Pathogenesis and Management of Polyomavirus Infection in ...

INVITED ARTICLE

IMMUNOCOMPROMISED HOSTS David R. Snydman, Section Editor

Pathogenesis and Management of Polyomavirus Infection in Transplant Recipients Eun Jeong Kwak,1,4 Regis A. Vilchez,5,6 Parmjeet Randhawa,3,4 Ron Shapiro,2,4 Janet S. Butel,6 and Shimon Kusne1,2,4 Departments of 1Medicine, 2Surgery, and 3Pathology, University of Pittsburgh Medical Center, and 4Thomas E. Starzl Transplantation Institute, Pittsburgh, Pennsylvania; and Departments of 5Medicine and 6Molecular Virology and Microbiology, Baylor College of Medicine, Houston, Texas

Polyomaviruses (JC virus [JCV], BK virus [BKV], and simian virus 40 [SV40]) establish subclinical and persistent infections and share the capacity for reactivation from latency in their host under immunosuppression. JCV establishes latency mainly in the kidney, and its reactivation results in the development of progressive multifocal leukoencephalopathy. BKV causes infection in the kidney and the urinary tract, and its activation causes a number of disorders, including nephropathy and hemorrhagic cystitis. Recent studies have reported SV40 in the allografts of children who received renal transplants and in the urine, blood, and kidneys of adults with focal segmental glomerulosclerosis, which is a cause of end-stage renal disease and an indication for kidney transplantation. Clinical syndromes related to polyomavirus infection are summarized in the present review, and strategies for the management of patients who receive transplants are discussed. Polyomaviruses (JC virus [JCV], BK virus [BKV], and simian virus 40 [SV40]) are classified as members of the Polyomavirus genus in the family Polyomaviridae. Polyomaviruses are nonenveloped icosahedral DNA viruses with genomes that are ∼5 kb in size, which are divided into 3 functional regions [1, 2]. The early region encodes for 2 nonstructural proteins, the large and small tumor antigens. The late region encodes 3 viral capsid proteins, VP1, VP2, and VP3. The third segment is the noncoding regulatory region. Polyomaviruses BKV and JCV share 72% DNA sequence homology, and each shares ∼70% homology with SV40 [1, 2]. Polyomaviruses typically establish subclinical and persistent infections in their natural hosts. Persistence occurs in different organs, including the kidneys, brain, and spleen [1–4]. In addition, recent reports have identified JCV, BKV, and SV40 DNA sequences in B lymphocytes in HIVinfected and uninfected patients, which suggests that polyomaviruses are lymphotropic [5–11]. These 3 viruses share the capacity for reactivation from latency in their host under imReceived 29 April 2002; revised 7 June 2002; electronically published 14 October 2002 Financial support: R.A.V. is the recipient of the 2001 Junior Faculty Development Award from GlaxoSmithKline. Reprints or correspondence: Dr. Shimon Kusne, Div. of Infectious Diseases, University of Pittsburgh Medical Center, Falk Medical Bldg., Ste. 3A, 3601 Fifth Ave., Pittsburgh, PA 15213 ([email protected]). Clinical Infectious Diseases 2002; 35:1081–7 2002 by the Infectious Diseases Society of America. All rights reserved. 1058-4838/2002/3509-0010$15.00

munosuppression and may infect additional tissues when reactivated [1–4]. Both JCV and BKV were first isolated in 1971 and were named after the first patients from whom the isolates were obtained. The first JCV isolate was recovered from the brain of a patient with progressive multifocal leukoencephalopathy (PML) [12]. BKV was isolated in the urine of a patient who developed ureteral stenosis after undergoing renal transplantation [13]. The history of polyomavirus SV40 infection in humans is associated with the poliovirus vaccine. Both inactivated and live attenuated forms of the poliovirus vaccine were prepared in primary rhesus monkey kidney cells, some of which were obtained from animals naturally infected with SV40, a virus that was not known at the time. Different studies demonstrated that infectious SV40 survived the vaccine inactivation treatments, and, from 1955 to early 1963, millions of people worldwide were inadvertently exposed to live SV40 when they were given these SV40-contaminated vaccines [1, 2, 14–16]. Some adenovirus vaccines (types 3 and 7) that were distributed in the United States to military and civilian personnel during 1961–1965 also contained SV40 [17]. However, the prevalence of polyomavirus SV40 infection in the human population today is not known. This is because the exact number of subjects who actually received contaminated vaccines is unknown, because the dose of infectious SV40 present in different lots of vaccine is not available, and because it IMMUNOCOMPROMISED HOSTS • CID 2002:35 (1 November) • 1081

