This material may be protected by Copyright law (Title 17 U.S. Code)
`
`TRANSPLANTATION
`
`Etanercept plus methylprednisolone as initial therapy for acute graft-versus-host
`disease
`John E. Levine,1,2 Sophie Paczesny,3 Shin Mineishi,2 Thomas Braun,4 Sung W. Choi,1,2 Raymond J. Hutchinson,1
`Dawn Jones,3 Yasser Khaled,2 Carrie L. Kitko,1,2 Daniel Bickley,3 Oleg Krijanovski,2 Pavan Reddy,2 Gregory Yanik,1,2 and
`James L. M. Ferrara1,2
`
`1Department of Pediatrics, University of Michigan Medical School, Ann Arbor, MI; 2Department of Medicine, University of Michigan Medical School, Ann Arbor,
`MI; 3Blood and Marrow Transplantation Program, University of Michigan Comprehensive Cancer Center, Ann Arbor, MI; 4Department of Biostatistics, School of
`Public Health, University of Michigan, Ann Arbor, MI.
`
`Graft-versus-host disease (GVHD) is a
`principal cause of morbidity following
`allogeneic hematopoietic cell transplanta-
`tion (HCT). Standard therapy for GVHD,
`high-dose steroids, results in complete
`responses (CRs) in 35% of patients. Be-
`cause tumor necrosis factor-␣ (TNF␣) is
`an important effector of experimental
`GVHD, we treated patients with new-
`onset GVHD with steroids plus the TNF␣
`inhibitor etanercept on a previously re-
`ported pilot trial (n ⴝ 20) and a phase 2
`trial (n ⴝ 41). We compared their out-
`Introduction
`
`comes with those of contemporaneous
`patients with GVHD (n ⴝ 99) whose initial
`therapy was steroids alone. Groups were
`similar with respect to age, conditioning,
`donor, degree of HLA match, and severity
`of GVHD at onset. Patients treated with
`etanercept were more likely to achieve
`CR than were patients treated with ste-
`roids alone (69% vs 33%; P < .001). This
`difference was observed in HCT recipi-
`ents of both related donors (79% vs 39%;
`P ⴝ .001) and unrelated donors (53% vs
`26%; P < .001). Plasma TNFR1 levels, a
`
`biomarker for GVHD activity, were el-
`evated at GVHD onset and decreased
`significantly only in patients with CR. We
`conclude that etanercept plus steroids as
`initial therapy for acute GVHD results in a
`substantial majority of CRs. This trial was
`referenced at www.clinicaltrials.gov as
`NCT00141713.
`(Blood.
`2008;111:
`2470-2475)
`
`© 2008 by The American Society of Hematology
`
`Allogeneic hematopoietic cell transplantation (HCT) is a curative
`treatment for a number of hematologic malignancies and genetic
`disorders. Despite the routine use of immunosuppressive agents
`that target T cells, such as calcineurin inhibitors, up to 50% of HCT
`recipients still experience significant graft-versus-host disease
`(GVHD) that requires treatment with high-dose systemic steroids,
`which have been the primary therapy of GVHD for more than
`25 years.1 The risk of mortality in patients who do not respond
`completely to initial therapy climbs exponentially,2,3 but complete
`response (CR) rates have remained approximately 35%.4-6
`Animal models have established that the pathophysiology of
`GVHD involves complex immunologic interactions between cellu-
`lar effectors, such as cytotoxic T lymphocytes and soluble effec-
`tors, such as inflammatory cytokines.7 One of the most important
`inflammatory cytokines involved in this process is tumor necrosis
`factor-␣ (TNF␣), which mediates GVHD both through the amplifi-
`cation of donor immune response to host tissues as well as direct
`toxicity to target organs.8,9 These preclinical data served as the
`rationale to use an anti-TNF␣ agent, either infliximab or etaner-
`cept, to treat steroid-refractory GVHD, with complete remissions
`achieved in 18% to 62% of patients.10-13 Infliximab, a monoclonal
`antibody directed at TNF␣, binds to both soluble and membrane-
`bound TNF␣, resulting in clearance of both circulating TNF␣ and
`T cells.14 Etanercept, a soluble dimeric TNF␣ receptor 2, competes
`for TNF␣ binding and renders it inactive.14,15 This mechanism of
`
`action combined with its relative ease of administration by
`subcutaneous injection and generally minor side effects,15 make
`etanercept attractive as primary GVHD therapy. We therefore
`tested the hypothesis that etanercept plus systemic corticosteriods
`would result in CR rates of at least 50% when used as primary
`therapy for GVHD.
