Polyomaviruses

History
Research dates to the 1950s Murine leukemia virus passaged
1)

through mice, developed salivary gland tumors and not leukemia

agent different from MLV, caused

multiple tumor types in newborn mice >

polyomavirus Greek poly (many) and

-oma, denoting cancer
1)

Gross, J. Exp Med, 1953, 414-21

History

Next member Simian virus 40 (SV40), 1960 Screening samples from polio vaccine production (rhesus monkey kidney cells)

no cytopathic effects, virus resistant to formalin inactivation

1971, human polyomaviruses from immunocompromised patients

JC , brain from patient with PML1) BK from Urine (renal tx patient) 2)

1) Padgett, 1971, Lancet, 1257-60 2) Gardner, 1971, Lancet, 1253-57

Polyomavirus infection of different hosts

Host
Human Monkey Cattle Rabbit

Virus
BK virus JC virus
Simian virus (SV40) Simian agent 12 Lymphotropic papovavirus

Host
Mouse Hamster

Virus
Mouse polyomavirus K virus

Hamster papovavirus

Athymic Rat polyomavirus Bovine polyomavirus Rat Rabbit kidney Budgerigar vacuolating virus Parakeet fledgling disease virus

SV40, model for basic eukaryotic cell processes

For many years SV40 used in basic

research to study basic eukaryotic processes (transcription, splicing)

SV40 most heavily mutagenized

DNA molecule per base pair

Polyomavirus in humans studied with

the HIV epidemic and progress in transplantation

Classification
Family
Polyomaviridae

Genus
Polyomavirus

Virion Structure


small, nonenveloped
three capsid proteins VP1,VP2 and VP3 double-stranded covalently closed circular DNA 5kbp genome length icosahedral symmetry

relative resistant to heat formalin and lipid solvent

only VP1 exposed on surface of capsid
180000x

EM magnification 152k and 280k, Urine K Bienz, Basel

Genome Structure
Genome divided in 3 parts

early region - expressed before onset of replication late region - expressed after replication

SV40, BK and JC, 70-5% homology

regulatory region - origin of replication, promoters for early and late transcription
non overlapping late/early, transcribed from opposite strand

large T antigen binds to tumor suppressor proteins Rb and p53, initiate cell cycle
Imperiale et al, Fields Virology 5th Edition, Vol2,p2266

Chapter 61, Polyomaviruses, Fields Virology, 5th ed 2007

Viral entry and trafficking

BK

JC

SV40

Terminal (2,3)-linked sialic Sialic acid Receptors (2,6)-linked acid independent sialic acid CoUnknown Receptors Serotonin receptor (5HT-2a) MHC class I
Dugan, A.S., Eash, S. & Atwood, W.J. Transplant Infectious Disease 8 (2), 62-67

Epidemiology

Seroconversion for BK virus beween age 5-7

JC virus later

High seroprevalence >80%

Antibodies to both viruses remain lifelong But fluctuate in titer Unclear if this is due to re-infection or reactivation

Imperiale et al, Fields Virology 5th Edition, Vol2,p2284

Clinical Features of Polyomavirus infection

Progressive Multifocal Leukoencephalopathy (PML)

Haemorhagic Cystitis (post HSCT)

BK Nephropathy or Polyomavirus-associated nephropathy (PVAN)

rare, encephalitis, pneumonitis, retinitis and vasculopathies

Progressive Multifocal Leukoencephalopathy

Features
An end-stage complication of HIV, caused by the JC virus PML is rare in the general community, but relatively common in HIV infection (affecting 4% of all AIDS patients). Routine testing for HIV should be considered for any patient with PML Evolution occurs over weeks to months

CD4<100

Progressive mulltifocal leukoencephalopathy (PML) Presenting Signs and Symptoms

Afebrile, alert, no headache Progressively impaired speech, vision, motor

function
Cranial nerve deficit and cortical blindness Cognition affected relatively late

