Sirolimus in Combination with Cyclosporine Or Tacrolimus Plus

Home , Sirolimus

Biology of Blood and Marrow Transplantation 14:531-537 (2008) Q 2008 American Society for Blood and Marrow Transplantation 1083-8791/08/1405-0001$32.00/0 doi:10.1016/j.bbmt.2008.02.009

Sirolimus in Combination with Cyclosporine or Tacrolimus Plus Methotrexate for Prevention of Graft-versus-Host Disease following Hematopoietic Cell Transplantation from Unrelated Donors

Terry Furlong, Hans-Peter Kiem, Frederick R. Appelbaum, Paul A. Carpenter, H. Joachim Deeg, Kristine Doney, Mary E. D. Flowers, Marco Mielcarek, Richard A. Nash, Rainer Storb, Paul J. Martin

Division of Clinical Research, Fred Hutchinson Cancer Research Center, and the Department of Medicine, University of Washington School of Medicine, Seattle, Washington

Correspondence and reprint requests: Terry Furlong, Fred Hutchinson Cancer Research Center, 1100 Fairview Avenue N, LF-200, P.O. Box 19024, Seattle, WA 98109-1024 (e-mail: [email protected]).

Received August 7, 2007; accepted February 13, 2008

ABSTRACT In 2 consecutive prospective clinical trials, we evaluated the efﬁcacy of sirolimus together with a calcineurin inhibitor (cyclosporine or tacrolimus) and low-dose methotrexate for prevention of graft-versus-host disease (GVHD) after unrelated hematopoietic cell transplantation (HCT). Nine patients received sirolimus with cyclosporine, and 17 received sirolimus with tacrolimus. The incidence of grade II-IV GVHD was 77%, with the median onset at day 7 after HCT. Because of toxicity, administration of sirolimus was discontinued earlier than planned in 11 patients, but after the onset of GVHD. Three patients developed renal failure requiring hemodialysis. Accrual in both studies was terminated because of lack of efﬁcacy. In these studies, the addition of sirolimus to regimens containing a calcineurin inhibitor and methotrexate appeared to cause toxicity and provided no detectable improvement in preventing GVHD. Ó 2008 American Society for Blood and Marrow Transplantation

KEY WORDS Sirolimus Graft-versus-host disease Hematopoietic cell transplantation

INTRODUCTION activation-induced apoptosis. The proliferation of T cells driven by other lymphokines, such as IL-4, IL- Acute graft-versus-host disease (GVHD) is a major cause of morbidity and nonrelapse mortality (NRM) 12, IL-7, and IL-15, is also inhibited by sirolimus after allogeneic hematopoietic cell transplantation [11]. Sirolimus inhibits lipopolysaccharide-induced (HCT) [1]. Current pharmacologic approaches to pre- proliferation of B cells resistant to TAC and CSP vent acute GVHD (aGVHD) generally involve the use [11,12]. Although both TAC and sirolimus have the of a combination of immunosuppressive medications, same intracellular receptor, it is hypothesized that such as cyclosporine (CSP) or tacrolimus (TAC), the drug-immunophilin complexes interact differently together with an antimetabolite, such as methotrexate with other molecules to form functionally distinct (MTX) [2-6]. Despite the use of these regimens, 35%- complexes and speciﬁc immunosuppressive effects 80% of recipients develop aGVHD, often requiring peculiar to each drug. Calcineurin inhibitors used in systemic immunosuppressive treatment [7-10]. combination with sirolimus cause an additive or syn- Sirolimus is a macrocyclic lactone fermentation ergistic suppression of immune cell function [13]. product with antifungal and immunosuppressive ac- Sirolimus is effective for preventing rejection of tivity. The immunosuppressive effect of sirolimus is renal and hepatic allografts [14-17], and has been eval- mediated through a mechanism different from TAC uated for treatment of aGVHD [18] and chronic or CSP. Sirolimus blocks IL-2-induced proliferation GVHD (cGVHD) [19,20]. Results of several recent of T cells but does not affect signals that result in studies have suggested that sirolimus is effective for