`
`Methods
`
`Study cohort
`
`A total of 738 patients who underwent allogeneic HCT transplantation at
`the University of Michigan between April 1, 2000, and September 30, 2006,
`provided informed consent for the collection of blood and plasma samples
`during their HCT course. Blood samples were obtained at onset of GVHD
`and prior to initiation of systemic therapy and 4 weeks later in 160 (55%) of
`291 patients who developed acute GVHD requiring treatment during this
`time period. Between September 2001 and September 2006, 61 of these
`patients provided additional informed consent and were enrolled in one of
`2 prospective consecutive clinical trials to test the combination of etaner-
`cept (supplied by Amgen, Thousand Oaks, CA) and methylprednisolone as
`the initial treatment of GVHD during this time period. The first trial
`enrolled 20 patients to establish the safety of etanercept administration as
`part of primary GVHD therapy.16 A follow-up phase 2 clinical trial with
`identical eligibility criteria and identical treatment design enrolled an
`additional 41 patients. Based on the observed CR rate of 75% in the pilot
`
`Submitted September 14, 2007; accepted November 21, 2007. Prepublished
`online as Blood First Edition paper, November 27, 2007; DOI 10.1182/blood-
`2007-09-112987.
`
`The publication costs of this article were defrayed in part by page charge
`payment. Therefore, and solely to indicate this fact, this article is hereby
`marked ‘‘advertisement’’ in accordance with 18 USC section 1734.
`
`The online version of this article contains a data supplement.
`
`© 2008 by The American Society of Hematology
`
`2470
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`BLOOD, 15 FEBRUARY 2008 䡠 VOLUME 111, NUMBER 4
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`

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`BLOOD, 15 FEBRUARY 2008 䡠 VOLUME 111, NUMBER 4
`
`ETANERCEPT PLUS METHYLPREDNISOLONE FOR GVHD
`
`2471
`
`trial,16 the follow-up study was designed to provide 80% power that the true
`CR rate was at least 50% when results from both trials were combined. The
`clinical trials were approved and reviewed by the University of Michigan
`Institutional Review Board (IRB) and the University of Michigan Cancer
`Center Data and Safety Monitoring Board. Eligibility criteria included
`diagnosis of new onset acute GVHD, clinical grades 2 to 4, that was
`confirmed on biopsy and treated for less than 72 hours with 2 mg/kg/day
`methylprednisolone prior to first dose of etanercept; an absolute neutrophil
`count greater than 500/mm3 for at least 3 days; absence of uncontrolled
`infection; and absence of inotropic agents for blood pressure support. The
`group of contemporaneous patients (99 of 160) who had blood samples
`obtained at onset of GVHD and 4 weeks later, whose initial GVHD therapy
`was steroids alone and who did not participate in an etanercept trial, served
`as a control group for the study subjects. Patients who did not have blood
`samples obtained prior to initiation of steroids for treatment of GVHD were
`not considered as control patients. To be a control patient, the patient must
`also have met the eligibility critieria for participation on an etanercept
`treatment trial. The reasons for lack of participation in 99 patients include
`the absence of an IRB-approved trial at the time of onset of GVHD
`(n ⫽ 49), patient/physician refusal (n ⫽ 23), and more than 72 hours of
`steroid therapy for GVHD (n ⫽ 22) and simultaneous infection (n ⫽ 5). An
`additional 191 patients who underwent transplantation between April 2000 and
`September 2006 who never developed GVHD and who had blood samples
`drawn at the appropriate time after HCT form a non-GVHD control population.
`
`GVHD treatment
`
`GVHD prophylaxis for all recipients included tacrolimus (titrated to a goal
`level of 8 to 12 ng/mL). After myeloablative HCT, patients also received
`minidose methotrexate (5 mg/m2 per dose intravenously on days 1, 3, 6, and
`11). After reduced-intensity HCT, patients also received mycophenolate
`mofetil (500 mg/m2 per dose every 8 hours) from days 0 to 28. All patients
`who were treated for GVHD received methylprednisolone 2 mg/kg per day
`and continued their GVHD prophylaxis agents at
`therapeutic dosing.
