater HCT complexity suggesting a need for early assessments and targeted interventions for this vulnerable population. Our findings suggest the utility of frailty and mental distress screening along with multidisciplinary interventions in pre-HCT to limit the morbidity of HCT.

Amphiregulin as monitoring biomarker for life-threatening acute graft-versushost disease: secondary analysis of two prospective clinical trials.

Not available.

Levels of Urinary Mercapturic Acids of Acrolein, Methacrolein, Crotonaldehyde, and Methyl Vinyl Ketone in Relationship to Chronic Obstructive Pulmonary Disease in Cigarette Smokers of the Subpopulations and Intermediate Outcome Measures in COPD Study (SPIROMICS).

Cigarette smoking is an established cause of chronic obstructive pulmonary disease (COPD). Numerous studies implicate acrolein, which occurs in relatively high concentrations in cigarette smoke and reacts readily with proteins, as one causative factor for COPD in smokers. Far less is known about the possible roles in COPD of the related α,ß-unsaturated carbonyl compounds of cigarette smoke crotonaldehyde, methacrolein, and methyl vinyl ketone. In the study reported here, we analyzed mercapturic acids of these α,ß-unsaturated compounds in the urine of 413 confirmed cigarette smokers in the Subpopulations and Intermediate Outcome Measures in COPD Study (SPIROMICS)─202 with COPD and 211 without COPD. The mercapturic acids analyzed were 3-hydroxypropyl mercapturic acid (3-HPMA) from acrolein, 3-hydroxy-1-methylpropyl mercapturic acid (HMPMA-1) from crotonaldehyde, 3-hydroxy-2-methylpropyl mercapturic acid (HMPMA-2) from methacrolein, and 3-hydroxy-3-methylpropyl mercapturic acid (HMPMA-3) from methyl vinyl ketone. In models adjusting for age, sex, race, pack years of tobacco use, and BMI, all four mercapturic acids were increased in individuals with COPD but not significantly. Stratified by the GOLD status, there were increased levels of the metabolites associated with GOLD 3-4 compared to that with GOLD 0, with the methacrolein metabolite HMPMA-2 reaching statistical significance (adjusted odds ratio 1.23 [95% CI: 1.00-1.53]). These results highlight the possible role of methacrolein, which has previously received little attention in this regard, as a causative factor in COPD in cigarette smokers.

Human Effectors of Acute and Chronic GVHD Overexpress CD83 and Predict Mortality.

PURPOSE: Acute and chronic GVHD remain major causes of transplant-related morbidity and mortality (TRM) after allogeneic hematopoietic cell transplantation (alloHCT). We have shown CD83 chimeric antigen receptor (CAR) T cells prevent GVHD and kill myeloid leukemia cell lines. In this pilot study, we investigate CD83 expression on GVHD effector cells, correlate these discoveries with clinical outcomes, and evaluate critical therapeutic implications for transplant recipients. EXPERIMENTAL DESIGN: CD83 expression was evaluated among circulating CD4+ T cells, B-cell subsets, T follicular helper (Tfh) cells, and monocytes from patients with/without acute or chronic GVHD (n = 48 for each group), respectively. CD83 expression was correlated with survival, TRM, and relapse after alloHCT. Differential effects of GVHD therapies on CD83 expression was determined. RESULTS: CD83 overexpression on CD4+ T cells correlates with reduced survival and increased TRM. Increased CD83+ B cells and Tfh cells, but not monocytes, are associated with poor posttransplant survival. CD83 CAR T eliminate autoreactive CD83+ B cells isolated from patients with chronic GVHD, without B-cell aplasia as observed with CD19 CAR T. We demonstrate robust CD83 antigen density on human acute myeloid leukemia (AML), and confirm potent antileukemic activity of CD83 CAR T in vivo, without observed myeloablation. CONCLUSIONS: CD83 is a promising diagnostic marker of GVHD and warrants further investigation as a therapeutic target of both GVHD and AML relapse after alloHCT.