Abstract
While mycophenolate mofetil (MMF) has replaced corticosteroids as immunosuppression in cord blood transplantation (CBT), optimal MMF dosing has yet to be established. We intensified MMF dosing from every 12 to 8 hours to augment graft-versus-host disease (GVHD) prophylaxis in double-unit CBT (dCBT) and evaluated outcomes according to the total daily MMF dose/kg in 174 double-unit CBT recipients (median age 39 years, range 1–71) transplanted for hematologic malignancies. Recipients of a MMF dose ≤ the median (36 mg/kg/day) had an increased day 100 grade III-IV acute GVHD (aGVHD) incidence compared with patients who received > 36 mg/kg/day (24% versus 8%, p = 0.008). Recipients of ≤ the median dose who had highly HLA-allele (1-3/6) mismatched dominant units had the highest day 100 grade III-IV aGVHD incidence of 37% (p = 0.009). This finding was confirmed in multivariate analysis (p = 0.053). In 83 patients evaluated for mycophenolic acid (MPA) troughs, those with a mean week 1-2 trough < 0.5 mcg/mL had an increased day 100 grade III-IV aGVHD of 26% versus 9% (p = 0.063), and those who received a low total daily MMF dose and had a low week 1-2 MPA trough had a 40% incidence (p = 0.008). Higher MMF dosing or MPA troughs had no impact on engraftment after myeloablation. This analysis supports intensified MMF dosing in mg/kg/day and MPA trough level monitoring early post-transplant in dCBT recipients.
Keywords: cord blood transplantation, graft-versus-host disease, mycophenolate mofetil, pharmacokinetics
Introduction
Mycophenolate mofetil (MMF) is an inactive morpholinoethyl ester prodrug that inhibits the proliferation of T- and B-lymphocytes and is designed to enhance the bioavailability of the active form, mycophenolic acid (MPA). MMF has been a successful immunosuppressive agent in adult donor allogeneic hematopoietic stem cell transplantation.1 Consequently, MMF was introduced in unrelated donor cord blood (CB) transplantation (CBT) to avoid the multiple toxicities of corticosteroids.2 However, some centers have had concerns that MMF could be associated with myelosuppression in CBT recipients.3,4 Furthermore, MPA's pharmacokinetics are characterized by a high intra- and inter- patient variability5,6, and inadequate MPA levels have been correlated with a reduced efficacy of prophylaxis and treatment of acute graft-versus-host disease (aGVHD).7,8
To augment MMF exposure, MMF has been increased from 1 gram (gm) every 12 hours to 1 gm every 8 hours in adult allograft recipients.9–11 As aGVHD is a leading cause of morbidity and transplantrelated mortality (TRM) in CBT12–14, intensified MMF dosing has also been adopted in CBT.15 Dosing in patients ≥ 50 kilograms (kg), however, has not taken into account the recipient's weight. Hence the association between MMF dosing and both aGVHD prevention and the potential toxicities to the bone marrow and gastro-intestinal tract merit investigation.16 As there is also no established target MPA exposure for CBT recipients, the association between MPA pharmacokinetic (PK) parameters and these end-points also require evaluation. We have, therefore, investigated the transplant outcomes in 174 double-unit CBT (dCBT) recipients treated for hematologic malignancies according to the total MMF dose in mg/kg/day. We hypothesized that MMF doses above the median total daily dose would be associated with a reduced incidence of grade III-IV aGVHD without an adverse effect upon neutrophil engraftment. In a subset of patients the association between total MPA trough levels and day 100 aGVHD and engraftment were also evaluated.
Methods
Patients and Graft Characteristics
Patients were transplanted at Memorial Sloan Kettering Cancer Center (MSKCC) between 10/1/2005 and 5/31/2013. Patients eligible for this analysis included all consecutive adult and pediatric recipients of first allografts who underwent dCBT for the treatment of hematologic malignancies with the exclusion of acute leukemia patients transplanted with ≥ 20% bone marrow blasts. All patients received double-unit CB grafts per institutional standard practice. CB units were selected on the basis of 4-6/6 HLA-A,-B antigen, -DRB1 allele match to the recipient, a cryopreserved total nucleated cell (TNC) dose of at least 1.5 × 107/kg/unit, and the bank of origin as previously described.17 Unit-unit HLA-match was not considered. High resolution HLA-A,-B,-C,-DRB1,-DQ allele typing of CB units was performed routinely for research purposes and usually did not influence unit selection. All patients provided written informed consent for transplantation and transplant outcome analysis on protocols approved by the MSKCC Institutional Review and Privacy Board.