`Generally, steroids continued at the starting dose for the first 7 days and
`were then tapered as tolerated in patients with CRs. A CR was defined as the
`resolution of all manifestations of GVHD (all organs stage 0). Second-line
`agents were added for progression of GVHD, for lack of response after 5 to
`7 days, or for slow responses as deemed necessary by the treating physician.
`A flare of GVHD was defined as achievement of CR with later worsening of
`GVHD by at least one clinical grade that required additional systemic
`therapy. Acute and chronic GVHD was staged and graded according to
`published GVHD scales.17
`The primary endpoint of the 2 etanercept trials was the complete
`response rate at 4 weeks. Patients who required additional therapy for
`GVHD in the first 4 weeks were considered treatment failures for the
`primary endpoint. None of these patients ever achieved a CR. Patients who
`died before day 28 regardless of GVHD status at the time of death were also
`regarded as treatment failures. A total of 20 patients (4 patients treated with
`etanercept plus steroids and 16 patients treated with steroids alone) had
`improvement in their GVHD scores by day 28 but not a CR (and therefore
`primary endpoint failures), but went on to a CR after day 28. In 5 of
`20 patients (all in the steroids-alone group), additional therapy was begun
`after day 28, and these CRs were included in analyses other than the
`primary endpoint analysis. No other patient receiving additional GVHD
`therapy achieved a CR.
`Etanercept was given subcutaneously twice weekly for 8 weeks at a
`dose of 0.4 mg/kg per dose (maximum dose, 25 mg) based on historic data
`from our center that CRs can occur up to 8 weeks following initiation of
`systemic treatment. Doses were held and not replaced until resolution of
`bacteremia, hemodynamic instability, or fever above 100.5°F for more than
`5 days. A total of 15 patients had one or 2 doses held for fever or for a
`positive blood culture. Etanercept was permanently discontinued in patients
`whose GVHD required additional systemic therapy in the first 28 days of
`treatment; these patients were categorized as nonresponders. Etanercept
`was also permanently discontinued in 5 patients who missed more than
`2 doses of etanercept. A total of 4 patients, all evaluable for the primary
`endpoint, were removed from the study after day 28 because of disease
`relapse or progression. All patients who enrolled in the study were included
`in the analyses even if they did not complete study treatment.
`
`Infection prophylaxis consisted of norfloxacin 400 mg twice daily and
`fluconazole 100 mg daily. Infection prophylaxis changed in 2005. The last
`36 patients treated with etanercept plus steroids and the last 48 patients
`treated with steroids alone received levofloxacin 500 mg daily instead of
`norfloxacin and voriconazole 200 mg twice daily instead of fluconazole.
`IgG levels were monitored monthly beginning on day 100 and intravenous
`immunoglobulin 400 mg/kg replacement therapy was given for IgG levels
`lower than 4.0 g/L (400 mg/dL). Patients received sulfamethoxazole/
`trimethoprim or pentamadine for PCP prophylaxis starting on day 30.
`Acyclovir 400 mg twice daily was given for herpes simplex and varicella-
`zoster prophylaxis in seropositive patients. Cytomegalovirus DNA was
`monitored weekly by quantitative polymerase chain reaction (PCR), and
`preemptive therapy with antiviral agents were begun in the event of a
`positive assay.
`
`Cytokine analysis
`
`Blood obtained at the time of onset of GVHD and 4 weeks later was
`separated into cellular and plasma components and frozen for later batch
`processing. The plasma component of each blood sample was analyzed for
`soluble TNF receptor 1 (sTNFR1) levels by the Immunologic Monitoring
`Core Laboratory of the University of Michigan Cancer Center using
`cytokine enzyme-linked immunosorbent assay (R&D Systems, Minneapo-
`lis, MN). The assays were performed according to the manufacturer’s
`protocol, and all samples and standards were run in duplicate.