is difficult to follow a large population for decades after virus exposure. However, the recent demonstration of SV40 genomes in some cancers and nonmalignant diseases in humans suggests that the virus may be etiologically meaningful in the development of those diseases [3]. Polyomaviruses JCV and BKV also have the ability to induce tumors in laboratory animals [2, 18], and they have been associated with some solid tumors in humans (in particular, brain cancers) [18–20], although less frequently than has SV40 [1, 3]. In recent years, there has also been increased recognition of the significant morbidity associated with the nonmalignant clinical syndromes related to polyomaviruses in kidney transplant and bone marrow transplant (BMT) recipients. The aims of this review are to summarize the data regarding nonmalignant clinical syndromes related to polyomavirus infections in renal transplant recipients and BMT recipients and to describe strategies for the management of such patients.

EPIDEMIOLOGY AND PATHOGENESIS Polyomaviruses JCV and BKV cause primary infections at an early age; the highest incidence of infection occurred between the ages of 1 and 6 years, and the infections appear to be mostly asymptomatic [1, 2]. An association of primary polyomavirus infection with mild respiratory tract disease, mild pyrexia, and transient cystitis has been reported [21], but the route of infection of these viruses has not been firmly defined. Approximately 80% of the adult population worldwide is seropositive for JCV and BKV [1, 2]. Serological studies (from 1970 to the present) that used a specific plaque reduction neutralization assay detected polyomavirus SV40 antibodies both in persons old enough to have been exposed to contaminated poliovirus vaccines and in those born after vaccines were no longer contaminated with SV40 [3, 22–24]. SV40 seroprevalence rates have ranged from 2% to 20% in different surveys conducted in the general population. This result supports the findings of an early study that indicated that SV40 causes infections in humans [3], and it raises the possibility that the virus may be circulating in the population not exposed to contaminated vaccines, although at a lower rate than are the polyomaviruses JCV and BKV. PML, the best-known clinical syndrome caused by JCV, was rarely seen before the HIV epidemic and was predominantly a disease of patients with hematological malignancies or iatrogenic deficiency of cell-mediated immunity that resulted from the receipt of such medications as steroids [12, 25]. The incidence of PML peaked during the height of the HIV epidemic, with 4%–5% of HIV-infected patients affected by PML. However, with the advent of HAART, a subsequent decrease in the incidence of PML has been observed among HIV-infected patients. Although

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PML is not common, it has been reported in both organ transplant [26, 27] and BMT recipients [28, 29]. In addition, JCV has also been shown as a coinfectious agent in renal transplant patients with BKV interstitial nephritis [30]. Whether JCV can cause nephropathy in kidney transplant recipients without preexisting BKV-induced renal disease is unknown. Polyomavirus BKV causes disease of the genitourinary tract in association with hematological malignancies, congenital immunodeficiency, and Wiskott-Aldrich syndrome [2]. The group most commonly affected by BKV is transplant recipients, including renal transplant and BMT recipients. An early prospective study of renal transplant recipients showed that ∼45% of kidney transplant patients had serological evidence of BKV reactivation [31], but only 2.5%–5% of the patients developed symptomatic illness, mostly tubulointerstitial nephritis [32, 33]. More recently, a study by Randhawa et al. [34] showed that BKV infection led to allograft failure in 36% of the affected patients. Augmented immunosuppression seems to play a role in polyomavirus reactivation; studies indicate that lowering of immunosuppression is associated with a decrease in the BKV virus load and a reduction in allograft inflammation [34–36]. Although there is concern that the use of tacrolimus and mycophenolate mofetil as primary immunosuppressive agents may increase the risk for BKV reactivation in renal transplant recipients [37], this may simply represent the widespread use of these medications in most transplantation centers. Multiple studies in different countries have indicated that polyomavirus SV40 is significantly associated with some cancers in humans, including malignant mesothelioma, ependymomas, osteosarcoma, and non-Hodgkin lymphoma [3, 38, 39]. These are the same types of tumors that are induced by the virus in laboratory animals [3]. In addition, SV40 causes PML and meningoencephalitis in monkeys with simian immunodeficiency virus infection, as well as interstitial nephritis in rhesus monkeys [4, 40]. Very few published studies have examined the frequency of SV40 infections among transplant recipients. A report by Shah et al. [41] found that 18% of adult kidney transplant patients had specific neutralizing antibody to SV40. Recently, SV40 DNA was detected and identified in the allografts of children who received renal transplants [24] and in the urine, blood, and kidneys of adult patients with focal segmental glomerulosclerosis [42], a known cause of end-stage renal disease. Moreover, it was demonstrated that SV40 seropositivity in children increased with age (P p .01) and was significantly associated with kidney transplantation (P p .001 ) [43]. However, studies are required to determine (1) the prevalence of polyomavirus SV40 infections in transplant recipients, and (2) whether the virus may be transmitted from person to person.