Conditioning and GVHD Prophylaxis
Pre-transplant conditioning varied according to the patient's age, diagnosis, remission status, extent of prior therapy, and co-morbidities, and consisted of high-dose, reduced intensity (but functionally myeloablative), and non-myeloablative regimens as previously described.13,18 Intravenous calcineurin-inhibitors (CNI), predominantly cyclosporine-A (CSA), and MMF comprised GVHD prophylaxis starting on day -3 intravenously with no anti-thymocyte globulin (ATG). CSA was dosed to achieve a trough level 200–400 ng/ml and tacrolimus was dosed to achieve trough levels of 5–12 ng/mL. Before September 2009, 81 patients (47%) received MMF 1 gram intravenously every 12 hours (those ≤ 12 years or < 50 kg received 15 mg/kg/dose). From September 2009, intensified MMF dosing was instituted to augment GVHD prophylaxis. Ninety-three patients (53%) received MMF at 1 gram intravenously every 8 hours for patients both > 12 years and ≥ 50 kg (or 15 mg/kg/dose if > 12 years but < 50 kg). Patients ≤ 12 years received 20 mg/kg/dose. Granulocyte-colony-stimulating factor (5 mcg/kg/day rounded to vial size) was given post-CBT to all patients until neutrophil recovery.
Standard practice to taper MMF was to reduce the dose at approximately 75–100 days posttransplant if there was no aGVHD. In patients dosed every 8 hours, this interval was maintained for as long as possible unless there were adherence problems with this schedule in which case a switch to twice daily dosing was made. The taper was performed at 10–25% decrements in all patients with the goal of being off MMF by 4-6 months; though this schedule was ultimately at the physician's discretion. For example, increased relapse risk may have resulted in a hastened taper schedule with attention to the development of aGVHD symptoms. The most frequent reason to delay the MMF taper other than aGVHD was significant renal impairment preventing therapeutic cyclosporine levels. Dosing modifications were very rarely made for delayed engraftment early after transplant. However, later in the transplant course, dosing reductions were sometimes done for cytopenias usually in the context of concurrent valganciclovir therapy.
Sample Collection and Analysis for MPA Trough Levels
Serial weekly total MPA trough levels were collected on days 1, 8, 15, 22, 29 and 36 after dCBT from a subset of 85 patients transplanted from 8/1/2009–11/30/2012. Troughs were evaluated due to the substantial logistical advantage over the serial blood samples required for AUC measurements. Samples were collected primarily for research purposes. However, in 8/85 (9%) patients dosing was increased to every 8 hours in week 2 as a result of week 1 trough results. Samples of ≥ 1 mL serum were collected in a no additive red top tube. Peripheral blood was collected in adults whereas the sample in children was drawn from a non-MMF infused line. The blood was allowed to clot for 20–30 minutes at 15–28° C, centrifuged at 2700 r.p.m. for 10 minutes, and sent for analysis by liquid chromatography-mass spectrometry methodology at Quest Diagnostics Laboratories, Teterboro, NJ, USA. The result was available within 3–4 days.
Study Definitions
Neutrophil recovery was defined as the first of 3 consecutive days with an absolute neutrophil count ≥ 0.5 × 109/L whereas platelet recovery was the first of 3 consecutive days ≥ 20 × 109/L and at least 7 days without transfusion support. Engraftment was defined as sustained donor-derived count recovery with donor chimerism > 90% (both units combined). aGVHD was diagnosed clinically with histologic confirmation when appropriate. The grade of aGVHD was based on the International Bone Marrow Transplant Registry classification19 except grades A-D were labeled grades I-IV. Grading was reviewed by a transplant clinician panel to reach a consensus of maximum aGVHD grade. Chronic GVHD was defined according to the National Institutes of Health consensus criteria.20 Relapse was defined as recurrence or progression of disease over pre-transplant baseline, and TRM was death from any cause in continued remission. Progression-free survival (PFS) was defined according to standard criteria.