`
`Statistical analysis
`
`Comparisons of patient characteristics between treatment groups were
`based upon a 2-sample t test for continuous variables and a chi-squared test
`of association for categoric variables. Comparison of infection and 4-week
`CR rates between treatment groups were based upon a chi-squared test of
`association. Mean and median steroid dose at 4 weeks were compared by
`2-sample t test and nonparametric tests, respectively. Time to complete
`response was estimated using standard competing risks methodology.18 The
`significance of differences in mean TNFR1 levels was based upon a paired t
`test. Effects of potential factors such as patient age,
`transplantation
`conditioning regimen (myeloablative vs reduced intensity), degree of HLA
`match, donor source (related vs unrelated), and post-HCT day of onset of
`GVHD on CR at 4 weeks were examined in a Cox proportional-hazards
`model. All reported P values are 2-sided.
`
`Results
`
`Patient characteristics
`
`The characteristics of patients without GVHD (n ⫽ 191), patients
`with GVHD treated with steroids alone (n ⫽ 99), and patients with
`GVHD who were treated with etanercept plus steroids (n ⫽ 61) are
`provided in Table 1. Patients in the 2 treatment groups were similar
`with respect to age, transplantation conditioning intensity (myeloa-
`blative vs reduced intensity), donor type (related vs unrelated),
`degree of match (matched vs mismatched), and severity of organ
`involvement of GVHD at onset. There were no statistically
`significant differences between patients treated on the pilot trial and
`the follow-up phase 2 trial (Table S1, available on the Blood website; see
`the Supplemental Materials link at the top of the online article).
`
`Clinical responses
`
`The primary endpoint of the clinical trial was CR (resolution of all
`GVHD manifestations) at 4 weeks after the initiation of treatment.
`Patients treated with etanercept plus steroids were significantly
`more likely to achieve CR 4 weeks later than were patients treated
`with steroids alone (69% vs 33%; P ⬍ .001; Figure 1A). The
`benefit of etanercept persisted so that by 12 weeks after initiation of
`GVHD treatment, 77% of patients treated with etanercept plus
`
`

`

`2472
`
`LEVINE et al
`
`BLOOD, 15 FEBRUARY 2008 䡠 VOLUME 111, NUMBER 4
`
`Table 1. Patient characteristics according to GVHD treatment group and for non-GVHD controls
`Non-GVHD controls
`Steroids alone
`Etanercept plus steroids
`n ⴝ 191
`n ⴝ 99
`n ⴝ 61
`
`P for difference between 2
`treatment groups
`
`Median age, y (range)
`Patients ⬍ 18 y, %
`Myeloablative conditioning, %
`Unrelated donor, %
`Mismatched, %
`Advanced malignancy, %*
`Nonmalignant disease, %
`GVHD at start of treatment, %
`Grade 2
`Grades 3-4
`Skin
`Skin only target organ
`Liver
`GI tract
`Upper GI only target organ
`
`44(1-69)
`15
`77
`32
`6
`22
`6
`
`—
`—
`—
`
`—
`—
`—
`
`49(1-71)
`9
`68
`46
`16
`25
`1
`
`69
`31
`69
`49
`15
`44
`5
`
`51(7-65)
`10
`61
`31
`11
`31
`0
`
`67
`33
`61
`34
`15
`61
`3
`
`.77
`.88
`.37
`.06
`.60
`.42
`.43
`
`.84
`
`.60
`.08
`.47
`.16
`.60
`
`GI indicates gastrointestinal; and —, not applicable.
`*Chronic myelogenous leukemia in chronic phase, myelodysplastic syndrome, multiple myeloma, acute leukemia in CR, and lymphoma in partial response (PR) or CR
`were considered nonadvanced disease for the purposes of analysis. Leukemia or lymphoma in untreated relapse or refractory to therapy were considered advanced disease
`for the purposes of analysis.
`
`steroids had achieved CR compared with 50% of patients treated
`with steroids alone (P ⬍ .001). Differences in steroid dosing did
`not account for differences in the response rate at day 28 because
`the mean steroid dose on day 28 in patients treated with etanercept
`plus steroids was 0.9 mg/kg (median, 0.7 mg/kg; range, 0.05-2
`mg/kg per day), similar to the 1 mg/kg (median, 0.9 mg/kg; range,
`0-2 mg/kg per day) in patients initially treated with steroids alone.