531 532 T. Furlong et al. preventing GVHD after allogeneic HCT [21-23].In2 The protocols restricted enrollment to patients phase II clinical trials, we attempted to confirm the with high-risk diagnoses, allowed a single class 1 allele efficacy and safety of sirolimus for preventing aGVHD disparity, and did not restrict the type of donor graft after unrelated HCT, first in combination with CSP used, because it was felt that these criteria would not and MTX, and subsequently in combination with significantly affect the protocol endpoints. For similar TAC and MTX. reasons, the eligibility criteria allowed the use of either of our standard myeloablative preparative condition- PATIENTS AND METHODS ing regimens. The first study evaluated the immunosuppressive Treatment Plan and Supportive Care regimen of CSP, sirolimus, and low-dose MTX (CSP Table 1 describes the treatment plan for each group). Enrollment began in December 2001, and group. Four pediatric patients in the TAC group was terminated in May 2002. The second study evalu- received 13.2 Gy TBI, and all other patients received ated the combination of TAC, sirolimus, and low-dose 12.0 Gy TBI. The first study used CSP as the calci- MTX (TAC group). Patients were enrolled between neurin inhibitor because CSP/MTX was our standard February 2003 and March 2005. Results for both stud- GVHD prophylaxis regimen. Because we initiate ies were analyzed as of March 3, 2007. The trials were CSP therapy 1 day before the infusion of the donor approved by the institutional review board at Fred graft, sirolimus was also started on the same day. Siro- Hutchinson Cancer Research Center (FHCRC), and limus blood levels were targeted at 4-14 ng/mL, which all patients signed consent documents. was slightly higher than the levels targeted by the inves- tigators at the Dana Farber Cancer Institute (DFCI). Patients Because of the high incidence of early-onset GVHD The primary inclusion criteria for patient selection observed in the CSP group, the second study were (1) the use of a preparative conditioning regimen substituted TAC for CSP, with the treatment plan containing total-body irradiation (TBI), followed by and dosing schedule made identical to the regimen cyclophosphamide or busulfan followed by cyclophos- used at the DFCI. When early enrollment in this study phamide; (2) an estimated creatinine clearance $70 showed a similar pattern to that seen in the CSP study, mL/min; (3) total serum bilirubin concentration within there was a concern that immunosuppression with siro- the normal range; and (4) hepatic transaminase levels limus may have been inadequate. Therefore, the proto- less than twice the upper limit of normal. Patients col was revised to increase the target level of sirolimus with diagnoses of chronic myelogenous leukemia in to 5-12 ng/mL. Sirolimus trough levels were measured the chronic phase, de novo acute leukemia in first re- 2-3 times weekly by chromatography-tandem mass mission, and refractory anemia were excluded. HLA- spectrometry, and dose adjustments made to maintain mismatching at DRB1 or DQB1 was not allowed, levels within the targeted range. The tablet formula- although a single class 1 allele disparity was allowed. tion of sirolimus was used throughout both studies. Histocompatibility with the respective donors was In the CSP group, in the absence of toxicity, adminis- determined by high-resolution HLA-A, B, C, and tration of sirolimus was discontinued on day 30. In DRB1 typing and by intermediate-resolution HLA- the TAC group, the dose of sirolimus was gradually DQB1 typing. tapered beginning on day 57, and administration was