Statistical Analysis
For analysis purposes, patients were a priori divided into two dosing groups: above or below the median total daily mg/kg MMF dose. The strategy to analyze outcomes according to above versus below the median total daily dose was chosen as deriving a threshold based on the observed data would require a separate validation cohort. Descriptive analyses were performed for patient demographics and differences across low versus high MMF groups and compared using the Wilcoxon rank-sum test for continuous covariates. Chi-square and Fisher's exact test were used for categorical covariates as indicated. The cumulative incidence method for competing risks was used to calculate the cumulative incidence of engraftment, GVHD, TRM, and relapse. Gray's test was used to compare the incidence of select outcomes across MMF dose levels. PFS was calculated using Kaplan-Meier methodology and compared across MMF dose levels using a log rank test.
In the subset analyses of MPA trough levels, descriptive statistics summarized the changes in MPA trough levels over the first 6 weeks post-transplant and the trends across age and conditioning regimens. Fisher's exact test evaluated associations between patient clinical factors and mean week 1-2 trough levels dichotomized at 0.5 mcg/mL. To evaluate the association between MPA trough levels and dCBT outcomes, mean levels of week 1 and week 2 combined were dichotomized at < 0.5 mcg/mL and ≥ 0.5 mcg/mL for efficacy, and < 2 mcg/mL and ≥ 2 mcg/mL for toxicity. Starting from a landmark of 2 weeks post-transplant, cumulative incidence functions and Gray's test were used to estimate and compare engraftment and aGVHD by mean MPA trough levels. All analyses were conducted using R statistical software, version 3.1.1 (R Foundation for Statistical Computing, Vienna, Austria).
Results
Comparison of Patient Characteristics According to MMF Dosing
One-hundred and seventy-four patients were analyzed. The majority (n = 136, 78%) received myeloablative conditioning. The median total daily dose of MMF was 36 mg/kg. Patients who received low (below the median) and high (above the median) total daily doses were comparable in terms of age, recipient CMV status, disease type and risk, conditioning regimen intensity, and graft characteristics (dominant or engrafting unit infused CD34+ cell dose and 6 HLA-allele match) (Table 1). However, patients in the low dosing group were more likely to be male, heavier, and receive MMF every 12 hours.
Table 1.
Patient and Graft Demographics According to Total Daily MMF Dose/kilogram (Below versus Above the Median, mTDD, mg/kg)
| Characteristic | mTDD ≤ 36 (n = 101) | mTDD > 36 (n = 73) | p-value |
|---|---|---|---|
|
| |||
| Median Age (years, range) | 42 (1–66) | 36 (2–71) | 0.181 |
|
| |||
| Recipient Gender, N (%) | |||
| Male | 70 (69%) | 27 (37%) | < 0.001 |
| Female | 31 (31%) | 46 (63%) | |
|
| |||
| Median Weight (kg, range) | |||
| Overall | 80 (7–125) | 54 (10–77) | < 0.001 |
| Male | 82 (7–125) | 63 (11–76) | < 0.001 |
| Female | 70 (13–104) | 53 (10–77) | < 0.001 |
|
| |||
| Recipient CMV Serostatus, N (%) | |||
| Negative | 51 (51%) | 27 (37%) | 0.107 |
| Positive | 50 (50%) | 46 (63%) | |
|
| |||
| Disease Type, N (%) | |||
| Acute leukemia | 52 (52%) | 48 (66%) | 0.173 |
| MDS/CML/Other MPD | 7 (7%) | 3 (4%) | |
| NHL/ HL/ CLL | 42 (42%) | 22 (30%) | |
|
| |||
| Disease Risk, N (%) | |||
| Standard | 11 (11%) | 9 (12%) | 0.958 |
| High | 90 (89%) | 64 (88%) | |
|
| |||
| Conditioning Intensity, N (%) | |||
| Myeloablative | 78 (77%) | 58 (80%) | 0.869 |
| Non-myeloablative | 23 (23%) | 15 (21%) | |
|
| |||
| Dosing Interval, N (%) | |||
| Every 12 hours | 71 (70%) | 10 (14%) | < 0.001 |
| Every 8 hours | 30 (30%) | 63 (86%) | |
|
| |||
| Dominant Unit Median Infused CD34+ Cell Dose (range) (× 105/kg) | |||
| Larger unit | 1.30 (0.26–6.42) | 1.24 (0.41–4.77) | 0.793 |
| Smaller unit | 0.68 (0.13–1.52) | 0.66 (0.08–2.12) | 0.976 |
|
| |||
| Unit 6-Allele HLA-Match, N (%)* | 0.130 | ||
| 1-3/6 | 75 (37%) | 42 (29%) | |
| 4-6/6 | 127 (63%) | 104(71%) | |
mTDD, median total daily dose; CMV, cytomegalovirus; MDS, myelodysplastic syndrome; CML, chronic myelogenous leukemia; MPD, myeloproliferative disorder; NHL, Non-Hodgkin's Lymphoma; HL, Hodgkin's Lymphoma; CLL, chronic lymphocytic leukemia.