`We performed a univariate analysis comparing CR rates according
`to conditioning regimen (myeloablative vs reduced intensity), a
`factor known to influence GVHD.19 Conditioning regimen did not
`affect CR rates in patients treated with etanercept plus steroids
`
`(Table S3). In multivariate analyses, the only 2 variables associated
`with increased likelihood of CR were the use of etanercept and a
`related donor stem-cell source. Patient age, conditioning regimen,
`degree of HLA match, and the day of onset of GVHD all had no
`statistically significant association with response. The CR rates in
`the 9 patients who received less than 8 doses of etanercept by day
`28 (primary endpoint) were not statistically different from the
`remaining patients (P ⫽ .33).
`All patients without CR to initial treatment were treated with
`additional agents, most commonly mycophenolate, at a median of
`18 days (range, 3-47 days) from initiation of treatment. None of
`
`A Time to CR :
`.,
`u
`C .,
`"0 ·u
`.E
`~
`i
`'3
`E
`::,
`(J
`
`.-(cid:173)
`---
`
`All recipients
`
`J---... ---.. -_.
`
`___ .J.J!_
`
`P < .0001
`
`0 1 2 3 4 5 6 7 8 9 10 11 12
`WEEKS
`
`D Overall Survival
`
`All recipients
`
`B
`.,
`u
`C .,
`"0 ·u
`.E .,
`>
`i
`:i
`E
`::,
`0
`
`E
`
`Related donor recipients
`
`C
`
`Unrelated donor recipients
`
`.-----·
`_,
`,,••
`T
`.. f ___ _...
`.:-
`····
`.. ----
`
`P= .001
`
`0 1 2 3 4 5 6 7 8 9 10 11 12
`WEEKS
`
`Related donor recipients
`
`F
`
`-, -----..
`
`P= .51
`
`<ii
`> -~
`
`::::,
`(/)
`
`c
`Q)
`~
`Q) c..
`
`I----.-----------------.
`-·
`
`P = .0005
`
`•• ----
`
`--··
`
`0 1 2 3 4 5 6 7 8 9 10 11 12
`WEEKS
`
`Unrelated donor recipients
`
`·(cid:173) ---~. ·--
`·--·(cid:173) '•--·-.. ··---·--------.
`
`P = .05
`
`0
`
`30
`
`60
`
`90
`DAYS
`
`120
`
`150
`
`180
`
`0
`
`30
`
`60
`
`90
`DAYS
`Figure 1. Time to CR for patients with GVHD treated with steroids alone or etanercept plus steroids. (A) Time to CR for all patients with GVHD treated with steroids alone
`(n ⫽ 99; dotted line) or etanercept plus steroids (n ⫽ 61; solid line). (B) Time to CR for patients who underwent related-donor HCT treated with steroids alone (n ⫽ 53) or with
`etanercept plus steroids (n ⫽ 42). (C) Time to CR for patients who underwent unrelated-donor HCT treated with steroids alone (n ⫽ 46) or with etanercept plus steroids
`(n ⫽ 19). The 95% confidence intervals for CR rate at 4 weeks are shown as error bars in panels A to C. Overall survival curves through 6 months from initiation of GVHD
`treatment by treatment group for all patients (D), patients who underwent related-donor HCT (E), and patients who underwent unrelated-donor HCT (F). The 95% confidence
`intervals for survival at 6 months are shown as error bars in panels D to F.
