Infections after the use of alemtuzumab in solid organ transplant recipients: a comparative study

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Diagnostic Microbiology and Infectious Disease 66 (2010) 7 – 15 www.elsevier.com/locate/diagmicrobio

Infections after the use of alemtuzumab in solid organ transplant recipients: a comparative study Nasia Safdara,b,⁎, Jeannina Smitha,b , Valerie Knasinskia,b , Colleen Sherkowa,b , Casey Herrfortha,b , Stuart Knechtlec , David Andesa,b,d a

Department of Medicine, Section Infectious Diseases, University of Wisconsin, Madison, WI 54792, USA b Department of Surgery, University of Wisconsin, Madison, WI 54792, USA c Section of Transplant Surgery, University of Wisconsin, Madison, WI 54792, USA d Medical Microbiology, University of Wisconsin, Madison, WI 54792, USA Received 18 December 2007; accepted 20 August 2009

Abstract We undertook a retrospective cohort study comparing infection in solid organ transplant recipients receiving alemtuzumab (n = 726) versus basiliximab (n = 215) or antithymocyte globulin (ATG) (n = 85). Eighty-one percent of patients had kidney transplants. Overall, 33% of patients in the alemtuzumab group (240/724) developed infection compared with 40% (87/215) in the basiliximab group (odds ratio [OR], 0.72; 95% confidence interval [CI], 0.53–0.99; P = .04). The frequency of infection was similar in the alemtuzumab and ATG groups (33% versus 36%, respectively, P = .53). The frequency of fungal infections, most caused by Candida spp., was similar in the alemtuzumab and basiliximab groups (10% versus 9%); disseminated fungal infection occurred in 68% of the patients with fungal infection receiving alemtuzumab and in 30% of the patients with fungal infection receiving basiliximab (OR, 4.76; 95% CI, 1.58–14.28; P = .003). Basiliximab posed a higher risk than alemtuzumab for infection. Disseminated candidal infections were more common in patients receiving alemtuzumab. © 2010 Elsevier Inc. All rights reserved. Keywords: Alemtuzumab; Infection; Transplantation; Fungal; Bacterial; Viral

1. Introduction Major advances in transplantation techniques and understanding of transplant biology have greatly improved the survival of renal transplant recipients. Each year, approximately 15 000 renal transplants are performed in the United States, with a 5-year survival exceeding 80% (http://www. unos.org [United Network for Organ Sharing, n.d.]). However, infection remains a major challenge in the solid organ transplant recipient and now exceeds rejection as the precipitating factor for hospitalization in patients in the first 2 years after a solid organ transplant (Dharnidharka et al., 2004). In recent years, lymphocyte depletion has been used with increasing frequency as an induction strategy after transplantation and for treatment of rejection (Shapiro et al.,

⁎ Corresponding author. Tel.: +1-608-263-1545; fax: +1-608-263-4464. E-mail address: [email protected] (N. Safdar). 0732-8893/$ – see front matter © 2010 Elsevier Inc. All rights reserved. doi:10.1016/j.diagmicrobio.2009.08.017

2005a, 2005b). Alemtuzumab (Campath-IH; Berlex, Montville, NJ) is a humanized monoclonal antibody directed against CD52, a cell surface antigen expressed on B and T lymphocytes, monocytes, and natural killer (NK) cells (Ferrajoli et al., 2001; Flynn and Byrd, 2000). It is a powerful cytolytic agent and is used therapeutically in bone marrow transplantation (Dumont, 2002; Hale, 2002) and several autoimmune diseases (Marsh and Gordon-Smith, 2001). Infusion of alemtuzumab results in marked decrement in circulating levels of NK cells, B cells, T lymphocytes, macrophages, and monocytes. Although numbers of NK cells, B cells, and monocytes typically return to normal levels within 3 to 6 months of alemtuzumab use, CD4+ and CD8+ T cells may remain low for years (Ferrajoli et al., 2001). In a trial of alemtuzumab in patients with rheumatoid arthritis, there was profound and persistent peripheral blood lymphopenia in the alemtuzumab-treated patients, affecting predominantly the CD4+ subset. Median CD4+ and CD8+ peripheral blood lymphocyte counts at 73 to 84 months after