The HLA-match grade reflects 202 units in the low MMF patient group and 146 units in the high group.
Association between MMF Dosing and aGVHD
Transplant outcomes according to MMF dosing below versus above the median total daily MMF dose/kg are shown in Table 2. There was no difference in the cumulative incidences of grade II-IV aGVHD at day 100 in the low and high dosing groups (53% versus 47%, p = 0.345). The incidence of severe (grade III-IV) aGVHD at day 100, however, was 3-fold higher in recipients of a total daily MMF dose below the median versus those in the high group (24% versus 8%, p = 0.008). One-year chronic GHVD was not different in the two groups (12% versus 11%, p = 0.422).
Table 2.
dCBT Outcomes by Total Daily MMF Dose/kilogram (Below versus Above the Median)
| Variable | Cumulative Incidence (95%CI) | p-value |
|---|---|---|
|
| ||
| Day 45 Neutrophil Engraftment * | ||
| Low MMF dose (n = 78) | 95% (86–98) | 0.990 |
| High MMF dose (n = 58) | 95% (83–99) | |
|
| ||
| Day 100 aGVHD II-IV | ||
| Low MMF dose (n= 101) | 53% (42–62) | 0.345 |
| High MMF dose (n = 73) | 47% (35–58) | |
|
| ||
| Day 100 aGVHD III-IV | ||
| Low MMF dose (n = 101) | 24% (16–33) | 0.008 |
| High MMF dose (n = 73) | 8% (3–16) | |
|
| ||
| 1-Year Chronic GVHD | ||
| Low MMF dose (n = 101) | 12% (7–19) | 0.422 |
| High MMF dose (n = 73) | 11% (5–20) | |
|
| ||
| Day 180 TRM | ||
| Low MMF dose (n = 101) | 20% (13–28) | 0.371 |
| High MMF dose (n = 73) | 21% (12–31) | |
|
| ||
| 1-year Relapse | ||
| Low MMF dose (n = 101) | 14% (8–21) | 0.518 |
| High MMF dose (n = 73) | 15% (8–24) | |
|
| ||
| 1-year PFS | ||
| Low MMF dose (n = 101) | 63% (53–72) | 0.225 |
| High MMF dose (n = 73) | 59% (47–69) | |
GVHD, graft verus host disease; TRM, transplant related mortality; PFS, progression-free survival
Myeloablative CBT recipients only
In univariate analysis of variables potentially associated with the incidence of day 100 grade II-IV aGHVD, aGVHD was associated with gender (57% in male versus 42% in female patients, p = 0.027) and CMV serostatus (58% in seronegative versus 44% in seropositive dCBT recipients, p = 0.032). The results of univariate analysis of grade III-IV aGVHD are shown in Table 3. Male gender was associated with a higher incidence of severe aGVHD (25% versus 8%, p = 0.003). Despite males being heavier, recipient weight (below or above the median) was not significant, and analyzing the patient's weight as a continuous variable or in quartiles also showed no differences in aGVHD incidence. Additionally, the association between gender and grade III-IV aGVHD persisted in both the subset of patients whose weight was below the median and those whose weight was above the median. Among the subset of male patients only, there appeared to be no association between weight and grade III-IV aGVHD (data not shown). Finally, in male patients there was no difference in the gender of the dominant unit and therefore gender mismatch did not explain increased aGVHD incidence in males.
Table 3.