`
`120
`
`150
`
`180
`
`90
`DAYS
`
`0
`
`30
`
`60
`
`120
`
`150
`
`180
`
`

`

`BLOOD, 15 FEBRUARY 2008 䡠 VOLUME 111, NUMBER 4
`
`ETANERCEPT PLUS METHYLPREDNISOLONE FOR GVHD
`
`2473
`
`Table 2. CR rates at 4 weeks according to treatment group
`Steroids alone
`Etanercept plus steroids
`
`Overall, no. (%)
`95% CI
`Skin
`95% CI
`Liver
`95% CI
`GI
`95% CI
`
`33/99 (33)
`24, 42
`32/68 (47)
`35, 59
`3/15 (20)
`0, 40
`21/44 (48)
`33, 63
`
`42/61 (69)
`57, 80
`30/37 (81)
`68, 94
`6/9 (67)
`36, 98
`29/37 (78)
`65, 92
`
`Table 3. Infection rates according to treatment group
`Steroids alone
`Etanercept plus steroids
`(n ⴝ 99)
`(n ⴝ 61)
`
`Bacterial, no. (%)
`Gram-positive
`Gram-negative
`Anaerobic
`Fungal, no. (%)
`Viral, no. (%)
`
`84 (85)
`22 (22)
`4 (4)
`19 (19)
`33 (33)
`
`49 (80)
`13 (21)
`2 (3)
`17 (28)
`19 (31)
`
`P
`
`⬍ .001
`
`⬍ .001
`
`.03
`
`.005
`
`P
`
`.46
`.89
`.79
`.20
`.78
`
`these patients achieved a CR by day 28. The higher response rate
`seen in patients treated with etanercept plus steroids translated into
`improved survival at 100 days from initiation of GVHD treatment
`(etanercept plus steroids, 82% vs steroids alone, 66%; P ⫽ .04). At
`6 months from initiation of treatment, a higher proportion of
`patients treated with etanercept plus steroids were still alive 69%,
`compared with 55% of patients treated with steroids alone, but this
`difference did not meet the criteria for statistical significance
`(P ⫽ .08; Figure 1D).
`Because of the trend toward an increased proportion of unre-
`lated donor transplantations in the group treated with steroids
`alone, we analyzed the results by stem cell source. The superiority
`of etanercept was evident in transplant recipients from both related
`donors (79% vs 39%; P ⫽ .001; Figure 1B) and unrelated donors
`(53% vs 26%; P ⬍ .001; Figure 1C). This latter difference is
`particularly noteworthy, because studies have shown that acute
`GVHD in recipients of unrelated donor transplants is more difficult
`to treat than GVHD in recipients of related donor transplants.4 As
`can be seen in the time to CR curves, different response patterns
`were observed between recipients of related and unrelated donors.
`The time to CR was significantly faster in recipients of related
`donor transplants who were treated with etanercept plus steroids,
`but by 12 weeks, nearly equivalent proportions of patients in both
`groups achieved a CR (etanercept plus steroids, 80%; steroids
`alone, 70%). It is therefore not surprising that survival 6 months
`from initiation of GVHD treatment was similar in both treatment
`groups (Figure 1E). Recipients of unrelated donor transplants who
`failed to achieve a CR by day 28 were likely to never achieve CR
`(Figure 1C). The superior efficacy of etanercept plus steroids in this
`group thus translated into a survival advantage 6 months later
`(P ⫽ .05; Figure 1F).
`Superior CR rates for each GVHD target organ (skin, liver,
`gastrointestinal tract) were observed in patients 4 weeks following
`treatment with etanercept plus steroids (Table 2). Furthermore,
`patients treated with etanercept plus steroids were twice as likely to
`achieve CR whether the grade of GVHD at presentation was grade
`2 (75% vs 37%) or grades 3 to 4 (50% vs 26%). There were no
`statistically significant differences in CR rates between patients
`treated on the pilot trial and the follow-up phase 2 trial (Table S2).
`Importantly, most patients who achieved a CR remained in
`remission: Only 10% (4 of 41) of responding patients treated with
`etanercept plus steroids experienced a flare of GVHD. This rate of
`recurrence is similar to that in patients treated with steroids alone
`(12%; 4 of 33 patients). Chronic GVHD developed in 57% of
`patients treated with etanercept plus steroids (extensive: 53%),
`which was not statistically different from the 56% incidence of
`chronic GVHD (extensive: 53%) observed in patients treated with
`steroids alone (Figure S1).
`We also examined relapse rates by the overall incidence of
`relapse in patients who underwent transplantation for malig-
`nancy without adjustment or censoring. Although the rate of
`
`relapse in patients treated with etanercept plus steroids (10 of
`61; 16%) was lower than that observed in patients treated
`with steroids alone (21 of 98; 21%), the difference was not
`statistically significant (P ⫽ .44).
`
`Infections
`
`The infection rates in the first 100 days from initiation of GVHD
`treatment were not different between patients treated with etaner-
`cept plus steroids or treated with steroids alone for bacterial,
`invasive fungal or viral
`infections (Table 3). There were no
`mycobacterial infections observed in any patients.