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therapy were 185 and 95 cells/μL, respectively (Issacs et al., 2001). More recently, alemtuzumab has come into use for induction immunosuppression and treatment of rejection for solid organ transplantation in some transplantation centers (Gourishankar et al., 2002; Magliocca and Knechtle, 2006; Morris and Russell, 2006; Shapiro et al., 2005a, 2005b). Long-lasting lymphopenia has been confirmed in patients treated with alemtuzumab for induction of immunosuppression in solid organ transplantation administered maintenance mycophenolate mofetil or sirolimus in addition to calcineurin inhibitors (Knechtle et al., 2003, 2004). Depletion of lymphocytes would be expected to result in an increased risk of opportunistic infections. Some series have found that alemtuzumab use was associated with an increase in the frequency of unusual infections (Abad et al., 2003; Magliocca and Knechtle, 2006; Martin et al., 2006; Nath et al., 2005; Silveira et al., 2007). However, others have not confirmed these findings (Barth et al., 2006), and thus, whether or not alemtuzumab use translates into an increased risk of infection in solid organ transplant recipients is unclear. We performed a retrospective cohort study to determine the risk of infection in patients receiving alemtuzumab induction compared with basiliximab/daclizumab or antithymocyte/antilymphocyte globulin at time of transplantation. We focused particularly on fungal infections as the main outcome. 2. Methods 2.1. Sources of data Using data from a prospectively maintained transplant database, supplemented by chart review and pharmacy billing records, we identified all 1738 patients who had undergone renal, liver, and kidney–pancreas transplantation at the University of Wisconsin, Madison, WI, between January 1, 2002, and December 31, 2005. For this retrospective cohort study, the cohort was constructed by including patients who received at least 1 dose of alemtuzumab as the alemtuzumab group. For retransplanted patients, only the first transplant was included. A comparator group was constructed by randomly selecting transplant recipients, using a random numbers table from a list of the transplant recipients who did not receive alemtuzumab but received basiliximab/daclizumab, antithymocyte globulin (ATG), and/or antilymphocyte globulin (OKT3). The study was approved by the institutional review board. Data collection included demographics, information pertaining to transplantation, and infection. Relevant definitions are provided in Table 1. Definitions for fungal infections were adapted from Ascioglu et al. (2002). All patients were given perioperative prophylaxis with cefazolin (renal transplants) and ceftriaxone (liver transplants). Selective digestive decontamination was not used. The alemtuzumab group received 1 (20 mg) or 2 doses

Table 1 Definition of infectious complications Infection

Definition

BK virus CMV viremia

Presence of BK viral load in blood or urine Positive quantitative serum polymerase chain reaction CMV disease Clinical signs and symptoms and histopathologic evidence of CMV (tissue invasive disease) or viral syndrome Severe CMV disease Tissue invasive disease requiring inpatient or intensive care Recurrent CMV Positive serum quantitative polymerase chain disease reaction and tissue invasive disease after a negative serum polymerase chain reaction test and resolution of clinical signs and symptoms of CMV after treatment of prior episode Fungal infection Histopathologic or cytopathologic examination (mold) showing hyphae from needle aspiration or biopsy specimen with evidence of associated tissue damage (either microscopically or unequivocally by imaging), or positive culture result for a sample obtained by sterile procedure from normally sterile and clinically or radiologically abnormal site consistent with infection, excluding urine and mucous membranes Fungal infections Histopathologic or cytopathologic examination (yeast) showing yeast cells (Candida spp. may also show pseudohyphae or true hyphae) from specimens of needle aspiration or biopsy excluding mucous membranes, or positive culture result on sample obtained by sterile procedure from normally sterile and clinically or radiologically abnormal site consistent with infection, excluding urine, sinuses, and mucous membranes, or microscopy (India ink, mucicarmine stain) or antigen positivity for Cryptococcus spp. in cerebrospinal fluid Disseminated fungal Evidence of invasive disease from N2 organ infection systems or bloodstream infection Fungemia Blood culture that yields fungi or yeast Pneumonia Respiratory symptoms and new infiltrate on chest radiograph Bacteremia Positive peripheral blood culture with a pathogenic organism. More than 2 positive blood cultures if organism was a common contaminant such as coagulase-negative Staphylococcus Bacteriuria N1000 CFU/mL Rejection Based on histopathology or attending physician notes if biopsy was contraindicated Leukopenia b3.0 × 103/μL Disseminated varicella Multiple dermatomal involvement with clinical disease and/or microbiologic evidence of varicella Adapted from Ascioglu et al. (2002). Both proven and probable fungal infections were included.