Univariate Analysis of Day 100 Grade III-IV aGVHD According to Patient and Graft Characteristics
| Variable | Cumulative Incidence (95% CI) | p-value |
|---|---|---|
|
| ||
| Age | ||
| 0–15 years (n = 35) | 26% (13–41) | 0.129 |
| ≥ 16 years (n= 139) | 15% (10–22) | |
|
| ||
| Recipient Gender | ||
| Male (n = 97) | 25% (17–34) | 0.003 |
| Female (n = 77) | 8% (3–15) | |
|
| ||
| Engrafting Unit Gender in Males * | ||
| Male (n = 54) | 26% (15–38) | 0.478 |
| Female (n = 40) | 20% (9–34) | |
|
| ||
| Median Weight (kg) | ||
| ≤ 67.5 (n = 87) | 17% (10–26) | 0.973 |
| > 67.5 (n = 87) | 17% (10–26) | |
|
| ||
| Recipient CMV Serostatus | ||
| Negative (n = 78) | 24% (16–34) | 0.026 |
| Positive (n = 96) | 12% (6–19) | |
|
| ||
| Conditioning Intensity | ||
| Myeloablative (n = 136) | 18% (12–25) | 0.800 |
| Non-myeloablative (n = 38) | 16% (6–29) | |
|
| ||
| Median MMF Dose (nig/kg/day) | ||
| ≤ 36 (n = 101) | 24% (16–33) | 0.008 |
| > 36 (n = 73) | 8% (3–16) | |
|
| ||
| Dominant Unit-Recipient 6-Allele HLA-Match | 0.105 | |
| 1-3/6 (n = 48) | 25% (14–38) | |
| 4-6/6 (n = 126) | 14% (9–21) | |
|
| ||
| Median Total Daily MMF Dose & Dominant Unit-Recipient 6-Allele HLA-Match | ||
| Low (≤ 36) and Worse (1-3/6) (n = 30) | 37% (20–54) | 0.009 |
| High (> 36) and Worse (1-3/6) (n = 18) | 6% (0–23) | |
| Low (≤ 36) and Better (4-6/6) (n = 71) | 18% (10–28) | |
| High (> 36) and Better (4-6/6) (n = 55) | 9% (3–19) | |
CMV, cytomegalovirus
Dominant unit gender unknown in 3 patients
In univariate analysis, there was also a significant association between grade III-IV aGVHD and the combination of MMF dose and dominant unit-recipient HLA-match (p = 0.009, Table 3). Recipients of total daily MMF doses below the median who had dominant units with a high degree of HLA-mismatch (n = 30) had the highest incidence of day 100 grade III-IV aGVHD at 37%. By contrast, patients with a high MMF dose had an incidence of severe aGVHD of < 10%. Severe aGVHD incidence in those with a low MMF dose but better HLA-allele matched engrafting unit was intermediate at 18%. Differences in severe aGVHD by dominant unit-recipient HLA-match alone did not reach significance.
In a multivariate analysis that included age, gender, CMV serostatus, and the four category combination of MMF dose and HLA-mismatch, males were significantly more likely to develop grade III-IV aGVHD (p = 0.041, Table 4). The risk of grade III-IV aGVHD also differed by MMF dose and degree of HLA-mismatch combination with the highest risk in those who received a low MMF dose and higher HLA-mismatch (p = 0.053).
Table 4.
Multivariate Analysis of Variables Potentially Associated with Day 100 Grade III-IV aGVHD Risk
| Variable | HR (95% CI) | p-value |
|---|---|---|
|
| ||
| Age, year | ||
| 0–15 (n = 35) | Reference | 0.217 |
| ≥ 16 (n = 139) | 0.58 (0.24–1.38) | |
|
| ||
| Recipient Gender | ||
| Male (n = 97) | Reference | 0.041 |
| Female (n = 77) | 0.38 (0.15–0.96) | |
|
| ||
| Recipient CMV Serostatus | ||
| Negative (n = 78) | Reference | 0.190 |
| Positive (n = 96) | 0.58 (0.26–1.31) | |
|
| ||
| Median Total Daily MMF Dose & Dominant Unit-Recipient 6-Allele HLA-Match | ||
| Low (≤ 36) and Worse (1-3/6) (n = 30) | Reference | 0.053 |
| High (> 36) and Worse (1-3/6) (n = 18) | 0.23 (0.03–1.84) | |
| Low (≤ 36) and Better (4-6/6) (n = 71) | 0.46 (0.20–1.07) | |
| High (> 36) and Better (4-6/6) (n = 55) | 0.26 (0.09–0.75) | |
CMV, cytomegalovirus
Association between MMF Dosing and Engraftment and Survival Endpoints
Given that most non-myeloablative dCBT recipients had transient autologous recovery, the effect of MMF dosing on donor engraftment was only compared among myeloablated dCBT recipients. The cumulative incidences of day 45 neutrophil engraftment were nearly identical regardless of MMF dosing, with each being 95% in the low and high MMF dosing groups at medians of 23 and 24 days, respectively (p = 0.990, Table 2). The cumulative incidences of day 180 platelet engraftment were also similar: 82% (95%CI: 71–89, median 49 days, range 29–162) in the low dose group and 86% (95%CI: 72–94, median 46 days, range 30–137) in the high dose group (p = 0.270).