Table 1. Treatment Plan

Cyclosporine Group Tacrolimus Group

Conditioning regimen Total body irradiation 12 Gy (6 fractions) 12-13.2 Gy (6-8 fractions) Cyclophosphamide 120 mg/kg (2 divided doses) 120 mg/kg (2 divided doses) or Busulfan 16 doses (targeted 800- 900 ng/mL) 16 doses (targeted 800-900 ng/mL) Cyclophosphamide 120 mg/kg (2 divided doses) 120 mg/kg (2 divided doses) GVHD prophylaxis Sirolimus* 12 mg p.o. loading dose day 21 12 mg p.o. loading dose day 23 followed by 4 mg p.o. daily followed by 4 mg p.o. daily Calcineurin inhibitor Cyclosporine—1.5 mg/kg/q12 h i.v. Tacrolimus—0.02 mg/kg/day i.v. start day 21 start day 23 Methotrexate 5 mg/m2 i.v. days 1, 3, 6, and 11 5 mg/m2 i.v. days 1, 3, 6, and 11 Targeted levels Sirolimus 4-14 ng/mL 3-12 ng/mL or 5-12 ng/mL Calcineurin inhibitor Cyclosporine—150-450 ng/mL Tacrolimus— 5-10 ng/mL *Sirolimus dosing for patients \1.5 m2 was 6 mg/m2 loading dose followed by 2 mg/m2 daily. Sirolimus, a Calcineurin Inhibitor Plus MTX for Prevention of Acute GVHD 533

Table 2. Patient and Transplant Characteristics According to Treatment Table 3. Acute GVHD and c GVHD Group CSP Group TAC Group CSP Group TAC Group Characteristic (n 5 9) (n 5 17) (n 5 9) (n 5 17) Onset of aGVHD, median day after 6 (5-23) 7 (3-100) Patient age, median years (range) 39 (20-65) 33 (5-52) HCT (range) Disease type and stage, n Acute GVHD peak grade, n AML . CR1 or persistent disease 0 3 0-I 2 4 ALL . CR1 or persistent disease 1 7 II 4 10 CML—beyond chronic phase 2 3 III-IV 3 3 AML from MDS 1 1 Chronic GVHD requiring systemic 613 RAEB* 33treatment, n NHL 2 0 None 1 1 HLA typing, n Not evaluated 2 3 HLA-matched 8 16 HLA-mismatched 1 1 Stem cell source, n G-CSF-mobilized blood cells 7 15 Toxicities Marrow 2 2 Assessment of safety focused on identifying toxic- Conditioning regimen, n ities attributable to sirolimus. These included cytope- TBI/cyclophosphamide 5 12 Busulfan/cyclophosphamide 4 5 nia, hyperlipidemia, hypercholesterolemia, hemolytic Donor age, median years (range) 37 (18-48) 33 (27-58) uremic syndrome (HUS), and transaminase elevation. Gender match (donor/recipient) The onset of engraftment was defined as the first of 3 Male/male 3 8 consecutive days after HCT when the absolute neutro- Male/female 1 4 phil count surpassed 0.5 Â 109/L. Female/male 1 5 Female/female 4 0 RESULTS AML indicates acute myeloid leukemia; CR1, first complete remis- sion; ALL, acute lymphoblastic leukemia; CML, chronic mye- Patient and Transplant Characteristics loid leukemia; MDS, myelodysplastic syndrome; RAEB, refractory anemia with excess blasts; NHL, non-Hodgkin lym- All patients had high-risk hematologic malignan- phoma. cies. Twenty-four patients had HLA-matched donors, *One patient had RA after treatment of RAEB2 with 5-azacytidine. and 22 received granulocyte-colony stimulting factor (G-CSF)-mobilized peripheral blood stem cells. Nine discontinued on day 180, unless the patient required patients were given sirolimus with CSP, and 17 were continued treatment for GVHD or experienced toxic- given sirolimus with TAC (Table 2). ity related to sirolimus. All patients received antifungal prophylaxis with GVHD fluconazole. Tacrolimus and CSP levels were drawn Grade II-IV aGVHD was observed in 20 (77%) of 1-3 times per week, and dose adjustments were made the 26 patients, 7 of 9 in the CSP group and 13 of 17 in to maintain therapeutic levels. In the absence of the TAC group (Table 3). Grade II GVHD was GVHD, TAC and CSP doses were tapered starting observed in 14 patients (54%), grade III in 6 (23%), days 50-57 after transplantation, and administration and grade IV in none. GVHD was diagnosed in all 9 was discontinued on day 180. This tapering schedule patients who received busulfan and cyclophospha- is similar to our standard practice. If patients devel- mide. The median onset of GVHD occurred at day 6 oped GVHD, tapering of the calcineurin inhibitor in the CSP group and day 7 in the TAC group. Sixteen was delayed and left to the discretion of the attending of the 20 patients with GVHD had histologic confir- physician. Typically, tapering of CSP or TAC doses mation of the diagnosis, including 13 with gastrointes- was not begun until patients had discontinued primary tinal involvement. Ten patients had positive skin or treatment for acute or chronic GVHD. intestinal biopsies at the time of onset of GVHD or within 1 week after the onset. All patients diagnosed Assessment of GVHD with GVHD required glucocorticoid therapy, and 4 Acute GVHD was graded according to previously required secondary therapy. Because of the need for described criteria [24]. Biopsy samples were obtained, treatment of aGVHD, only 4 patients began tapering when appropriate, to corroborate the clinical diagnosis CSP or TAC doses as planned at week 8 after HCT. of GVHD. Follow-up was censored for evaluation of Chronic GVHD requiring systemic immunosuppres- aGVHD at the time of hematologic relapse or death. sive treatment developed in 6 of 7 patients in the Chronic GVHD was diagnosed according to criteria CSP group and in 13 of 14 patients in the TAC group closely similar to those recommended by the NIH who could be evaluated. Two patients in the CSP Consensus Conference [25]. group and 3 patients in the TAC group could not be 534 T. Furlong et al.