(40 mg) of alemtuzumab intravenously, the first given before reperfusion. The group that did not receive alemtuzumab received basiliximab (2 doses of 20 mg on operative day and day 4), daclizumab (1 mg/kg on day 0 and 1 dose every other week for a total of 5 doses), OKT3, or ATG/thymoglobulin. Thymoglobulin (Sang Stat, Fremont, CA) was administered at a dose of 1.5 mg/kg daily starting on day 0 (day of transplant) and continued until calcineurin inhibitor levels were therapeutic or a maximum

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of 14 doses was given. The operating surgeon made the decision to use a particular antibody for induction therapy on the basis of perceived efficacy, side effect profile, and cost (Knechtle et al., 2004). Thymoglobulin was the generally preferred antibody for retransplants, although not limited to retransplants. A minimum of 4 doses of thymoglobulin was given. Muromonab-CD3 (OKT3; Ortho Biotech, Raritan, NJ) was given at a dose of 5 mg/day for a minimum of 7 doses and maximum of 14 doses. ATG (Upjohn, Kalamazoo, MI) was given at a dose of 15 mg/kg iv for a minimum of 4 daily doses and a maximum of 14 doses. For kidney transplants, maintenance immunosuppression in all groups consisted of either tacrolimus or cyclosporine started when serum creatinine level declined to less than 3.0 mg/dL. Use of cyclosporine versus tacrolimus was at the discretion of the transplant surgeons and physicians who cared for these patients. Mycophenolate was used in all patients starting on day 1 at a dose of 1000 mg bid and reduced for symptoms of diarrhea or low white blood cell count (less than 3.0). Prednisone was tapered to 10 mg by 3 months in patients without rejection. For liver transplants, maintenance immunosuppression consisted of tacrolimus with or without a second immunosuppressant such as mycophenolate at a dose of 2 to 3 g per day in 2 divided doses. The tacrolimus doses were adjusted to maintain levels of 5 to 10 ng/mL. Antifungal prophylaxis included clotrimazole troches or nystatin swish and swallow for 3 months. Fluconazole antifungal prophylaxis is not routinely used at our institution for any of these organ transplant groups. For cytomegalovirus (CMV) prophylaxis in kidney transplant recipients, donor positive/recipient negative (D+/R−) received valganciclovir, donor negative/ recipient negative (D−/R−) received acyclovir 400 mg bid, and donor negative/recipient positive (D−/R+) received acyclovir 800 mg qid if induction agent was basiliximab, but if alemtuzumab or thymoglobulin was used, valganciclovir instead of acyclovir is now used. Preemptive therapy or monitoring for CMV is not used at our institution. Liver transplant recipients received valganciclovir for CMV prophylaxis. In the years before valganciclovir became available, oral ganciclovir was used. The duration of CMV prophylaxis was 90 days after transplantation. Treatment of rejection was initiated with high-dose corticosteroids. For steroid refractory rejection, OKT3 was used. As the study was over a 4-year period, changes in immunosuppression that occurred over time included more basiliximab use compared with ATG and more alemtuzumab use in the latter half of the study period. We also undertook a literature review of series and trials of alemtuzumab in solid organ transplantation using the search terms Campath, alemtuzumab, and liver, kidney, pancreas, transplant in PubMed from 1998 to December 2006. The purpose of the literature review was to compare the results of the available studies with regard to infection. To be included, studies had to provide data on infection

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and Campath use in solid organ transplant patients. In cases of multiple reports on the same type of transplant on patients from a single center, the largest study was chosen. Trials that did not use a control population where alemtuzumab was not used were not included. Because of the clinical heterogeneity in types of outcomes, types of transplant, and dosing regimens of alemtuzumab, a formal metaanalysis to combine the results was not undertaken (The Tricontinental Mycophenolate Mofetil Renal Transplantation Study Group, 1996). 2.2. Statistical analysis We divided the study population into 3 groups based on induction therapy: the first group consisting of patients who received alemtuzumab, the second group consisting of patients who received either basiliximab or daclizumab, and the third group comprising patients who received either ATG or OKT3. The 3 groups were compared with regard to baseline characteristics. Baseline data in the 3 groups was compared using χ2 or Fisher exact test for categoric variables (such as type of transplant, gender, and infections) and 1-way analysis of variance for continuous variables (such as time of observation after transplantation). For infectious and related outcomes, such as incidence of overall infection, fungal infection, viral infection, and bacterial infection, and patient and graft survival and rejection, the alemtuzumab group was compared with each of the other 2 groups, using χ2 or Fisher exact test for categoric variables and Student t test for continuous variables. Main outcomes of interest in univariate analyses were overall infection, fungal infection, disseminated fungal infection, CMV viremia, severity of CMV disease, leucopenia, urinary tract infection, pneumonia, bacteremia, surgical site infection, rejection, and patient and graft survival. Multivariate logistic regression models were used to estimate independent predictors of types of infection found to be significant in univariate analysis and those that are clinically significant. The dependent outcome for multivariate analysis was disseminated fungal infection. A multivariable model was constructed using backward elimination. Criteria for backward elimination of a variable were P value N .05. Analyses were conducted using SAS software (version 8.1, 2000; SAS, Cary, NC) (Lemeshow and Hosmer, 1982).