There were no significant differences in the cumulative incidences of TRM or relapse by dosing group, and the PFS were also similar (Table 2). Of 49 TRM deaths, GVHD was the most common cause (n = 22) followed by organ toxicity (n = 13). Infection as the primary cause of death was uncommon occurring in only 7 patients overall. There were no differences in the percentage of patients dying of GVHD in low versus high MMF dosing groups. Eleven of 22 patients who died of GVHD did so prior to day 180. Of these 22 patients, their aGVHD onset was prior to day 100 in all but one patient, and all 22 patients who died of GVHD had either aGVHD (n = 20) or aGVHD followed by overlap syndrome (n = 2).
Subset Analysis of Total MPA Trough Levels
Serial MPA troughs during weeks 1–6 were available in 85 patients. These showed an increase in exposure over time for all patients: week 1 median MPA trough was 0.9 mcg/mL and week 6 median was 1.3 mcg/mL. Two patient groups had consistently lower levels in the first 6 weeks post-transplant: children < 16 years of age whose levels ranged 0.3–1.2 mcg/mL lower than adults, and recipients of myeloablative conditioning whose levels ranged 0.4–0.8 mcg/mL lower than those who received nonmyeloablative conditioning.
Based on the known importance of therapeutic early post-transplant week CNI levels21, we analyzed day 100 aGVHD risk according to the mean week 1-2 MPA trough levels using a level ≥ 0.5 mcg/mL. We chose this concentration because a mean week 1-2 trough > 0.5 mcg/mL is clinically significant for efficacy of aGVHD therapy.8 Eighty-three patients had complete blood samples in the first 2 weeks post-transplant. Clinical factors such as age, gender, above or below 120% of ideal body weight, concomitant CNI, and albumin and bilirubin levels above or below the upper limit of normal were not associated with trough levels of < 0.5 or ≥ 0.5 mcg/mL, and CNI levels were therapeutic for all patients in the first two weeks (data not shown). Fifty-six of 69 (81%) patients who received MMF every 8 hours had trough levels ≥ 0.5 mcg/mL compared with 8/14 (57%) in the 12 hour dosing group (p = 0.078).
There was no difference in the cumulative incidence of day 100 grade II-IV aGVHD in MPA trough level groups of < 0.5 and ≥ 0.5 mcg/mL (Figure 1a). Patients with troughs of < 0.5 mcg/mL, however, had a grade III-IV aGVHD incidence of 26% versus 9% (p = 0.063, Figure 1b). dCBT recipients (n = 10) who received a low total daily MMF dose (≤ median of 43 mg/kg/day in this 83 patient subset) and had a low week 1-2 MPA trough level (< 0.5 mcg/mL) had an incidence of grade III-IV aGVHD at day 100 of 40% (95%CI: 7–73) as compared with that of 10% (95%CI: 3–16) in the 73 patients with other dose and trough level combinations (i.e. high MMF dose regardless of trough level or trough ≥ 0.5 mcg/mL regardless of dose, p = 0.008).
Figure 1a. Cumulative incidence of day 100 II-IV aGVHD by MPA Trough Level (n = 83).
In a day 14 landmark analysis week 1-2 MPA trough levels ≥ 0.5 mcg/mL had no association with grades II-IV aGVHD by day 100.
Figure 1b. Cumulative incidence of day 100 III-IV aGVHD by MPA Trough Level (n = 83).
