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1 (2010) 45, & 2010 Macmillan Publishers Limited All rights reserved /10 $ REVIEW CMV central nervous system disease in stem-cell transplant recipients: an increasing complication of drug-resistant CMV infection and protracted immunodeficiency SM Reddy 1, DJ Winston 2, MC Territo 2 and GJ Schiller 2 1 David Geffen School of Medicine, University of California, Los Angeles, CA, USA and 2 Division of Hematology-Oncology, Department of Medicine, UCLA Medical Center, Los Angeles, CA, USA We report on two patients with no active GVHD and on moderate doses of immunosuppressive drugs who unexpectedly developed fatal CMV meningoencephalitis after umbilical cord blood transplantation. A review of these two cases along with nine other cases of CMV central nervous system (CNS) disease after allogeneic SCT that were mostly reported within the last 8 years suggests that this severe complication of CMV infection may be increasing. CMV CNS disease after allogeneic SCT is a late-onset disease (median time of onset, 210 days) and is usually manifested as encephalitis in the absence of other sites of CMV disease. The development of CMV CNS disease is associated with risk factors (T-cell depletion, anti-thymocyte globulin, umbilical cord blood transplantation) that cause severe and protracted T-cell immunodeficiency (8 of 11 cases), a history of recurrent CMV treated with multiple courses of preemptive ganciclovir or therapy (11 of 11 cases), and ganciclovir-resistant CMV infection (11 of 11 cases). Despite therapy with a combination of antiviral drugs (ganciclovir, and cidofovir), mortality is high (10 of 11 cases). Given this high mortality, extended with current or novel antiviral drugs and strategies to enhance CMV immunity need to be considered in high-risk patients. (2010) 45, ; doi: /bmt ; published online 1 March 2010 Keywords: CMV; central nervous system; cord blood; SCT Introduction Before the availability of an effective preventive therapy, CMV disease was a common cause of morbidity and Correspondence: Dr DJ Winston, Division of Hematology-Oncology, Department of Medicine, UCLA Medical Center, Room , CHS, Le Conte Avenue, Los Angeles, CA 90095, USA. dwinston@mednet.ucla.edu Received 2 September 2009; revised and accepted 14 January 2010; published online 1 March 2010 mortality after allogeneic hematopoietic SCT. Currently, the use of ganciclovir either as or as preemptive therapy has eliminated most cases of CMV disease during the initial several months after transplantation. 1,2 On the other hand, late-onset CMV disease after the completion of or preemptive therapy is being recognized more frequently in high-risk patients. 1 The common manifestations of CMV disease after allogeneic SCT have been interstitial pneumonia and gastrointestinal disease. In contrast, central nervous system (CNS) disease due to CMV infection has been very rarely reported. 3 In this report, we describe two stable patients without active GVHD and on relatively low doses of immunosuppressive drugs who unexpectedly developed life-threatening CMV meningoencephalitis after umbilical cord blood transplantation in A review of these two patients along with previously reported cases of CMV CNS disease after SCT suggests that this complication of CMV infection may be increasing as a consequence of antiviral resistance in a more susceptible patient population with protracted immunodeficiency. Case reports Patient 1 A 55-year-old man with myelodysplastic syndrome of intermediate risk type 2 received an unrelated 5/6 histocompatible- single umbilical cord blood transplant ( CD34 cells/kg of body weight) in January The patient was CMV-seropositive, whereas the donor was CMV-seronegative (R þ /D ), as determined by the maternal CMV IgG serostatus. A reducedintensity preparative conditioning regimen of BU, fludarabine and anti-thymocyte globulin (total dose of 3 mg/kg over days 4 and 3 before transplantation) was administered. Cyclosporine, mycophenolate and weekly (500 mg/ kg) were given for prevention of GVHD. The was continued until day 100 after transplantation. The patient s initial post-transplantation course was relatively uncomplicated. After engraftment, on day 48 after transplantation, the patient developed asymptomatic CMV. CMV DNA level in the blood was