3. Results 3.1. Patient demographics The patients were followed longitudinally at our institution for a mean of 13 months (SD, 8.3). Patient demographics and clinical features are shown in Table 2. There were 215 patients in the basiliximab/daclizumab group (henceforth called basiliximab group, 9 patients received daclizumab), 726 patients in the alemtuzumab group, and 85 in the ATG or OKT3 group (henceforth called ATG group, 1 patient received OKT3). Because of

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Table 2 Features of the study population

Sex, n (%) Male Age at time of transplant, mean years (SD) Transplant type, n (%) Kidney Kidney–pancreas Liver Liver–kidney Pancreas Heart–kidney Islet cell Immunosuppression, n (%) Prednisone Mycophenolate Cyclosporine Tacrolimus Sirolimus Time of observation after transplantation, mean months (SD) PCP prophylaxis, n (%) Viral prophylaxis Acyclovir (recipient positive and donor negative/recipient negative) Valganciclovir (donor positive/recipient negative, recipient positive if alemtuzumab givena Unknown Fungal prophylaxis CMV serostatus Donor positive/recipient negative Recipient positive Donor negative/recipient negative Unknown

Basiliximab (n = 215)

Alemtuzumab (n = 726)

ATG or ALG (n = 85)

126 (59) 52.05 (15.3)

430 (59) 52.86 (17.6)

49 (58) 50.06 (16.5)

174 (81) 32 (15) 4 (2) 2 (1) 3 (1)

592 (82) 88 (12) 20 (3) 5 (1) 17 (2) 2 (0) 2 (0)

71 (84) 7 (8) 1 (1) 6 (7)

215 (100) 159 (74) 126 (59) 78 (36) 21 (10) 13.06 (8.3) 200 (93) 210 (98) 140 (65) 42 (20)

716 (99) 701 (97) 349 (48) 350 (48) 92 (13) 13.13 (8.49) 699 (96) 699 (96) 490 (67) 203 (28)

85 (100) 71 (84) 48 (56) 39 (46) 7 (8) 13.68 (10.22) 80 (94) 82 (96) 42 (49) 21 (25)

19 (9) 193 (90)

33 (5) 682 (94)

22 (26) 78 (92)

42 (20) 111 (52) 48 (22) 14 (7)

102 (14) 518 (71) 88 (12) 20 (3)

8 (9) 51 (60) 7 (8) 19 (22)

P

.42 .55 .73

.12 b.0001 .01 .03 .31 .82 .11 .79 .16

.21 .09⁎

ALG = antilymphocyte globulin; PCP = P. jiroveci pneumonia. a Oral ganciclovir was used before availability of valganciclovir. ⁎ P value comparing CMV status across the 3 groups.

limitations in data availability, we were not able to obtain an equal number of patients in all 3 groups. The distribution of men and women was similar in all 3 groups. The mean age for the 3 groups was 52 years. Most patients had a kidney transplant (81% in all 3 groups, P = .73). The next most common organ transplant was kidney–pancreas. All patients received prednisone. Ninety-six percent of patients in the alemtuzumab group received mycophenolate compared with 74% in the basiliximab group and 83% in the ATG group (P ≤ .0001). More patients in the basiliximab group received cyclosporine (59%) compared with 48% in the alemtuzumab group and 56% in the ATG group (P = .01). Tacrolimus was used most frequently in the alemtuzumab group (48% compared with 37% for basiliximab group and 46% for the ATG group, P = .03). Sirolimus was used in 13% of patients in the alemtuzumab group compared with 10% in the basiliximab group and 1% in the ATG group (P = .31). The mean time of observation after transplantation was 13 months in all groups (P = .82). There were no significant differences in the groups regarding fungal, viral, or Pneumocystis jiroveci pneumonia prophylaxis (P N .05 for all comparisons).