CLINICAL SYNDROMES Polyomavirus nephritis and hemorrhagic cystitis. Depending on the method used for diagnosis, polyomavirus viruria can be detected in 10%–45% of renal transplant recipients [1, 2, 32]. BKV reactivation usually occurs within the first 3 months after kidney transplantation, but late reactivation (occurring 1–2 years after transplantation) has been reported [44]. Of more importance, data show that reactivation and replication of polyomavirus BKV lead not only to viruria but, also, to tubulointerstitial nephritis, which can lead to severe allograft dysfunction and graft loss [32]. BKV infection is recognized to be more common among seropositive kidney transplant recipients who received an organ from seropositive donors than among those who received an organ from seronegative donors. This finding was demonstrated in a study by Andrews et al. [44] in which the prevalence of BKV infection in renal transplant recipients increased from 7.3% to 33.7% when the kidney donor was BKV seropositive instead of seronegative, and it supports the idea that reactivation of latent polyomavirus in the donated organ plays a role in viral pathogenesis. Alternatively, either primary infection of the transplanted kidney caused by BKV harbored in the recipient or de novo primary infection of the recipient may occur. However, because most individuals (∼80%) are BKV seropositive, reactivation of persisting virus in the renal allograft is the most likely explanation. It has been suggested that prolonged cold ischemia time could potentiate the effect of BKV in the transplant kidney. This is supported by studies showing that BKV-induced nephritis is less common among recipients of organs from living related donors [45], although this association is not uniformly observed in all cases [46]. Another well-established BKV syndrome in renal transplant patients is ureteral stenosis [13]. The virus may exert a direct cytopathic effect on the ureteral epithelium, resulting in ulceration and inflammation, which leads to obstructive uropathy. Obstruction can also occur as a result of accumulation of tubular casts secondary to tubular necrosis. More recently, an unusual case of fatal BKV-induced vasculopathy was reported in a renal transplant recipient [47]. Autopsy showed widespread BKV infection with concomitant necrotizing endothelial injury. Although JCV DNA has been detected in the kidneys of a subset of renal transplant recipients [30], a role for JCV as an independent etiologic agent of kidney disease has not been proven because JCV has been detected only in patients with established BKV nephropathy. Coinfection with JCV and BKV has also been reported in pregnant women with viruria and in the urine and brain tissue of HIV-infected patients with PML and patients with HIV-related nephropathy [2]. Hemorrhagic cystitis is the most common genitourinary

manifestation of polyomavirus BKV infection in BMT recipients. As many as 50% of all BMT recipients shed BKV in the urine, and 64% of patients with viruria develop late-onset hemorrhagic cystitis unrelated to cyclophosphamide therapy [2]. In seropositive BMT recipients, urinary shedding of BKV occurs 2–8 weeks after transplantation [48, 49]. Of more importance, a study by Arthur et al. [48] demonstrated that patients who excreted BKV in the urine had a 4-fold increased incidence of hemorrhagic cystitis, compared with patients without BKV viruria. JCV has been isolated from the urine of some BMT patients, but no specific clinical syndrome associated with this virus has been found in this patient population. PML. This demyelinating disease of the CNS caused by polyomavirus JCV presents with rapidly progressive focal neurological deficits, which may include hemiparesis, paresthesia, visual field deficits, ataxia, and cognitive and behavioral changes. In patients with advanced HIV infection, the disease progresses relentlessly, with death occurring within 6 months after diagnosis. PML occurs much less commonly in HIVseronegative patients. In addition to solid-organ and BMT recipients, this group includes individuals with hematological malignancies, patients receiving high-dose steroid therapy for autoimmune diseases, and patients with other diseases affecting cell-mediated immunity, such as idiopathic CD4 lymphopenia [1, 2].