2 980 CMV CNS disease in stem-cell transplant recipients copies/ml, as determined by a quantitative real-time PCR method using reagents from Roche Diagnostics (Indianapolis, IN, USA). Preemptive therapy with i.v. ganciclovir (5 mg/kg every 12 h) was started. Blood CMV DNA became undetectable on day 55, and i.v. ganciclovir was changed to a maintenance dose of 6 mg/kg per day, from Monday through Friday. However, on day 92, surveillance testing revealed recurrent CMV ( copies/ml) on maintenance ganciclovir; therefore, the dose of i.v. ganciclovir was increased to 5 mg/kg every 12 h. CMV then decreased to 3020 copies/ml on day 115, and i.v. ganciclovir was changed back to a maintenance dose of 6 mg/kg per day, from Monday through Friday. On day 146, when surveillance testing showed the blood CMV DNA level had increased to copies/ml on maintenance ganciclovir, ganciclovir resistance was suspected clinically. Thus, on day 152, treatment was changed from ganciclovir to i.v. (90 mg/kg every 12 h). The patient also developed grade 2 GVHD of the gastrointestinal tract on day 159, which resolved on prednisone (15 mg per day). On day 184, blood CMV DNA became undetectable on the which was subsequently discontinued on day 191 owing to nephrotoxicity (increase of serum creatinine from 0.9 to 1.6 mg per 100 ml). On day 215, the patient was admitted to the UCLA Medical Center for progressive lethargy and weight loss. He had no active GVHD on cyclosporine, mycophenolate and prednisone (15 mg/day). Serum trough cyclosporine level was 26 ng/ml. His absolute CD4 count was 49/mm 3. Blood CMV DNA was copies/ml. Brain magnetic resonance imaging showed multiple small infarcts. Lumbar puncture revealed a CSF RBC of 1.0/mm 3, WBC of 12/mm 3 (100% lymphocytes), glucose level of 64 mg per 100 ml and protein level of 55 mg per 100 ml. CMV DNA was detected in the CSF by a qualitative PCR method using reagents from Invitrogen Corporation (Carlsbad, CA, USA). All CSF cultures, serologies and PCR assays for other organisms were negative. Despite treatment with i.v. ganciclovir alone, followed by alone and then cidofovir plus for CMV meningoencephalitis, the patient had progressive deterioration of his neurological function over the next 6 weeks. On day 252, blood CMV DNA level was copies/ml, and repeat CSF PCR was still positive for CMV DNA. The patient expired on day 259. An autopsy was not done. Patient 2 A 41-year-old man diagnosed with relapsed bi-phenotypic acute leukemia received an unrelated 5/6 histocompatible single umbilical cord blood transplant ( CD34 cells/kg of body weight) in May Both the recipient and donor were CMV-seropositive (R þ /D þ ). The donor CMV IgG serostatus was determined by using maternal blood. The preparative conditioning regimen was a combination of CY, fludarabine, anti-thymocyte globulin (total dose of 2.75 mg/kg of anti-thymocyte globulin over days 3, 2, and 1 before transplantation) and total-body irradiation. Cyclosporine, mycophenolate and weekly (500 mg/kg) were given for prevention of GVHD. The was continued until day 100 after transplantation. The patient also received prophylactic i.v. ganciclovir (6 mg/kg per day, Monday Friday) from the time of engraftment (day 27 after transplantation) until day 100 after transplantation. The patient s initial posttransplantation course was relatively uncomplicated. He developed grade 2 GVHD of the skin on day 64 after transplantation, which resolved on administration of prednisone (40 mg/day). On day 159, the patient developed asymptomatic CMV. The quantitative PCR test showed copies/ ml of CMV DNA in the blood. Preemptive therapy with oral valganciclovir (900 mg every 12 h) was started. On day 177, blood CMV DNA had increased to copies/ml, and oral valganciclovir was changed to i.v. ganciclovir (5 mg/kg every 12 h). On day 186, blood CMV DNA level increased to copies/ml. Ganciclovir resistance was suspected clinically, and ganciclovir was changed to i.v. (90 mg/kg every 12 h). Blood CMV DNA became undetectable on day 201. Despite nephrotoxicity (increase of serum creatinine from 1.7 to 2.7 mg per 100 ml), was continued at a reduced induction dose of 60 mg/kg every 12 h, which was later changed to a maintenance dose of 60 mg/kg once daily on day 229. On day 239, the patient was admitted to the UCLA Medical Center for disorientation and poor memory. He was still receiving maintenance. There was no active GVHD on cyclosporine, mycophenolate and a low dose of prednisone (3 mg/day). Serum trough cyclosporine level was o30 ng/ml. His absolute CD4 count was 43/mm 3. Blood CMV DNA was not detectable. Brain magnetic resonance imaging showed several foci of restricted diffusion along the ventricles and the ependyma, consistent