3.2. Univariate analysis 3.2.1. Alemtuzumab compared with basiliximab Overall, 33% of patients in the alemtuzumab group (240/ 724) developed infection compared with 40% (87/215) in the basiliximab group (odds ratio [OR], 0.72; 95% confidence interval [CI], 0.53–0.99; P = .04) (Table 3). However, CMV viremia was significantly more frequent in the alemtuzumab group (25% versus 7%), with a 4-fold increased risk (OR, 4.27; 95% CI, 2.50–7.31; P b .0001). No significant differences were found in the severity of CMV disease, and recurrent CMV was less frequent in the alemtuzumab group, 58% versus 87% (OR, 0.19; 95% CI, 0.09–0.38; P b .0001). Varicella–zoster localized disease and disseminated zoster were more common in the alemtuzumab group, but the results did not achieve statistical significance (24/724 versus 3/215 for localized disease and 11/24 and 1/3 for disseminated zoster, respectively). BK virus was detected in 14% of alemtuzumab patients (101/713). However, comparison with basiliximab and ATG was not possible because of lack of availability of data for all patients. Among bacterial infections, bacteremia and bacteriuria were significantly

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Table 3 Outcomes after induction with alemtuzumab compared with basiliximab and alemtuzumab compared with ATG in univariate analyses Outcome

No. of patients

Alemtuzumab (n = 724)

Basiliximab/ daclizumab (n = 215)

ATG/ALG (n = 85)

OR (95% CI) comparing alemtuzumab to basiliximab

P

OR (95% CI) comparing alemtuzumab to ATG or ALG

P

Infection, n (%) CMV viremia, n (%) Severe CMV disease Recurrent CMV VZV disease VZV disseminated Wound infection Bacteremia Pneumonia Urinary tract infection Fungal infection Disseminated fungal infection Rejection Patient survival Graft survival Leukopenia

358 215 117 134 30 13 152 101 88 307 96 56 309 988 915 345

240/724 (33) 186/724 (26) 101/186 (54) 108/186 (58) 24/724 (3) 11/24 (46) 102/724 (14) 62/720 (9) 64/724 (9) 202/711 (28) 72/724 (10) 49/72 (68) 211/724 (29) 698/724 (96) 638/725 (88) 272/724 (38)

87/215 (40) 16/214 (7) 9/16 (56) 14/16 (88) 3/215 (1) 1/3 (33) 35/214 (16) 28/214 (13) 19/215 (9) 76/213 (36) 20/214 (9) 4/20 (20) 75/215 (35) 206/215 (96) 196/215 (91) 53/215 (25)

31/85 (36) 13/85 (15) 7/13 (54) 12/13 (92) 3/85 (4) 1/3 (33) 15/85 (18) 11/85 (13) 5/85 (6) 29/85 (34) 4/85 (5) 3/4 (75) 23/85 (27) 84/85 (99) 81/85 (95) 20/85 (24)

0.72 (0.53–0.99) 4.27 (2.50–7.31) 0.90 (0.32–2.52) 0.19 (0.09–0.38) 2.40 (0.71–8.07) 1.69 (0.13–21.26) 0.83 (0.55–1.27) 0.62 (0.38–1.00) 1.27 (0.85–1.89) 0.66 (0.53–0.83) 1.07 (0.63–1.81) 4.76 (1.58–14.28) 0.76 (0.55–1.05) 0.91(0.49–1.68) 1.40 (0.83–2.36) 1.89 (1.50–2.37)

.04 b.0001 .84 b.0001 .14 .68 .40 .05 .23 .0004 .78 .003 .10 .99 .19 b.0001

0.86 (0.54–1.38) 1.92 (1.04–3.57) 1.00 (0.32–3.10) 0.11 (0.04–0.88) 1.06 (0.31–3.62) 1.18 (0.06–21.17) 0.76 (0.42–1.38) 0.63 (0.32–1.26) 1.56 (0.87–4.00) 0.76 (0.47–1.25) 0.44 (0.15–1.25) 0.65 (0.06–6.66) 1.09 (0.65–1.81) 0.39 (0.05–3.00) 0.36 (0.32–1.01) 1.96 (1.16–3.31)

.53 .03 .99 .01 .91 .90 .37 .18 .35 .27 .11 .71 .74 .39 .04 .01

ALG = antilymphocyte globulin; VZV = varicella–zoster virus. No outcome data were recorded on 2 patients in the alemtuzumab group, and these 2 were excluded from the analysis.