evaluated because of early death (n 5 4) or recurrent Table 4. Sirolimus and Calcineurin Inhibitor Levels and Toxicity malignancy (n 5 1). CSP Group TAC Group (n 5 9) (n 5 17) Sirolimus, CSP, and Tacrolimus Levels Sirolimus level (ng/mL), mean* 11.7 8.8 Twenty-three patients had sirolimus blood levels Sirolimus levels,* n (%) drawn on the day the donor graft infusion was finished . Target range 26 (24) 35 (18) or within 24 hours. Eighteen patients had sirolimus Within target range 80 (74) 133 (71) levels within or above the range targeted for each \ Target range 2 (2) 20 (11) Sirolimus level at onset of group, including 7 of 8 patients in the CSP group, in GVHD, n (%)† whom sirolimus therapy was started on the day before Patients with level . target range 3 (43) 0 (0) transplantation. Mean sirolimus blood levels through Patients with level within 3 (43) 8 (67) day 30 after transplantation (or through the time of target range discontinued administration, if earlier) were 11.7 ng/ Patients with level \ target range 1 (14) 4 (33) Cyclosporine level (ng/mL), mean* 324 — mL in the CSP group and 8.8 ng/mL in the TAC Cyclosporine levels,* n (%) group, with only 2% and 11% of all levels below the . Target range 22 (22) — targeted range in each group, respectively. At the onset Within target range 65 (65) — of GVHD, 5 patients (26%) had sirolimus levels below \ Target range 13 (13) — the target range. The mean CSP and TAC levels for Cyclosporine level at onset of GVHD, n (%) the same 30 day period were 324 ng/mL and 9.6 ng/ Patients with level . target range 1 (14) — mL, respectively. Thirteen percent of all CSP levels Patients with level within 6 (86) — were below the targeted range, and no patients had target range levels below the targeted range at the onset of Patients with level \ target range 0 (0) — GVHD. Twelve percent of all TAC levels were below Tacrolimus level (ng/mL), mean* — 9.6 Tacrolimus levels,* n (%) the targeted range, and 2 patients had levels below the . Target range — 81 (32) targeted range at the onset of GVHD (Table 4). Within target range — 140 (56) \ Target range — 30 (12) Toxicity and Early Discontinuation of Sirolimus Tacrolimus level at onset of GVHD, n (%) Eleven of 26 patients (42%) discontinued adminis- Patients with level . target range — 2 (15) tration of sirolimus early because of adverse events, Patients with level within — 9 (70) including cytopenias (5), HUS or renal failure (4), target range Patients with level \ target range — 2 (15) abnormal liver function tests (1), and hyperlipidemia Reasons for discontinuation of (1). The administration of sirolimus was discontinued sirolimus, n in 1 patient because of an elevated blood level (Table Cytopenia 2 3 4). Nine patients had sirolimus levels drawn at the Renal failure 1 3 time of the adverse event, and 3 had levels above the tar- Abnormal liver function tests 0 1 Elevated sirolimus level 1 0 geted range. In 2 patients, who discontinued sirolimus Hyperlipidemia 0 1 therapy on days 277 and 351, the levels of sirolimus were unknown. Four of 5 patients with cytopenias *Sirolimus, TAC, and CSP levels include all measurements through day 30 after HCT or until administration was discontinued, if earlier. had resolution of neutropenia or thrombocytopenia af- The mean levels are for the same time period. ter the administration of sirolimus was discontinued. †One patient in the TAC Group stopped sirolimus administration One patient with neutropenia had recurrent malig- before the onset of aGVHD. nancy 21 days after the administration of sirolimus was discontinued. Another patient with thrombocytopenia also had evidence of mild HUS, and this likely patients who developed HUS or renal failure, 3 had contributed to the thrombocytopenia. One patient blood levels of sirolimus or the calcineurin inhibitor had elevated alanine and asparate transaminase values above the specified target range at the onset of renal in- in the