`
`Plasma levels of TNFR1 as a biomarker for GVHD response to
`treatment
`
`TNFR1 levels in the plasma are reproducibly measured, correlate
`with levels of TNF␣,20 and are elevated in patients who develop
`GVHD.21 Plasma TNFR1 levels in patients at the onset of GVHD
`were more than 3 times higher than in patients without GVHD
`(Figure 2; P ⬍ .001). As expected, TNFR1 levels at the onset of
`GVHD were equivalent in both treatment groups of patients. In the
`group who received steroids alone, TNFR1 levels fell by more than
`50% toward baseline after 4 weeks of treatment in the 33% of
`patients who achieved a CR, but did not change in the remaining
`67% of patients (P ⫽ .001). Results were strikingly similar in the
`
`P = .001
`P= .009
`~ ~
`
`10000
`
`8000
`
`6000
`
`4000
`
`2000
`
`0
`
`.J
`E
`CJ
`C.
`
`... 0::
`u. z
`
`I-
`
`No GVHD Onset CR NoCR Onset CR NoCR
`
`Percent
`
`Treatment
`
`I 100% 133% s1o;. I i 100% 169% 31 % I
`
`Steroids alone
`
`Steroids + etanercept
`
`Figure 2. Mean TNFR1 plasma levels at initiation of GVHD treatment and 4
`weeks later. Mean TNFR1 plasma levels were significantly lower in patients without
`GVHD (䡺; n ⫽ 190) at time points similar to the onset of GVHD (f) compared with
`patients treated with steroids alone (n ⫽ 99) and etanercept plus steroids (n ⫽ 61;
`P ⬍ .001). At 4 weeks after initiation of treatment, mean TNFR1 plasma levels in
`D
`) were significantly lower than at initiation of treatment both in
`patients in CR (
`patients receiving steroids alone (33 of 99; 33%) and patients receiving etanercept
`) were unchanged in
`plus steroids (42 of 61; 69%). Mean TNFR1 plasma levels (
`patients not in CR in both patients treated with steroids alone (66 of 99; 67%) and
`patients treated with etanercept plus steroids (19 of 61; 31%). Error bars are mean
`plus or minus SEM.
`
`-
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`group who received etanercept plus steroids: TNFR1 levels fell
`significantly in the 69% of patients achieving a CR after 4 weeks
`of therapy, but did not change in the 31% who still had active
`disease (P ⫽ .009).
`
`Discussion
`
`Current standards for the treatment of acute GVHD rely primarily
`on steroids alone as initial therapy, and reserve additional agents for
`steroid refractory disease. Previous studies in steroid refractory
`GVHD have shown that anti-TNF␣ agents have significant effi-
`cacy, but most patients still die from GVHD or its complica-
`tions.10-13 Our results show that the combination of etanercept plus
`steroids as initial treatment for GVHD results in significantly better
`CR rates 4 weeks later compared with steroids alone. These CRs
`were durable and could not be explained by differences in the
`steroid dose at 4 weeks. Of note, etanercept plus steroids improved
`the outcome for recipients of both related donor and unrelated
`donor transplants, which translated into significantly improved
`survival at 6 months for unrelated donor transplant recipients. The
`improvements in outcome were realized without an increased
`incidence of serious infections, chronic GVHD, or relapse. These
`comparisons are based on nonrandomized patient populations, and
`factors other than treatment with etanercept may have contributed
`to the observed results. Although the similarity in outcomes for the
`patients treated with steroids alone are very similar to published
`results from other centers,4-6 a prospective multicenter randomized
`study will be necessary to confirm these encouraging results.
`Animal studies have demonstrated that TNF␣ plays a critical role in
`both the gastrointestinal tract22,23 and the skin,24 but its role in the liver is
`more controversial.8,22 In our study, the addition of etanercept to steroids
`resulted in high response rates in all 3 target organs, suggesting that
`TNF␣ is involved in the clinical pathophysiology of all 3. Moreover, our
`data confirm mechanistic studies of GVHD pathophysiology in animal
`models that have delineated both TNF␣-dependent and TNF␣-
`independent pathways of disease,8 because TNF␣ inhibition increases
`response rates but does not completely eliminate GVHD. All patients
`who received etanercept also received steroids, and therefore the relative
`importance of these pathways in clinical GVHD remains to be
`determined in future studies.