more frequent in the basiliximab group than the alemtuzumab group. Although, overall, the frequency of fungal infection was not appreciably different in the 2 groups (10% in the alemtuzumab group versus 9% in the basiliximab group), disseminated fungal infection occurred in 68% of the patients with fungal infection in the alemtuzumab group and in 30% of the patients with fungal infection in the basiliximab group (OR, 4.76; 95% CI, 1.58–14.28; P = .003). Almost all the fungal infections in either group were caused by Candida spp. Fungal infection in the entire study population was associated with high mortality (OR, 9.10; 95% CI, 4.33– 19.11; P ≤ .0001). As anticipated, leucopenia (white blood cell count, b3.0) was more common in the alemtuzumab group (37% versus 25%, P b .0001). There were no significant differences regarding the type and time of transplant, rejection, or patient and graft survival. 3.2.2. Alemtuzumab compared with ATG Overall, the frequency of infection was similar in the alemtuzumab and ATG groups (33% versus 36%, respectively; P = .53) (Table 3). However, CMV viremia was significantly more frequent in the alemtuzumab group (25% versus 15%), with an almost 2-fold increased risk (OR, 1.92; 95% CI, 1.04–3.57; P = .03). No significant differences were found in the severity of CMV disease, but recurrent CMV was less frequent in the alemtuzumab group, 58% versus 92% (OR, 0.11; 95% CI, 0.04–0.88; P = .01). Disseminated varicella was more common in the alemtuzumab group, but the results did not achieve statistical significance. Data on BK virus were not available for the majority of ATG patients and, thus, could not be examined. Among bacterial and fungal infections, there were no significant differences in the 2 groups. Patients with fungal

infection had a 6-fold higher risk of death in the alemtuzumab group than patients without fungal infection (OR, 6.20; 95% CI, 2.47–15.53). Leucopenia was more common in the alemtuzumab group (37% versus 23%, P = .01). There were no significant differences regarding the type and time of transplant, rejection, or patient survival; graft survival was slightly better in the ATG group (95% versus 88%, P = .04). 3.2.3. Microbiology The types of organisms found in patients developing fungal infection are shown in Table 4. The majority of fungal infections, including almost all disseminated infections, were caused by Candida spp. Interestingly, only patients in the

Table 4 Microbiology of fungal infections in the study population Infection

Alemtuzumab group (n = 72)a

No-alemtuzumab group (n = 24)

Aspergillus fumigatus Aspergillus terreus Aspergillus niger Aspergillus spp. Candida albicans Candida glabrata Candida spp. Candida guilliermondii Candida krusei Blastomycosis Histoplasmosis Cryptococcus Total

4 (5.3%) 0 0 8 (10.6) 5 (6.6%) 8 (10.6%) 45 (60%) 0 0 3 (4.0%) 2 (2.6%) 0 75

6 (11.3%) 2 (3.7%) 2 (3.7%) 2 (3.7%) 7 (13.2%) 5 (9.4%) 22 (41.5%) 2 (3.7%) 3 (5.6%) 0 0 2 (3.7%) 53

a Total adds up to more than the number of patients with fungal infection (128) because some patients had more than 1 fungal infection.

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Table 5 Statistically significant independent predictors of disseminated fungal infection in study population receiving either alemtuzumab or basiliximab induction at time of transplantation Variable

OR (95% CI)

P

Alemtuzumab CMV viremia

10.02 (2.69–37.31) 3.64 (1.13–11.67)

.0006 .02

alemtuzumab experienced clinical infection with endemic fungi (blastomycosis and histoplasmosis), although the numbers were small. 3.2.4. Multivariate analysis To determine whether alemtuzumab was an independent predictor of disseminated fungal infection, multivariate models were constructed using all patients in the basiliximab and alemtuzumab groups using variables that were clinically and/or statistically significant as potential factors for the main outcome of disseminated fungal infection. Alemtuzumab use increased the risk of disseminated fungal infection 10-fold and that of CMV viremia 3 and a half-fold, after adjusting for maintenance immunosuppression, gender, and leucopenia (Table 5). Multivariate analysis compared alemtuzumab with ATG as a risk factor for disseminated fungal infection was not undertaken, because in univariate analysis, there was no significant difference in disseminated fungal infection seen when comparing ATG to alemtuzumab. 3.2.5. Results of literature review We identified 11 recent large studies in our literature review, which assessed the risk of infection after alemtuzumab in various types of solid organ transplantation and included a comparator group (Table 6). One was in children and 10 included only adults. Six studies focused on renal transplants, 3 included solitary kidney or kidney–pancreas transplants, and 1 included liver transplant. The only study in children included multivisceral transplantation. In all these studies, alemtuzumab was used for induction. In one, it was also used as maintenance immunosuppressive therapy. All but 1 (Vathsala et al., 2005) were nonrandomized interventional studies with historic controls. The comparator agents were ATG in 4 studies; basiliximab or daclizumab in 3; and combinations of cyclosporine, mycophenolate, azathioprine, tacrolimus, and corticosteroids, depending upon the type of transplant. In all of these studies, alemtuzumab was not associated with a statistically significant higher risk of infection than the comparator agent was. 4. Discussion Our study shows that although the overall risk of infection was not increased with the use of alemtuzumab and was lower than that with basiliximab, the infections that did occur were more severe and were more likely to be disseminated. Our interest was particularly in fungal infections, and we