blood in the absence of GVHD and with normal sufficiency. blood levels of TAC and sirolimus. After discontinuing Five patients in the CSP Group discontinued the administration of sirolimus, the transaminase administration of sirolimus as planned at day 30 after values returned to the normal range. One patient in HCT, and the remaining 4 patients discontinued the CSP group developed rhabdomyolysis with uncer- administration of sirolimus on days 13, 24, 26, and 27 tain attribution to sirolimus. because of presumed toxicity. In the TAC group, Three patients in the TAC group required hemo- administrationofsirolimuswasdiscontinuedatamedian dialysis. One of the patients who developed HUS had of 86 days (range: 9-474) after HCT. With 1 exception, previously experienced renal injury attributed to the all patients developed GVHD before sirolimus ad- cyclophosphamide conditioning regimen. Among 5 ministration was stopped. Among the 25 patients who Sirolimus, a Calcineurin Inhibitor Plus MTX for Prevention of Acute GVHD 535 could be evaluated, engraftment of neutrophils occurred enrolled only adults, whereas we enrolled adults and at a median of 16 days (range: 10-28) after HCT. children. The DFCI patients received a conditioning Nine patients received targeted oral busulfan ther- regimen of cyclophosphamide followed by fractionated apy as part of their preparative conditioning regimen, TBI, whereas patients who received a TBI-based regi- with levels targeted, in most cases, at 800-900 ng/ men at our center were given TBI followed by cyclo- mL. Seven patients had mean busulfan levels within phosphamide. It is unlikely that this scheduling of the targeted range. Two patients had mean levels of therapy would explain the high incidence of aGVHD 923 ng/mL and 952 ng/mL. seen in this study, because all patients who received busulfan and cyclophosphamide also developed aGVHD. Outcome The dose of cyclophosphamide was slightly lower for Nineteen of 21 patients (90%) who could be evalu- DFCI patients than for FHCRC patients, but the ated developed cGVHD. Twenty-one (81%) and 16 TBI exposure in the DFCI study (14 Gy) was slightly patients (62%) were alive at 6 months and 1 year after higher than in our study. The DFCI patients were HCT, respectively (Figure 1). Twelve (46%) remain given clotrimazole and nonabsorbable antibiotics until alive at a median of 1106 days after HCT. Three engraftment, whereas FHCRC patients were given flu- patients had recurrent malignancy and died. Eleven conazole and levofloxacin (adults) or ceftazidime (chil- patients died from other causes: respiratory failure dren) to prevent infection during neutropenia. There (not infection) (4), cardiac arrest (2), hepatic failure were no obvious differences in sirolimus blood levels (2), GVHD (1), infection (1) and multiorgan failure (1). between the 2 studies, and the ranges used in our studies for targeting sirolimus levels were identical to or DISCUSSION higher than those used in the DFCI study. Nearly all patients in our studies had levels of CSP or TAC within In these 2 studies, the addition of sirolimus to or above the targeted range at the onset of GVHD. Of a standard calcineurin inhibitor plus MTX regimen note, the dose of MTX used in these studies was lower provided no detectable improvement in preventing than the standard dose used at our center for GVHD GVHD after unrelated HCT. Accrual in the CSP prophylaxis after HCT with myeloablative condition- group was terminated when 7 of 9 patients developed ing regimens. Although early blood levels of CSP or aGVHD at a median of 6 days after HCT. When 13 TAC and sirolimus were within the targeted range in of 17 patients in the TAC group developed aGVHD most patients, it is possible that the reduced dosing of at a median of 7 days after HCT, the protocol was MTX contributed to inadequate immunosuppression closed to further enrollment. early after transplantation. The immunosuppressive