Validation of CD4+ CD57+ PD1+ T Cells in Bronchoalveolar Lavage as a Biomarker of Lung Allograft Dysfunction

Abstract

Our analysis on serial bronchoalveolar lavage (BAL) cells from 25 lung transplant recipients (LTRs) using cutting-edge mass cytometry (MC) technology revealed a strong association between the frequency of CD4+ CD57+ PD1+ cells and subsequent lung allograft dysfunction (LAD). Here we present validation of this cell subset as a biomarker of incident LAD in an additional 25 LTRs. MC was performed using a panel of 37 heavy metal-tagged antibodies against surface markers and intracellular cytokines. This technique was applied to BAL cells from 50 consecutive LTRs at their 3-, 6- and 9-month post-transplant surveillance bronchoscopies. LTRs were randomly assigned into two groups (Fig 1A). Semi-supervised identification algorithm on the first group of LTRs identified CD4+ CD57+ PD1+ cells as the highest discriminatory cluster between stable and LAD patients. We then tested best-fit threshold for frequency of the cell subset and applied it on the second group of LTRs for validation. Longitudinal analysis of the first randomized group of 25 LTRs demonstrated a higher frequency of CD4+ CD57+ PD1+ cells separating LAD (11.41±4.89%) from stable patients (4.43±2.11%; Fig 1B). Using a receiver operating characteristic curve, we identified a frequency of 7.8% as the best-fit threshold of the cell subset (Fig 1C). Survival analysis showed CD4+ CD57+ PD1+ T cells at 7.8% can discriminate stable from LAD patients in the first group of LTRs (Fig 1D). We then tested the performance of this threshold in the second group of LTRs and found those with BAL CD4+ CD57+ PD1+ cells above the threshold had significantly lower freedom from incident LAD (p=0.05, Fig 1E). Our data validated that emergence of CD4+ CD57+ PD1+ T cells precedes allograft dysfunction in a small cohort of LTRs. Further validation in larger cohorts, and molecular and functional studies on this cell population are underway and may lead to improved prediction of LAD and a better understanding of lung transplant immunobiology.