found that disseminated fungal infection and CMV viremia occurred more commonly in the alemtuzumab-treated group. However, patients who were given alemtuzumab were also more likely to receive mycophenolate, which may impact the risk of opportunistic infection. We adjusted for mycophenolate and other immunosuppression in our multivariate analysis and found that alemtuzumab was an independent risk factor for disseminated fungal infection. Candida spp. rather than molds accounted for the vast majority of fungal and disseminated fungal infections. In our center, antifungal prophylaxis is not routinely used. However, in many transplant centers in the United States and worldwide, azoles, including extended-spectrum azoles, are used routinely as antifungal prophylaxis (Singh et al., 2008). Although effective in reducing fungal infections, they may pose an increased risk of azole-resistant infections (Husain et al., 2003). Thus, ongoing surveillance is important in determining the impact of various induction strategies and prophylaxis regimens on the incidence and epidemiology of invasive fungal infections. We also found that infections unusual in the early posttransplant period, such as histoplasmosis and blastomycosis, occurred in the alemtuzumab group. There were no major differences in patient and graft survival noted in the 3 groups. We found that infections occurring more commonly in the basiliximab group compared with alemtuzumab included bacteremias, urinary tract infection, and recurrent CMV disease. The incidence of CMV disease may have been higher in patients with basiliximab because the prophylaxis regimen used in our study was acyclovir for all R+ patients, whereas a more effective regimen using valganciclovir was used in patients that received alemtuzumab. In other studies with a study design similar to ours, overall infection was not noted to be different in patients receiving basiliximab or alemtuzumab (Knechtle et al., 2004; Kaufman et al., 2005). However, these studies were limited in the amount of detail provided regarding the infections that occurred. Differences in patient populations and regimens for prophylaxis and treatment across institutions may account for these disparate findings. Although the risk of infection has been well described with the use of alemtuzumab in hematologic malignancies (Dearden, 2006; Enblad et al., 2004; Ravandi and O'Brien, 2005), relatively little is known in solid organ transplantation where the number of doses of alemtuzumab for induction is usually 1 (30 mg iv) or 2 compared with the multiple dosing over weeks used for treatment of hematologic malignancy, such as chronic lymphocytic leukemia. Uncontrolled series have reported conflicting data on the risk of infection with alemtuzumab in solid organ transplantation (Barth et al., 2006; Silveira et al., 2006; Thai et al., 2006). However, the studies are limited by small cohort size, lack of a comparator group, varying definitions of infections, and varying types and regimens of immunosuppression. Either most studies assessed only

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Table 6 Infectious complications associated with alemtuzumab in solid organ transplantation in large series and trials Study

Study design

Patient population receiving alemtuzumab (n)

Induction therapy or treatment of rejection

Comparator

Reported infections, alemtuzumab group versus comparator

Mean duration of follow-up

Tzakis et al. (2004) Malek et al. (2006)

Before–after study using historic controls Before–after study using historic controls

Adult liver transplants (40) Adult renal transplant patients (49)

Induction

10% versus 2% (P = NS) 16% versus 32% (P = NS)

12 months

Vathsala et al. (2005)

Randomized controlled trial

Adult renal transplant patients (20)

Induction

Knechtle et al. (2004)

Before–after study using historic controls

Adult renal transplant recipients (126)

Induction

Tacrolimus and corticosteroids Cyclosporine, mycophenolate, prednisone, and basiliximab in minority Cyclosporine, azathioprine, corticosteroids Basiliximab or thymoglobulin

Watson et al. (2005)

Before–after study using historic controls

Adult renal transplant recipients (33)

Induction

Tan et al. (2006)

Before–after study using historic controls

Adult renal transplant recipients (205)

Induction

Gruessner et al. (2005)

Before–after study using historic controls

Induction and maintenance

Sundberg et al. (2005)

Before–after study using historic controls

Kato et al. (2006)

Before–after study using historic controls

Kaufman et al. (2005) Kaufman et al. (2006)

Before–after study using historic controls Before–after study using historic controls

Adult pancreas kidney and solitary kidney transplant (75) Adult pancreas–kidney and solitary kidney transplant (16) Children with intestine, liver and intestine, multivisceral (18) Adult kidney transplant (123) Adult kidney–pancreas transplant (50)