DIAGNOSIS Cell culture for polyomaviruses is rarely helpful in establishing the clinical diagnosis of infection, because of the slow growth of these viruses, and because of the requirement for specialized cell lines. Serological analysis is useful for epidemiological purposes but, it is of minimal use in the diagnosis of clinical syndromes because most polyomavirus infections are believed to result from reactivation of latent infection. Polyomavirus nephritis and hemorrhagic cystitis. Demonstration of the characteristic “decoy” cells by cytologic testing of urine is suggestive of viruria with polyomaviruses. Decoy cells are epithelial cells with enlarged nuclei and large basophilic ground-glass intranuclear inclusions. However, their diagnostic usefulness is limited by the following: (1) suboptimal sensitivity [32, 50], (2) inability to distinguish between the 3 polyomaviruses able to infect humans (JCV, BKV, and SV40), and (3) potential for confusion with other viral infections (i.e., those due to cytomegalovirus or adenovirus) or malignancy [2]. Polyomavirus viruria can also be detected by electron microscopy (EM) of the urinary sediment, by ELISA, and by PCR. However, detection of viruria by use of these methods does not correlate well with clinical polyomavirus nephritis because asymptomatic subjects can shed the virus in their urine [1, 2]. On the other

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hand, for patients with biopsy-proven BKV infection, serial quantitative PCR analysis of urine and blood may be of use to follow patients and to assess response to therapy [1]. Diagnosis of polyomavirus infections in the renal allograft and the bladder must rely on histologic findings. In the earliest stages of disease, viral inclusions may be observed with little associated inflammation. Later phases of the disease are characterized by mononuclear infiltrates with focal invasion of the renal tubules and urothelium. Infected epithelial cells show enlarged nuclei, hyperchromatic chromatin, and intranuclear inclusions [32, 34, 36]. Cytoplasmic inclusion bodies are not seen, which helps distinguish polyomavirus infections from cytomegalovirus. Acute tubular necrosis is often present in the kidney. The histopathologic findings can be confused with the changes associated with acute allograft rejection, making the demonstration of polyomavirus within the renal tissue necessary for definitive diagnosis. Polyomaviruses in infected cells can be identified by a variety of techniques, including immunohistochemical analysis with a SV40-specific stain, in situ hybridization, or EM (figures 1 and 2). EM typically shows intranuclear, intracytoplasmic, and extracellular virus particles (size, 40–50 nm) arranged in small clusters or crystalline arrays. Ahuja et al. [51] indicated that patients with polyomavirus nephropathy had reduced numbers of cytotoxic T cells in their kidney allograft, compared with kidney transplant recipients with acute allograft rejection alone (7% vs. 24%; P p .01). In addition, the study revealed an increase in B cells in renal allografts with polyomavirus infection compared with those with acute rejection alone (21% vs. 6%; P p .03). These dif-

ferences are modest and subject to variability, and they are not helpful in establishing the differential diagnosis for individual patients. The routine use of PCR analysis of urine for the detection of polyomaviruses in kidney transplant recipients is not helpful because of the low specificity of PCR for the diagnosis of viral nephropathy. Blood PCR analysis can be of greater help in identifying patients at risk for BK nephropathy. In a study by Nickeleit et al. [52], BKV DNA was detected in the blood of all patients with BKV nephropathy and was found in only 8% of renal transplant patients who had no signs of nephropathy. Of more importance, 50% of the patients who were studied serially after transplantation had detectable BKV DNA in blood 16–33 weeks before nephropathy became clinically evident. The results of the blood BKV DNA test became negative, and nephropathy resolved in 33% of patients after the reduction of immunosuppression. Clearance of the virus occurred in 50% of the patients after allograft nephrectomy. Thus, blood PCR analysis for BKV DNA is a sensitive and specific screening test for BK nephropathy. However, tissue involvement needs to be confirmed by biopsy. PML. A clinical history compatible with PML in conjunction with MRI findings of asymmetric areas of increased T2 signal is strongly suggestive, but not diagnostic, of this disease. Definitive diagnosis requires a tissue sample with characteristic pathologic findings. These include multifocal areas of demyelination, nuclear enlargement, loss of normal chromatin pattern, and the presence of homogenous intranuclear basophilic staining material in oligodendrocytes. The majority of

Figure 1. Immunohistochemical stain demonstrating polyomavirus antigens in the tubular epithelium (arrow) of the kidney allograft. The infected tubules show infiltration by mononuclear cells, thus mimicking a diagnosis of acute cellular rejection.