with encephalitis. Lumbar puncture showed a CSF RBC of 14/mm 3, WBC of 42/mm 3 (98% lymphocytes), glucose level of 72 mg per 100 ml and protein level of 51 mg per 100 ml. The qualitative PCR test was positive for CMV DNA in the CSF. All CSF cultures, serologies and PCR tests for other organisms were negative. CSF cytological analysis was negative for malignant cells. Treatment with higher doses of (60 mg/kg every 12 h) in combination with i.v. cidofovir (5 mg/kg once weekly) was given for CMV meningoencephalitis. I.v. Ig (500 mg/kg every 48 h) was added on day 245. Despite this combination therapy over the next weeks, the patient experienced progressive deterioration of his neurological function and renal failure. He expired on day 277. An autopsy was not done. Review of reported cases We reviewed the literature for cases of CMV CNS disease previously reported in stem-cell transplant recipients, by using the PubMed database. Keywords used for this search were cytomegalovirus encephalitis, cytomegalovirus and encephalitis and stem, cytomegalovirus and encephalitis and transplant, cytomegalovirus and meningitis, and cytomegalovirus and CNS. All languages were searched. The period reviewed was The criteria used for the diagnosis of CMV CNS disease for our two cases and for the selection of the published cases have been published previously. 4 CMV CNS disease was defined by the presence

3 CMV CNS disease in stem-cell transplant recipients of CNS symptoms associated with the detection of CMV in CSF samples, by PCR or culture, or in brain tissue specimens, by culture, histopathological testing, immunohistochemical analysis or in situ hybridization. This review yielded nine cases of CMV CNS disease in SCT recipients, which meet the criteria for CMV disease of the CNS Data from these nine cases along with the two UCLA cases (a total of 11 patients) are summarized in Table 1. Except for one pediatric patient, all patients were adults (median age, 46 years). Except for one patient with metachromatic leukodystrophy, all patients had undergone allogeneic SCT for a hematologic malignancy. The source of stem cells was peripheral blood in seven patients, BM in one patient, cord blood in two patients and BM plus peripheral blood in one patient. The donor was related for four cases and unrelated for seven cases. Nine patients were CMV-seropositive before transplantation, whereas two patients were CMV-seronegative before transplantation. Anti-thymocyte globulin was used in 9 of the 11 patients either as part of the pre-transplantation conditioning regimen or as part of the post-transplantation GVHD prophylactic regimen. Four patients also received a T-celldepleted stem-cell graft, and thus 10 of the 11 patients received either a T-cell-depleted graft and/or were treated with anti-thymocyte globulin. At the time of onset of the CMV CNS disease, 7 of the 11 patients were being treated for GVHD, and the CD4 count was low (o70 cells/mm 3 ) in all 8 patients with reported results. Of note, only one patient (case 11) had received previous CMV with ganciclovir after engraftment. All these cases of CMV CNS disease were cases of lateonset CMV disease, occurring 166 or more days after transplantation (median time of onset 210 days, range days). Encephalitis was the predominant clinical presentation. Computed tomography scan or magnetic resonance imaging of the brain commonly showed multiple foci of restricted diffusion or infarction. Three patients had concomitant retinitis. Only two patients (case 5 with pneumonia, case 7 with colitis) had concomitant CMV disease outside the CNS. On the other hand, all 11 of the patients had a previous history of recurrent CMV treated with multiple courses of preemptive therapy using ganciclovir (11 patients), (11 patients) or cidofovir (1 patient). The diagnosis of CMV CNS infection was established by demonstrating CMV DNA in the CSF or brain tissue by current PCR methods. For the three patients with evidence of CMV infection in brain tissue, inclusion bodies were seen in three cases and immunohistochemical stains were positive for CMV in two cases. A ganciclovir-resistant mutant strain of CMV was found in eight patients, while three other patients (cases 6, 10 and 11) had increasing levels of CMV on ganciclovir consistent with ganciclovir resistance. Thus, all 11 of the patients had evidence of ganciclovir-resistant CMV infection. Despite treatment with ganciclovir (9 patients), (11 patients), cidofovir (5 patients) and Ig (5 patients), only 1 of these 