In a day 14 landmark analysis the incidence of grade III-IV aGVHD at day 100 was lower in dCBT recipients with week 1-2 MPA trough levels ≥ 0.5 mcg/mL.
To evaluate whether high MPA troughs correlated with myelosuppression or gastro-intestinal toxicity, engraftment and duration of TPN were analyzed. Concentrations of < 2 and ≥ 2 mcg/mL were chosen because ≥ 2 mcg/mL have been associated with leukopenia in solid organ transplant recipients.22 There was no difference in the cumulative incidence of day 100 neutrophil engraftment in myeloablated patients with mean week 1-2 MPA troughs of < 2 and ≥ 2 mcg/mL: 95% (95%CI:89–100) versus 100% (95%CI:78–100), p = 0.422. Also in myeloablated patients, there were no differences in platelet engraftment or duration of TPN (data not shown).
Discussion
Although MMF immunosuppression in combination with a CNI has been used in CBT as aGVHD prophylaxis since the year 20002, optimal dosing of MMF has not been established. In view of the significant incidence of aGVHD in our patient population13 we have intensified MMF dosing to every 8 hours in dCBT recipients. However, in this analysis we analyzed transplant outcomes according to the MMF dose in mg/kg/day as dosing has not accounted for recipient weight ≥ 50 kg, and dosing in small children has substantially changed over the time period of the study. Patients in each mg/kg/day dosing groups were well matched strengthening the multiple significant findings of this analysis. We found recipients who received a low mg/kg/day dose of MMF had a significantly higher incidence of day 100 grade III-IV acute GVHD. Recipients of a low dose who had a highly HLA-allele mismatched dominant unit were at the worst risk. Finally, patients with mean week 1-2 MPA trough concentrations < 0.5 mcg/mL had an increased day 100 grade III-IV aGVHD with no effect on hematopoietic recovery or gastro-intestinal toxicity as measured by TPN use. To our knowledge, this is the first large and comprehensive analysis to address the issue of MMF dosing in mg/kg/day and include assessment of the clinical significance of MPA trough levels early post-transplant in CBT patients.
Important in this analysis, when combining the MMF total daily dose with the degree of dominant unit-recipient HLA-match, we noted that a higher MMF dose could potentially reduce the detrimental impact of having a lesser HLA-matched engrafting unit. This is clinically relevant given our center's prior report that found a less well matched dominant unit was associated with a greater grade III-IV aGVHD risk.13 While our findings must be confirmed in larger studies, they suggest that a higher MMF dose is beneficial in recipients of grafts with a high degree of HLA-mismatch.
While it is established that small children metabolize MMF more rapidly than adults, we did not identify a significant association between age and grade III-IV aGVHD (Tables 3 and 4). This may have been due to partial abrogation of aGVHD by the substantial increase in MMF dose in small children during the study period, and it is an important question for further investigation. An unexpected observation in this series was that male patients had a significantly higher incidence of grade III-IV aGVHD than women. This was not due to an increased aGVHD risk in those with heavier weight or due to a greater risk in male patients engrafting with female units. This observation concerning gender is of great interest and remains unexplained.
The lack of difference in TRM between the low and high MMF dosing groups may have been due to aggressive supportive care administered to severe aGVHD cases. Additionally, as half of the 22 GVHD-related deaths were after day 180 this diluted the sample size to be able to identify differences at day 180. This question of whether augmented MMF dosing can reduce GVHD-related TRM is critically important, however, and requires further investigation in a larger study. Nonetheless, it is clear that the GVHD deaths in this report were related to aGVHD, and in nearly all of these patients the GVHD began prior to day 100. Therefore, aGVHD is the correct target for prevention, and as severe aGVHD increases patient morbidity and transplant costs it is a serious complication to abrogate. Importantly, in relation to other survival end-points, increased MMF dosing was not associated with an increase in deaths from infections or the incidence of relapse.