regimen of sirolimus, Our schedule for discontinuing administration of TAC, and MTX for patients in the TAC group of sirolimus, particularly in the CSP Group, was different our study was identical to that used at the DFCI, where from that used in the DFCI study. All patients in the the incidence of aGVHD after unrelated HCT was CSP Group had sirolimus therapy discontinued on or 26% [22]. It is difficult to explain why our results differ before day 30 after transplantation. In the TAC Group so greatly from the DFCI experience. Review of patient administration of sirolimus was discontinued at a me- selection criteria and treatment methods has identified dian of 86 days after transplantation. The impact several differences between the 2 studies, but none of of this schedule on the incidence of aGVHD is not sig- these differences can be easily invoked as likely expla- nificant, as all patients, except 1, in both studies devel- nations for the disparate results. The DFCI study oped GVHD before the discontinuation of sirolimus therapy. It is possible that the grading of GVHD differs between centers, and the threshold severity of GVHD required to prompt treatment may also differ. The incidence of grades II-IV aGVHD after unrelated HCT with myeloablative conditioning at our center has been approximately 80%, substantially higher than the rates reported from other centers [10].We have published evidence suggesting that sensitivity for making the diagnosis of gastrointestinal GVHD might be higher at our center than at other centers [10]. The consistent early presentation of GVHD in the present studies was highly atypical in our experience. The early onset of symptoms might suggest the development of an engraftment syndrome. If so, treat- Figure 1. Relapse and overall and event-free survival for all patients. ment with a short course of high-dose glucocorticoids 536 T. Furlong et al. should have been adequate. This approach was not study may be related to effects of intensive chemother- tested, and the patients were treated with glucocorti- apy before referral for transplantation, limited ability coids per our standard practice for management of to tolerate the conditioning and immunosuppressive aGVHD. Of note, a high incidence of engraftment syn- regimens, as well as the early onset of aGVHD. drome was not mentioned in the report from the DFCI. In conclusion, we found no evidence for improved Risk factors for the development of hyperacute prevention of GVHD after adding sirolimus to the GVHD following HCT have been identified by Saliba combination of methotrexate and a calcineurin inhi- et al. [26]. These factors include a donor/recipient (fe- bitor after unrelated HCT. Moreover, we found an male/male) gender mismatch, the use of HLA-matched unusually high incidence of complications that we be- unrelated donor grafts, the use of a myeloablative pre- lieve were caused by administration of sirolimus. Our parative conditioning regimen, and treatment with results sound a note of caution in future clinical trials more that 5 prior chemotherapy regimens prior to testing the safety and efficacy of sirolimus for GVHD transplantation. All of the patients in these studies re- prophylaxis after unrelated HCT. ceived unrelated donor grafts following a myeloablative preparative conditioning regimen. All had high-risk hematologic malignancies likely requiring multiple ACKNOWLEDGMENTS courses of chemotherapy prior to transplantation. Six This research was supported by grants CA18029, male patients received grafts from female donors. CA 15704, and HL36444 from the National Institutes Although the demographic characteristics of our study of Health, Department of Health and Human Ser- cohort included many of these risk factors, this group is vices, Bethesda, MD. similar to the population of patients who receive unrelated HCT at our institution, yet the onset of GVHD REFERENCES among the patients receiving sirolimus was distinctly earlier than what we normally expect. In a multicenter 1. Sullivan KM. Graft-versus-host disease. In: Blume KG, trial comparing CSP/MTX versus TAC/MTX for Forman SJ, Appelbaum FR, editors. Thomas’ Hematopoietic Cell GVHD prophylaxis following unrelated donor trans- Transplantation, 3rd Ed. Oxford, UK: Blackwell Publishing plantation [3], the median time of onset of GVHD Ltd.; 2004. p. 635-664. was much later than that observed in the current studies. 