Induction

Cyclosporine, azathioprine, prednisone—no induction agent Cyclosporine, azathioprine, prednisone—no induction agent Thymoglobulin

13 months

40% versus 30% (P = NS)

6 months

Similar incidence of infection in alemtuzumab group compared with basiliximab or thymoglobulin (P = NS) 30% versus 19% (P = NS)

36 months

60 months

No CMV infection in either group

41 months

29% versus 31% (P = NS)

6 months

Induction

ATG

12% versus 19% (P = NS)

9 months

Induction

Daclizumab

NR by group

36 months

Induction

Basiliximab

36 months

Induction

ATG

No difference in infection rates 6% 3-year CMV infection versus 19% 3-year CMV infection (P b 0.05)

36 months

NS = not significant; NR = not reported.

overall infection, without delving into the epidemiology of the ones that did occur, or they assessed only specific infections such as CMV. Moreover, the severity of infections was not reported in the studies. Similar to these studies, we found no overall increased risk of infection in our analysis; however, in-depth analysis of the infections occurring in our study showed that infections caused by Candida were more likely to be severe and disseminated. Fungal infections were associated with excess mortality in patients receiving alemtuzumab compared with ATG and in patients receiving basiliximab compared with alemtuzumab. Other recent studies of alemtuzumab use in solid organ transplantation have also reported the development of unusual and unusually severe infections; Peleg et al. (2007) assessed the risk of infection in 547 solid organ

transplant recipients who received alemtuzumab as either induction or for treatment of rejection. Ten percent of patients developed a total of 62 infections, with those receiving alemtuzumab induction less likely to develop infection than those receiving it as treatment of rejection, perhaps because of the multiple doses often used in treatment of rejection. Alemtuzumab used as treatment of rejection rather than induction was associated with a 3-fold greater risk of developing an opportunistic infection. In this study, the microbial etiology of infections was diverse, including nocardia, mycobacteria, and invasive molds. Silveira et al. reported data on cryptococcal disease in 834 liver and 727 kidney transplant recipients who received either alemtuzumab or ATG (780) or neither ATG and alemtuzumab (781). The cumulative incidence of cryptococcosis was 0.26% among those who did not receive ATG or alemtuzumab,

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0.3% (2/646) among those who received only 1 dose, and 2.24% (3/134) among those who received 2 doses (P = .03). However, the authors did not report analyses separately for ATG and alemtuzumab. These studies are not included in Table 6 because of lack of a nonalemtuzumab group. The primary goal of the current study was to compare alemtuzumab induction with other induction therapies to determine whether or not it posed a differential risk of infection. The fact that disseminated candidal infection and CMV was more common in the alemtuzumab group and that leucopenia was more frequent raises questions about optimal induction therapy in kidney transplant recipients. It should be acknowledged that preemptive monitoring and treatment are not used in our institution, and the prophylaxis protocol used acyclovir except in the very high-risk patients. Moreover, we did not collect data on mycophenolate dosage. This may impact the risk of CMV because 3 g of mycophenolate has been reported to increase the risk of CMV (1996). The current study has a number of limitations, including the retrospective design, which may have limited the availability of data. The inability to have a larger sample size for the alemtuzumab comparison groups because of lack of data availability limits the power of the study to detect small differences. We were not able to quantify the extent of lymphocyte depletion, although we did find that absolute leucopenia was much more frequent in the alemtuzumab group. The vast majority of our population was kidney transplant recipients, and very few had other types of transplants. Our average duration of follow-up was 13 months; longer follow-up would be desirable to determine how the risk of infection with induction immunosuppression using alemtuzumab changes over a longer period. Furthermore, ours is an observational study and is limited by the use of historic rather than concurrent controls, which may introduce confounding as a result of era-dependent differences in the groups under study. At the time the study was undertaken, the use of alemtuzumab as induction therapy was evolving, and there were no set criteria for use of alemtuzumab rather than basiliximab or daclizumab. The choice of 1 or 2 doses was also arbitrary and was at the discretion of the treating surgical team. As such, our data represent an exploratory analysis of the infection risk with various types of induction therapy. Future randomized trials comparing alemtuzumab to basiliximab and ATG are necessary to identify optimal induction therapy including the risks of infection with each. Our findings have important implications for the prevention of infection after transplantation, particularly fungal infection. The benefit of reducing rejection with a lymphocyte-depleting agent such as alemtuzumab must be balanced against the risk of severe infection. Future studies should undertake a detailed analysis of infections after the use of alemtuzumab and identify surrogate markers, such as the CD4 count to be followed serially to determine the risk and duration of risk for infection.

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