Previous reports support the efficacy of PK monitoring. Studies suggest that higher MPA exposure from an increased trough23, steady state concentration24, or AUC25,26 results in a lower aGHVD incidence although the ideal monitoring parameter has not been established. In this study, a mean week 1-2 MPA trough ≥ 0.5 mcg/mL was hypothesized as clinically relevant for prophylaxis given its relationship with therapeutic efficacy.8 We found that troughs < 0.5 mcg/mL were associated with an increased grade III-IV aGVHD risk complementing the findings of the larger 174 patient analysis. Moreover, that dCBT recipients who received a low total daily MMF dose/kg and had a low week 1–2 MPA trough had a very high incidence of grade III-IV aGVHD demonstrates that these patients in particular require increased MPA exposure. While we did not identify an age effect in the 174 patient analysis, young children are potentially at special risk given patients < 16 years of age had lower troughs in our analysis over the six week period as expected given their faster MMF metabolism.27 Indeed, Osunkwo and colleagues have proposed an increased MMF dosing to 900–1200 mg/m2 every 6 hours to achieve levels ≥ 1 mcg/mL, which corresponds to nearly 30 mg/kg/dose in children.23
From the stand-point of engraftment, an increased total daily MMF dose/kg had no deleterious effect. For this MPA trough level toxicity analysis, we choose a 2 mcg/mL cut-off based on solid organ transplantation literature.22 The lack of any association between 1-2 week troughs and engraftment supports the findings of the larger 174 patient analysis. The upper threshold for total MPA troughs in CBT has yet to be established, however, and the possible toxicity of very high levels could remain a clinical concern. While Sanz and colleagues reported a delay in neutrophil engraftment with the use of MMF compared with corticosteroids in patients receiving single CBT3, we have no evidence to indicate that intensified dosing as investigated in this MSKCC analysis is detrimental in dCBT. A potential risk of gastro-intestinal toxicity was also not substantiated, and there were no cases of MMF colitis.
Multiple questions concerning MMF pharmacokinetics remain unanswered. Though the majority of our patients remained on IV MMF for the duration of the PK subset period, the impact of changing from intravenous to oral drug was not addressed in this analysis and could be important given the potential drop in bioavailability.28 Additionally, whether intensified MMF dosing later in the transplant course can exacerbate myelosuppression is possible, especially in the setting of other myelosuppressive drugs such as valganciclovir. Finally, how to correctly taper MMF post-transplant is not established. All of these questions require formal future investigation.
This analysis supports intensified MMF dosing based on the total daily dose/kg even in adult dCBT recipients receiving relatively low unit cell doses. As a recognized adult dosage is 15 mg/kg/dose for patients < 50 kg, we propose patients ≥ 50 kg also receive 15 mg/kg/dose every 8 hours with a dose cap of 1.5 gm every 8 hours. This intervention could prove especially important in heavy patients and/or recipients of highly HLA-mismatched units. Furthermore, while the ideal dosing in children less than 12 years remains unknown, we now support intensified dosing of 30 mg/kg/dose (nearly 900 mg/m2/dose) as previously suggested in Bhatia et al29 to optimize MPA exposure in this population. Finally, obtaining MPA trough levels (ideally with rapid result availability) during the first two weeks after transplant to target a trough level ≥ 0.5 mcg/mL is also appropriate in dCBT recipients.
Highlights.
Higher total daily MMF dose/kg reduces severe aGVHD after CBT
Higher MMF dose can reduce the aGVHD risk of marked HLA- mismatch after CBT
Higher week 1-2 MPA troughs were associated with reduced severe aGVHD after CBT
Higher MMF dosing and MPA troughs did not impair neutrophil engraftment after CBT
Acknowledgments
Financial disclosure: this work was supported in part by the Gabrielle's Angel Foundation for Cancer Research (J.N.B.), the Memorial Sloan-Kettering Cancer Center Society (J.N.B.), the Translational and Integrative Medicine Research Grant (J.N.B.), P01 CA23766 from the National Cancer Institute, National Institutes of Health (J.N.B.), the Society of Memorial Sloan Kettering Cancer Center (J.N.B.), and the New York State Empire Clinical Research Investigator Program (D.M.P.).
Footnotes
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Conflict of interest statement: There are no conflicts of interest to report.
Authorship statement: S.H., D.M.P. and J.N.B. designed the study, interpreted the data, and wrote the manuscript. P.H., J.Z., and S.M.D. analyzed the data and wrote the manuscript. M.L. analyzed the data. M.P., S.M., N.A., N.A.K., R.O., S.P., A.S., A.H., R.J., M.M.v.B., S.G., M.A.P., and J.W.Y. wrote the manuscript.
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