2. Storb R, Deeg HJ, Whitehead J, et al. Methotrexate and cyclo- In the setting of myeloablative HCT for patients sporine compared with cyclosporine alone for prophylaxis of acute graft versus host disease after marrow transplantation for with high-risk malignancies, it is difficult to attribute leukemia. N Engl J Med. 1986;314:729-735. the occurrence and severity of any specific adverse 3. Nash RA, Antin JH, Karanes C, et al. Phase 3 study comparing event to sirolimus alone. On the other hand, some of methotrexate and tacrolimus with methotrexate and cyclospor- the adverse events that we observed in the current study ine for prophylaxis of acute graft-versus-host disease after mar- have been previously reported in GVHD treatment row transplantation from unrelated donors. Blood. 2000;96: and renal transplant studies with sirolimus [15,18-20]. 2062-2068. For example, neutropenia has been reported to occur 4. Ratanatharathorn V, Nash RA, Przepiorka D, et al. Phase III in 5%-19% of patients and thrombocytopenia in study comparing methotrexate and tacrolimus (Prograf, 9%-57% of patients. In the current studies, 4 of the 5 FK506) with methotrexate and cyclosporine for graft-versus- patients who experienced neutropenia or thrombocy- host disease prophylaxis after HLA-identical sibling bone mar- topenia had resolution of the cytopenia following row transplantation. Blood. 1998;92:2303-2314. 5. Przepiorka D, Khouri I, Ippoliti C, et al. Tacrolimus and mini- discontinuation of sirolimus therapy. In a study evalu- dose methotrexate for prevention of acute graft-versus-host dis- ating the use of sirolimus for treatment of cGVHD, 7 of ease after HLA-mismatched marrow or blood stem cell 19 patients discontinued sirolimus therapy because of transplantation. Bone Marrow Transplant. 1999;24:763-768. adverse events [20]. Six of these 7 patients developed 6. Uberti JP, Ayash L, Braun T, Reynolds C, Silver S, HUS or acute renal insufficiency, and the study was Ratanatharathorn V. Tacrolimus as monotherapy or combined closed to accrual earlier than originally planned be- minidose methotrexate for graft-versus-host disease prophylaxis cause of a high incidence of toxicity. Even with close after allogeneic peripheral blood stem cell transplantation: long- monitoring of blood levels, combination regimens of term effects. Bone Marrow Transplant. 2004;34:425-431. sirolimus and calcineurin inhibitors may have a very 7. Bensinger WI, Martin PJ, Storer B, et al. Transplantation of narrow therapeutic index. bone marrow as compared with peripheral-blood cells from Eleven of the 14 deaths in our study were related to HLA-identical relatives in patients with hematologic cancers. N Engl J Med. 2001;344:175-181. causes other than relapse, with 4 deaths within the first 8. Hansen JA, Gooley TA, Martin PJ, et al. Bone marrow trans- 100 days after transplantation. With the exception of 4 plants from unrelated donors for patients with chronic myeloid patients who died with noninfectious pulmonary com- leukemia. N Engl J Med. 1998;338:962-968. plications, the other causes of death varied, with no 9. Kernan NA, Bartsch G, Ash RC, et al. Analysis of 462 transplan- single dominant cause to explain the high rate of non- tations from unrelated donors facilitated by the National Mar- relapse deaths. The high rate of NMR observed in this row Donor Program. N Engl J Med. 1993;328:593-602. Sirolimus, a Calcineurin Inhibitor Plus MTX for Prevention of Acute GVHD 537