11 SCT patients with CMV CNS disease survived. The surviving patient (case 9) had encephalitis plus retinitis treated successfully with a combination of and cidofovir. This patient also had her immunosuppressive therapy tapered, and demonstrated both humoral and cellular immune responses to CMV during her recovery. Discussion CMV disease of the CNS has occurred most commonly in patients with advanced AIDS, 3,13 but is rarely reported in SCT or solid-organ transplant patients. The clinical features of CMV CNS disease in AIDS patients include encephalitis, ventriculoencephalitis and polyradiculomyelitis. CMV retinitis has been highly associated with the occurrence of CMV encephalitis in patients with AIDS, who often develop CNS disease with a ganciclovir-resistant CMV strain while receiving ganciclovir therapy for a previous retinitis. 13 In this review, we found that CMV CNS disease is a relatively late complication after allogeneic hematopoietic SCT. All cases occurred more than 4 months after transplantation (median time of onset, 210 days). Encephalitis alone is the predominant clinical manifestation. In contrast to AIDS patients, only one patient had a history of CMV retinitis, and only two patients had concomitant CMV disease outside the CNS (Table 1). Computed tomography and magnetic resonance imaging of the brain usually show changes suggestive of encephalitis or ventriculoencephalitis, but detection of CMV DNA by PCR in the CSF or brain tissue is required for definitive diagnosis. Of note, two patients, case 2 reported by Julin et al. 6 and case 11 from UCLA (Table 1), developed CMV CNS disease despite clearing of their CMV. This compartmentalization of CMV infection emphasizes the importance of not excluding CMV disease in SCT patients who have no detectable ongoing CMV. Several factors appear to contribute to the development of CMV CNS disease after allogeneic SCT. First, CMVspecific CTLs are of critical importance in protecting against severe CMV disease after allogeneic SCT. 14,15 Although CMV-specific CTL function was not measured in these patients with CMV CNS disease, most patients had received either T-cell-depleted stem-cell grafts or had been treated with a T-cell-depleting agent (anti-thymocyte globulin). A low CD4 count was reported in eight patients. In addition, the two UCLA patients developed CMV meningoencephalitis after umbilical cord blood transplantation, which is associated with a slower recovery of T-cell function compared with PBSC or BM transplantation. 16 Second, all of these patients with CMV CNS disease had a history of previous recurrent CMV treated with multiple courses of preemptive ganciclovir or therapy. All cases had either genotypic or clinical evidence of ganciclovir-resistant CMV infection. Thus, prolonged and difficult-to-control CMV caused by ganciclovirresistant viral strains in the setting of severe T-cell deficiency likely placed these patients at a greater risk for CMV disease. Third, the relatively low penetration of ganciclovir and into the CNS may have been a factor in the development of CNS disease, in contrast to pneumonia or other organ involvement, as the major consequence of this uncontrollable CMV infection. 17,18 Of interest, active GVHD, a common risk factor for late-onset 981

4 982 CMV CNS disease in stem-cell transplant recipients Table 1 Characteristics of CMV central nervous system disease in SCT recipients Case no. Reference Age (yrs) Sex Disease Type of transplant CMV serostatus Conditioning GVHD Antiviral T-cell depletion GVHD a Treatment of GVHD CD4 count a cells/ mm 3 CMV infection before CMV CNS disease 1 Seo 6 F ALL BMT+PBSCT, et al. 5 related, R+/D+ TBI, Cy, thiotepa ATG, steroids No Yes Yes Steroids, daclizumub o10 Recurrent 2 Julin 20 M MLD BMT, et al. 6 unrelated R+/D NA CsA, steroids Acyclovir No Yes MMF NA Recurrent 3 Hamprecht 48 F AML PBSCT, et al. 7 unrelated, R /D+ NA ATG, steroids No No No No Low Recurrent 4 Wolf 30 F AML PBSCT, et al. 8 related, haploidentical 5 Wolf 54 M AML PBSCT, et al. 8 related, haploidentical R+/D+ ATG, TBI, FDB, thiotepa R /D FDB, thiotepa, melphalan NA Acyclovir Yes Yes Steroids 30 Recurrent NA Acyclovir Yes Yes Therapy not specified NA Recurrent 6 Zeiser 55 M AML PBSCT, et al. 9 related, haploidentical R+/D+ ATG, FDB, thiotepa, melphalan NA No Yes No No 70 Recurrent 7 Battiwalla 46 M NHL PBSCT, et al. 10 unrelated, R+/D ATG, FDB, Bu MTX, tacrolimus No No Yes Steroids NA Recurrent 8 Miller 64 M AML PBSCT, et al. 11 unrelated R+. Donor NA ATG, pentostatin CsA