Differential Elimination of Anti-Thymocyte Globulin of Fresenius and Genzyme Impacts T-Cell Reconstitution After Hematopoietic Stem Cell Transplantation

Abstract

Anti-thymocyte globulin (ATG) is a lymphocyte depleting agent applied in hematopoietic stem cell transplantation (HSCT) to prevent rejection and Graft-vs.-Host Disease (GvHD). In this study, we compared two rabbit ATG products, ATG-Genzyme (ATG-GENZ), and ATG-Fresenius (ATG-FRES), with respect to dosing, clearance of the active lymphocyte binding component, post-HSCT immune reconstitution and clinical outcome. Fifty-eigth pediatric acute leukemia patients (n = 42 ATG-GENZ, n = 16 ATG-FRES), who received a non-depleted bone marrow or peripheral blood stem cell graft from an unrelated donor were included. ATG-GENZ was given at a dosage of 6-10 mg/kg; ATG-FRES at 45-60 mg/kg. The active component of ATG from both products was cleared at different rates. Within the ATG-FRES dose range no differences were found in clearance of active ATG or T-cell re-appearance. However, the high dosage of ATG-GENZ (10 mg/kg), in contrast to the low dosage (6-8 mg/kg), correlated with prolonged persistence of active ATG and delayed T-cell reconstitution. Occurrence of serious acute GvHD (grade III-IV) was highest in the ATG-GENZ-low dosage group. These results imply that dosing of ATG-GENZ is more critical than dosing of ATG-FRES due to the difference in clearance of active ATG. This should be taken into account when designing clinical protocols.

Keywords: ATG; Fresenius; Genzyme; acute GvHD; pediatrics; serotherapy.

Figures

Figure 1

Serum/plasma concentration of total Rabbit IgG (total ATG) and active ATG in ATG-GENZ (Genzyme, red) or ATG-FRES (Fresenius, green) treated patients. Total Rabbit IgG (μg/mL) of ATG-FRES treated patients (B) was higher in comparison with ATG-GENZ treated patients (A), because of the higher dose of ATG-FRES given, but clearance was comparable for both brands. Active ATG levels (AU/mL) of ATG-FRES treated patients (D), receiving 60 mg/kg (solid lines; n = 9) or 45 mg/kg (dashed lines; n = 7), decreased fast and was only in 1 out of 16 patients (6%) above 1 AU/mL at 3 weeks post-HSCT. ATG-GENZ treated patients (C) receiving 10 mg/kg (solid lines; n = 24) or 6–8 mg/kg (dashed lines; n = 18) showed more variance in clearance. At 3 weeks post-HSCT 15 out of 24 patients (63%) receiving 10 mg/kg had active ATG levels above 1 AU/mL vs. only 2 out of 18 patients (11%) receiving 6–8 mg/kg. The horizontal dashed line at 1 AU/mL marks the highest serum/plasma concentration at which T-cells can reappear in the blood. The vertical solid gray line is the expected time of neutrophil engraftment. Of note, in the 6 patients receiving ATG-GENZ and showing a steep decline of rabbit ATG IgG antibodies against rabbit IgG were detected.

Figure 2

Box plots of serum/plasma concentration of total Rabbit IgG (total ATG) and active ATG in ATG-GENZ (Genzyme, red) and ATG-FRES (Fresenius, green) treated patients. (A) Total Rabbit IgG concentration in ATG-FRES treated patients at week 0 (day of graft infusion) and week +3 (expected time of neutrophil engraftment) is significantly higher in comparison with ATG-GENZ treated patients. (B) Active ATG concentration at week 0 in ATG-FRES treated patients is significantly higher in comparison with ATG-GENZ treated patients, but at week +3 the serum/plasma level of active ATG in ATG-FRES treated patients is lower than in ATG-GENZ treated patients (0.17 vs. 0.65 AU/mL).

Figure 3

Box plots of serum/plasma concentration of active ATG in ATG-GENZ (Genzyme) and ATG-FRES (Fresenius) treated patients depending on the given total ATG dose. At week 0 (day of graft infusion) the ATG-GENZ patients receiving a total ATG dose of 10 mg/kg had a significantly higher serum/plasma level of active ATG than the 6–8 mg/kg receiving patients. Active ATG in ATG-FRES treated patients was at week 0 comparable between 45 and 60 mg/kg total ATG dose receiving patients and higher than in ATG-GENZ treated patients. At week +3 post-HSCT the serum/plasma level of active ATG in ATG-GENZ treated patients was still strongly dependent on the given dose (1.29 and 0.11 AU/mL), but the active ATG concentration in ATG-FRES treated patients was comparably low regardless of ATG dosing (0.12 and 0.21 AU/mL, respectively). The horizontal dashed line at 1 AU/mL is the highest serum/plasma concentration at which T-cells can recover.

Figure 4

Immune cell recovery post-HSCT in ATG-FRES (Fresenius) treated patients in comparison with high (ATG-GENZ-high) and low dose (ATG-GENZ-low) ATG-GENZ (Genzyme) treated patients. CD3 T-cell and CD4 and CD8 T-cell subset numbers differed significantly at one month post-HSCT between the two ATG treated groups. ATG-GENZ-high patients recovered significantly slower for CD3, CD4, and CD8 compared to the ATG-FRES and ATG-GENZ-low treated patients. In contrast to the T-cell recovery, NK-cells recovered fast after HSCT reaching normal levels within 1 month. The highest level of NK-cell numbers were observed at one month in the ATG-GENZ-high group. B-cell recovery in all ATG groups returned to normal levels at 6 months post-HSCT with a significantly higher number of B-cells at 2 and 3 months in the ATG-GENZ-high compared to the ATG-GENZ-low and ATG-FRES treated patients, respectively. Dotted horizontal lines represent the 5th and 95th percentiles of cell numbers in 28 healthy age-matched donors. Gray line: patients transplanted for acute leukemia with a graft from an HLA-identical sibling donor without receiving ATG in the conditioning.

Figure 5

The effect of ATG brand and dosing on acute GvHD. Although not significant, acute GvHD (grade II-IV) occurred less frequently in ATG Genzyme high dose (ATG-GENZ-high) receiving patients compared to patients receiving a low dose of ATG-Genzyme (ATG-GENZ-low) or ATG Fresenius (ATG-FRES). For severe acute GvHD (grade III-IV), there was a significant difference between the three ATG groups. Severe acute GvHD occurred most frequently in the ATG-GENZ-low group.