10. Martin PJ, McDonald GB, Sanders JE, et al. Increasingly fre- 19. Couriel DR, Saliba R, Escalon MP, et al. Sirolimus in combina- quent diagnosis of acute gastrointestinal graft-versus-host dis- tion with tacrolimus and corticosteroids for the treatment of ease after allogeneic hematopoietic cell transplantation. Biol resistant chronic graft-versus-host disease. Br J Haematol. Blood Marrow Transplant. 2004;10:320-327. 2005;130:409-417. 11. Dumont FJ, Staruch MJ, Koprak SL, Melino MR, Sigal NH. 20. Johnston LJ, Brown J, Shizuru JA, et al. Rapamycin (sirolimus) Distinct mechanisms of suppression of murine T cell activation for treatment of chronic graft-versus-host disease. Biol Blood by the related macrolides FK-506 and rapamycin. J Immunol. Marrow Transplant. 2005;11:47-55. 1990;144:251-258. 21. Cutler C, Kim HT, Hochberg E, et al. Sirolimus and tacrolimus 12. Sehgal SN. Sirolimus: its discovery, biological properties, and without methotrexate as graft-versus-host disease prophylaxis mechanism of action. Transplant Proc. 2003;35:7S-14S. after matched related donor peripheral blood stem cell trans- 13. Morris RE. Rapamycin: FK506’s fraternal twin or distant plantation. Biol Blood Marrow Transplant. 2004;10:328-336. cousin? Immunol Today. 1991;12:137-140. 22. Antin JH, Kim HT, Cutler C, et al. Sirolimus, tacrolimus, and 14. Groth CG, Backman L, Morales JM, et al. Sirolimus (rapamycin)- low-dose methotrexate for graft-versus-host disease prophylaxis based therapy in human renal transplantation: similar efﬁcacy and in mismatched related donor or unrelated donor transplanta- different toxicity compared with cyclosporine. Sirolimus Euro- tion. Blood. 2003;102:1601-1605. pean Renal Transplant Study Group. Transplantation. 1999;67: 23. Cutler C, Li S, Ho VT, et al. Extended follow-up of methotrex- 1036-1042. ate-free immunosuppression using sirolimus and tacrolimus in 15. Kahan BD. Efﬁcacy of sirolimus compared with azathioprine for related and unrelated donor peripheral blood stem cell trans- reduction of acute renal allograft rejection: a randomized multi- plantation. Blood. 2007;109:3108-3114. centre study. The Rapamune US Study Group. Lancet. 2000; 24. Przepiorka D, Weisdorf D, Martin P, et al. 1994 Consensus con- 356:194-202. ference on acute GVHD grading. Bone Marrow Transplant. 1995; 16. Kahan BD, Knight R, Schoenberg J, et al. Ten years of siroli- 15:825-828. mus therapy for human renal transplantation: the University 25. Filipovich AH, Weisdorf D, Pavletic S, et al. National Institutes of Texas at Houston experience. Transplant Proc. 2003;35: of Health consensus development project on criteria for clinical 25S-34S. trials in chronic graft-versus-host disease: I. Diagnosis and stag- 17. Trotter JF. Sirolimus in liver transplantation. Transplant Proc. ing working group report. Biol Blood Marrow Transplant. 2005; 2003;35:193S-200S. 11:945-955. 18. Benito AI, Furlong T, Martin PJ, et al. Sirolimus (rapamycin) for 26. Saliba RM, DeLima M, Giralt S, et al. Hyperacute GVHD: risk the treatment of steroid-refractory acute graft-versus-host factors, outcomes, and clinical implications. Blood. 2007;109: disease. Transplantation. 2001;72:1924-1929. 2751-2758.