Decrease in T-Lymphocyte Mitochondrial DNA Copy Number Is Associated with Acute Graft Versus Host Disease

Abstract

Introduction Mitochondria play an important role in T-lymphocyte activation and initiation of the inflammatory response, two pathways that are important in the pathogenesis of acute graft-versus-host disease (aGvHD). We evaluated whether change in mitochondrial DNA (mtDNA) copy number in CD3+ T-lymphocytes measured at two time-points pre and post aGvHD could be used as a biomarker for aGvHD diagnosis that correlates with clinical and/or biopsy proven aGvHD. Methods 99 patients who underwent an allogeneic hematopoietic cell transplant (allo-HCT) between March 2012 and December 2016, were diagnosed with aGvHD and had samples available before and after aGvHD diagnosis were selected for this analysis. We measured mtDNA copy number in CD3+ T-lymphocytes at two time points using real time quantitative polymerase chain reaction (PCR); once prior to diagnosis of aGvHD (median (Interquartile range): 11 (5 – 22) days prior to aGvHD diagnosis) and once after aGvHD diagnosis (median: 23 (12-34) days after aGvHD diagnosis) in 99 patients. 16% (n=16) of the patients had grade I aGvHD and 84% (n=83) had grade II-IV aGvHD. We used linear mixed models with random subject and batch effects to evaluate the change in mtDNA copy number before to after aGvHD diagnosis within each patient. We also used Fine and Gray regression models to evaluate whether reduction in mtDNA copy number after aGvHD diagnosis was associated with one year non relapse mortality (NRM). Results The median age was 58 (interquartile range 38 to 65). 35% patients underwent a transplant from a HLA-identical sibling donor, 19% matched unrelated donor, 2% mismatched related donor, 14% single and 30% double umbilical cord blood. Conditioning was myeloablative in 36% and reduced intensity in 64%. aGvHD was diagnosed at a median of 42 (range 14-180) days from HCT. 16 (16%) of the patients were diagnosed with grade 1, 38% with grade II, 34% grade III and 11% grade IV aGvHD. Overall, geometric mean mtDNA copy number decreased 14% after aGvHD diagnosis (95% CI: 3% to 26%; p=0.01). MtDNA copy number decreased from pre-aGvHD to post-aGvHD in all grades of aGvHD though a decrease was seen in a larger proportion of patients with grade IV aGvHD (9/11; 82%) vs. patients with grade I aGvHD (8/16; 50%). There were 25 patients who experienced NRM within the first year of transplant. Exploratory analysis showed that mortality was lower among patients in whom mtDNA copy number increased post-aGvHD diagnosis (6/38; 16%) as compared to those in whom mtDNA copy number decreased post-aGvHD diagnosis (19/61; 31%) (p=0.12). Conclusion This study shows that mtDNA copy number was more likely to decrease after diagnosis of aGvHD. Thus, change in mtDNA copy number among CD3+ T lymphocytes may be a useful biomarker to identify individuals with aGvHD. The prognostic significance of reduced mtDNA copy number on NRM needs to be evaluated in larger studies.