`One potential concern regarding the use of TNF inhibitors to
`treat GVHD is an increased risk of infections, particularly mycobac-
`terial and fungal infections.25 Invasive Aspergillus infections have
`been associated with use of the anti-TNF␣ monoclonal antibody
`infliximab for treatment of GVHD in 2 retrospective studies
`involving a total of 32 patients.11,26 Although the overall incidence
`of infection was significant, as would be expected in patients with
`GVHD receiving high-dose steroids, we did not observe any
`significant difference in infection rates, including fungal infections,
`in patients also treated with etanercept. The discrepancy between
`the 2 studies may be due to potential differences in the mechanism
`of action of the 2 drugs: infliximab can induce systemic elimination
`and clearance of monocytes and macrophages that express mem-
`brane-bound TNF␣, whereas etanercept does not.27 In addition, the
`recent availability of more effective prophylactic agents against a
`broad range of fungal species may have contained the overall
`infectious risk. We also did not observe increased morbidity and
`mortality from etanercept in patients who developed Gram-positive
`infections, which differs from prior reports in other disease settings.28
`Elevated TNFR1 plasma levels correlated with active GVHD
`both before and after treatment. TNFR1 levels in patients achieving
`
`CR was the same in both treatment groups, although the proportion
`of patients achieving CR in the etanercept group was significantly
`higher (Figures 1,2). These data raise the intriguing possibility that
`clinical GVHD manifestations occur above a certain critical
`threshold of TNFR1 levels. Reduction below that threshold, by any
`means, induces a CR; etanercept plus steroids was able to achieve
`that reduction twice as often as steroids alone. Interestingly,
`TNFR1 levels remained high in patients whose GVHD did not
`remit despite TNF inhibition. It is possible the TNF␣ in these
`patients was not completely neutralized, and that higher doses of
`etanercept than used in our study might be beneficial for some of
`these patients. We do not favor this explanation because analysis of
`a limited number of patients treated with etanercept demonstrated
`circulating TNFR2 levels at least 10-fold higher than physiologic
`levels (data not shown). An alternative explanation for the persis-
`tently high TNFR1 levels in patients with persistent GVHD is that
`some cellular effectors continue to express membrane-bound
`TNF␣ and shed TNFR1 despite the quenching of soluble TNF␣ by
`etanercept. This explanation is supported by a recent animal study
`demonstrating that although soluble TNF␣ is responsible for a major
`portion of GVHD morbidity and mortality, donor cells that express
`membrane-bound TNF␣ still cause significant clinical GVHD.29
`The number of unrelated donor transplantations continues to
`increase annually, and GVHD following unrelated donor HCT is
`more prevalent, more severe, and less responsive to treatment
`compared with related donor HCT. In this study, the post-GVHD
`survival for unrelated donor hematopoietic cell transplant recipi-
`ents treated with etanercept and steroids was similar to that of
`related donor HCT recipients, suggesting that
`treatment with
`etanercept and steroids may be particularly useful in the unrelated
`donor HCT setting.
`
`Acknowledgments
`
`We thank the patients, their families, and the clinical personnel who
`participated in this study; Jennifer Lay-Luskin and the BMT
`Program Team at the Clinical Trials Office at the University of
`Michigan for outstanding data collection and management; and the
`BMT Program research nurses for research support, without which
`this study would not have been possible.
`This work was supported by grants from the Food and Drug
`Administration (5R01FD002397-03-2), the National Institutes of
`Health (2P01CA039542-18 and 5P30CA046592), the Doris Duke
`Charitable Foundation, and Amgen. J.L.M.F. is the recipient of a
`Doris Duke Distinguished Clinical Scientist Award.
`
`Authorship
`
`Contribution: J.E.L., T.B., R.J.H., and J.L.M.F. designed and
`planned the study. T.B. was the study statistician. S.P. performed
`the cytokine assays. S.P., S.W.C., D.J., and C.K. were in charge of
`data collection and quality assurance. All authors participated in
`writing the report.
`Conflict-of-interest disclosure: J.E.L. reports research grant
`support from Amgen. J.L.M.F. reports research grant and sympo-
`sium support from Amgen. The other authors declare no competing
`financial interests.
`Correspondence: John E. Levine, 1500 E Medical Center Dr,
`5303 Cancer Center, Ann Arbor, MI, 48109-0941; e-mail:
`jelevine@umich.edu.
`
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