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Acute graft versus host disease (aGVHD) is an important complication of allogeneic hematopoietic stem cell transplantation (HSCT). A need exists for validated biomarkers that can effectively predict the onset of aGVHD prior to clinical symptoms to allow for pre-emptive therapy. Expansion of CD8+ effector memory T-cells (TEM) have been observed at day +28 after HSCT in patients with aGVHD, suggesting their role in aGVHD pathophysiology. CD38 is a marker of T-cell activation but is also crucial for leukocyte migration through the endothelium.
We hypothesized that peripheral blood expansion of activated CD8+ TEMs would be observed following HSCT prior to onset of aGVHD symptoms.
Peripheral venous blood was collected twice weekly following HSCT for 7 weeks in 49 consecutive pediatric and young adult HSCT recipients. Samples were incubated with fluorochrome-conjugated antibodies against CD45, CD3, CD8, CD38, CD45RA and CCR7, followed by red cell lysis and fixation and analyzed by flow cytometry on a FACS Canto II flow cytometer (BD Biosciences). Data were analyzed using FCS Express (De Novo Software). TEM cells were defined as CD3+ CD8+CCR7- CD45RA- lymphocytes (Figure 1). Patients were followed for 100 days for development of aGVHD.
Twenty- three patients of median age 13.5 years (range 1-33 years) developed grade I-IV aGVHD at a median of 37 days (15-79 days) after HCST (Table 1). Peripheral expansion of CD38 bright CD8+TEM cells was observed at a median of 8 days (range 1-34 days) prior to clinical symptoms of aGVHD (Figure 2 A). A receiver-operating characteristic curve analysis revealed an area-under the curve of 0.85 (Figure 2B). Expansion of absolute CD38 bright CD8+ TEM of > 35 cells/µL predicted aGVHD with a sensitivity of 82.6%, specificity 91.6%, positive predictive value of 90.5% and negative predictive value of 84.6 %. The cumulative incidence of aGVHD was 90% in patients with CD38 bright CD8+TEM expansion and 15% in patients without (p< 0.0001) (Figure 2C).
Expansion of CD38 bright CD8+ TEM populations is a novel predictor of aGVHD and CD38 up regulation prior to aGVHD onset could represent trafficking lymphocytes.
Patient | Maximum Absolute CD38 Bright CD8+ TEM (cells/µL) | Maximum Absolute CD38 Bright CD8+ TEM (Day Post HSCT) | Onset of acute GVHD (Day Post HSCT) | Onset of CD38 Bright CD8+ TEM >35cells/uL (Day Post HSCT) | Maximum grade of aGVHD | Organ(s) involved |
1 | 64 | 26 | 33 | 26 | 1 | Skin |
2 | 69 | 48 | 58 | 37 | 3 | GI Liver |
3 | 32 | 34 | 41 | 34 | 1 | Skin |
4 | 304 | 34 | 47 | 20 | 1 | Skin |
5 | 231 | 7 | 34 | 7 | 1 | Skin |
6 | 41 | 24 | 34 | 24 | 2 | Skin |
7 | 99 | 25 | 32 | 25 | 4 | Skin Liver |
8 | 98 | 16 | 23 | 16 | 3 | Skin GI |
9 | 90 | 52 | 69 | 17 | 3 | Skin Liver |
10 | 183 | 37 | 52 | 20 | 3 | Skin GI Liver |
11 | 42 | 33 | 42 | 33 | 3 | Skin GI |
12 | 36 | 11 | 15 | 11 | 4 | Skin GI |
13 | 82 | 15 | 29 | 15 | 4 | GI |
14 | 37 | 43 | 44 | 12 | 2 | Skin |
15 | 151 | 45 | 79 | 15 | 3 | Skin GI |
16 | 63 | 23 | 33 | 23 | 4 | Skin GI |
17 | 47 | 25 | 31 | 25 | 1 | Skin |
18 | 87 | 17 | 18 | 17 | 3 | GI |
19 | 69 | 56 | 59 | 56 | 1 | Skin |
20 | 2 | 25 | 28 | NA | 1 | Skin |
21 | 12 | 36 | 37 | NA | 1 | Skin |
22 | 70 | 28 | 38 | 21 | 1 | Skin |
23 | 128 | 42 | 45 | 28 | 3 | Skin GI |
Table 1. Grade and timing of aGVHD relative to absolute CD38 bright CD8+TEM cells/µL prior to aGVHD onset