Figure 2. Electron micrograph of a tubular epithelial cell infected by human polyomavirus BK. Virus particles (arrows) are seen in the cytoplasm as well as on the free luminal surface of the infected cell.

patients do not demonstrate inflammation, although a subset of patients with marked inflammatory changes has been described. When the characteristic histologic findings are present, the diagnosis is confirmed by identification of polyomavirus particles in enlarged oligodendrocyte nuclei by EM, by detection of viral antigens in the brain by immunofluorescence staining, or by assessment of virus gene sequences by in situ hybridization [1, 2]. When invasive diagnostic procedures are not performed, a presumptive diagnosis of PML can be made by demonstration of JCV DNA in the CSF by PCR, together with the observation of clinical and radiological findings compatible with PML. Indeed, a study by Ferrante et al. [53] indicated that JCV DNA was detected by PCR in the CSF of 90% of patients with PML. However, a negative PCR result cannot be used reliably to rule out PML.

TREATMENT AND MANAGEMENT There is no specific antiviral therapy for polyomaviruses. Currently, treatment is largely supportive and involves reduction in the doses of immunosuppressive agents. However, this approach requires close follow-up because it may lead to subsequent graft failure resulting from acute cellular rejection. A variety of antiviral agents have been studied in vitro for their effects against polyomaviruses. These include cidofovir, retinoic acid, topoisomerase inhibitors, 5-bromo 2-deoxyuridine, cytosine arabinoside (AraC), and IFN. The most promising drug

is cidofovir, an acyclic nucleoside phosphonate analogue that is known to have in vitro activity against the 3 polyomavirus that are able to infect humans (JCV, BKV, and SV40) [54, 55]. There are a few case reports of hemorrhagic cystitis in BMT recipients for which cidofovir was used with some success [56, 57]. In addition, a few renal transplant recipients have been treated with cidofovir for BKV nephropathy [58]. However, cidofovir has yet to be tested for the treatment of polyomavirus nephropathy in randomized clinical trials involving large numbers of patients. A retrospective study by De Luca et al. [59] suggested that the use of cidofovir in conjunction with HAART resulted in decreased JCV levels in the CSF as well as clinical improvement in HIV-infected patients with PML. In contrast, a recent prospective study [60] showed no benefit of cidofovir over HAART alone in HIV-infected patients with PML. However, there are few data on the treatment of PML in transplant recipients. Although reduction or withdrawal of immunosuppressive therapy may slow the progression of the disease in organ transplant patients, prospective controlled clinical trials are lacking to allow accurate evaluation. IL-2 has been used successfully in the treatment of PML in at least 1 BMT recipient [61]. There is no experience with the use of cidofovir as a treatment for PML in transplant recipients. A recent study showed that the use of AraC in HIVnegative patients with PML produced disease stabilization or improvement in one-third of the patients [62]. AraC has in vitro


activity against JCV [63]. However, prospective studies are needed to evaluate the use of AraC in transplant recipients.

CONCLUSION Polyomaviruses cause syndromes associated with significant morbidity and mortality among BMT and renal transplant recipients. Diagnosis usually requires biopsy of the affected organ and demonstration of the presence of polyomavirus in the tissue via EM or by immunohistochemical analysis. PCR testing may be helpful for early diagnosis before the development of clinical disease or for assessment of patients who have developed BKV-associated nephropathy. Prospective studies are required to answer many questions concerning polyomavirus infection in transplant recipients, including the relative importance of infection with each polyomavirus, the role of primary infection versus reactivation, and the means and frequency of monitoring infection, as well as whether polyomaviruses may affect liver, heart, and lung transplant recipients. In addition, controlled studies are needed to evaluate the efficacy of cidofovir in the treatment of polyomavirus-induced nephritis and hemorrhagic cystitis.

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