to tacrolimus No No Yes Steroids 0 Recurrent 9 Hubacek 18 F AML PBSCT, et al. 12 unrelated, mis R+/D Cy, Bu, melphalan ATG, MTX, CsA No No Yes Steroids 0 Recurrent Preemptive therapy IVIG oral GCV i.v. Ig, VGCV CMV CNS disease, onset and diagnosis Ventriculoencephalitis with retinitis, D220,, pp65+, culture+ Encephalitis, D166, CSF and brain tissue CMV PCR+ retinitis, D170, Encephalitis, D285, pneumonia, D201, Polyradiculopathy, D137, CSF CMV PCR+, encephalitis, D193, CSF CMV PCR+, histological and IHC stains of brain tissue CMV+ colitis, D210, CSF CMV PCR+ Ventriculoencephalitis, D180,, IHC stains of brain tissue CMV+ retinitis, D220, Drug resistance mutations detected UL54 in CSF in CSF and brain tissue in blood, not in CNS in blood and CNS in lung, not in CNS None detected UL54,, site not reported UL54 in blood, not in CSF in blood and CSF Treatment Outcome Foscarnet, leflunamide Survived

5 CMV CNS disease in stem-cell transplant recipients Table 1 Continued Treatment Outcome Drug resistance mutations detected CMV CNS disease, onset and diagnosis Preemptive therapy CMV infection before CMV CNS disease CD4 count a cells/ mm 3 GVHD a Treatment of GVHD T-cell depletion Antiviral Conditioning GVHD CMV serostatus Sex Disease Type of transplant Reference Age (yrs) Case no. Not tested CDV Meningoencephalitis, D215, No No No No 49 Recurrent CsA, R+/D ATG, FDB, Bu 10 This study 55 M MDS CBT, Not tested Foscarnet, Meningoencephalitis, D239, VGCV Ganciclovir No No No 43 Recurrent CsA R+/D+ ATG, TBI, Cy, FDB i.v.ig 11 This study 41 M AML CBT, a At time of CMV CNS disease presentation. infections after allogeneic SCT, was present in only 7 of 11 patients. Despite therapy with multiple antiviral drugs and, the mortality of SCT recipients with CMV CNS disease is very high (10 of 11 patients). This high mortality is likely related to the same factors contributing to the development of the disease impaired T-cell immunity, high rate of viral replication, relatively low penetration of antiviral drugs into the CNS and antiviral resistance. Only one patient survived (Table 1, case 9). The surviving patient was taken off immunosuppressive drugs, treated with a combination of plus cidofovir, and also showed humoral and cellular immune responses to her infection. Although a combination of ganciclovir and has been advocated for treatment of ganciclovir-resistant CMV disease in solid-organ transplant patients, 19 all the SCT patients in this review who were treated with this combination expired. All of the cases of CMV CNS disease in SCTs, including the two UCLA patients, were reported only within the last 8 years. Indeed, at UCLA, during the 34-year history of the SCT program ( ), we had not documented any cases of CMV CNS disease until During this 34-year period, 1941 patients underwent allogeneic SCT. There were 1379 adult patients (X18 years old) and 562 pediatric patients (o18 years old). In all, 1462 transplants were related and 479 transplants were unrelated. The source of stem cells was BM in 1492 cases, peripheral blood in 354 cases and cord blood in 95 cases. Most of the unrelated cord-blood transplants at UCLA were done within the last 5 years. Thus, this severe complication of CMV infection may become more common as a greater number of allogeneic SCTs associated with pronounced T-cell deficiency and delayed immune reconstitution are being performed. The deaths of the two UCLA cord blood transplant recipients without GVHD or other common risk factors for infection, as well as the overall high mortality of CMV CNS in allogeneic SCT patients, emphasize the need for effective preventive strategies. Only 1 of the 11 patients in this review had received anti-cmv from the time of engraftment until day 100 after transplantation, and none received CMV after day 100. In contrast, all 11 patients had received repeated courses of preemptive therapy associated with the emergence of ganciclovirresistant CMV strains. Thus, long-term preventive strategies with valganciclovir or a novel anti-cmv drug (maribavir or lipid conjugate of cidofovir) active against ganciclovirresistant CMV strains might be required in high-risk patients Reconstitution of CMV immunity using adoptively transferred CMV-specific T-cells or a CMV vaccine are other prophylactic strategies worthy of consideration Conflict of interest The authors declare no conflict of interest. Acknowledgements The work was carried out at UCLA Medical Center, Los Angeles, CA, USA. 983

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