Classical AIDS-associated PML in the preHAART era

CD4+ T cell counts < 200 l
non-enhancing white matter lesions on MRI, without edema and mass effect detectable JCV DNA by PCR in CSF Absence of inflammatory infiltrates in brain biopsy Progressive evolution and fatal outcome in few months

 PML can be contained if

Sufficient number of CD4+ T cells presence of JCV-specific CD8+ cytotoxic T lymphocytes in the blood these CD8+ T cells infiltrate the brain lesions and destroy JCV-infected cells

Inflammatory reaction
causes a break-down of the blood-brain-barrier decreases viral replication in the brain and JCV VL in CSF

Diagnostics

CT brain scan may be normal or remarkable for areas of diminished density or demyelination (deterioration of the covering of the nerve) PCR of CSF for detection of JC virus

PCR is positive in about 60-80% of the cases

Differential diagnosis:

Toxoplasmosis Primary CNS lymphoma

Definitive diagnosis is by brain biopsy

Pathologic characteristics of JC infection

Detection of JC DNA by in-situ hybridisation

Luxol fast stain for myelin with lesions caused by JC lytic infection (d=demyelination)

Crystaline array of assembled virion particles in the nucleus of infected oligodentrocytes

PML Radiological findings

CNS imaging may reveal changes typical of PML

is not specific

MRI more sensitive than CT scans

single or multiple hypo-dense lesions in the subcortical white matter

with or without surrounding oedema

MRI shows increased T2 signal with liitle/no enhancement with gadolinium CT generally nonenhancing Inflammatory response (enhancement) sometimes seen in patient on ART

Progressive Multifocal Leukoencephalopathy

Text

MRI showing subcortical white matter demyelination

Role of antivirals in treatment of PML

Cytarabine in Progressive Multifocal Leukoencephalopathy Associated with HIV Infection

Pre HART area clinical trial 57 patient randomized to

anti-retroviral drug or antiretroviral drug + Cyto-Ara iv or Cyto-Ara intrathecal

Median survival 8 wks Cyt-Ara 15 wks intrathecal Cyt-Ara, 11 anti-retroviral only

Hall, NEJM,1998, 1345

PML: Treatment
No effective therapy ART with increase in CD4 count may improve neurologic function Clinical trial data do not support use of vidarabine or cidofovir

6/05

Natalizumab-associated PM Natalizumab humanized monoclonal antibody

that binds to the a4 chain of the a41 and a47 integrins

reduce the migration of leukocytes from the peripheral blood into tissues
In MS, natalizumab reduction of leukocyte extravasation into the CNS After approval by FDA, 3 patient developed PML

Risk of PML after natalizumab treatment 0.1%

BK virus
Immunocompetent: Asymptomatic shedding in the urine (0.3%) Bone marrow transplant patient Haemorrhagic cystitis Renal transplant recipients Ureteral stenosis (~ 3%) BK nephropathy (BKVN)

Terminology
Term Definition Serological/virological exposure Including replicxative/non rep. state Virus multiplication (lytic infection) in non latency sites BK infection

BK replication

BK disease

Histological evidence of BK mediated organ pathology

BKVN a new disease?

1-5% of renal transplant recipients Allograft loss in 45% Virtually nonexistant prior to 1995 0/616 between 1985-95 vs 5/70 between 1995-97
(Binet et al. Transplantation, 99)

Time Course of Detection of BK Virus DNA in the Plasma of Representative RenalAllograft Recipients with BK Virus Nephropathy

Text

9 patients with BKN and 41 without Only 2 of control in plasma

neg plasma PCR

pos plasma PCR

X biopsy proofen + clinical signs

Nickeleit et al, NEJM,342:1309,2000

Clinical manifestation of BKVN

Typical manifests as
Slowly decrease of GFR (increase in creatinin) Failure to respond to anti-rejection therapy

Onset 28-40 weeks (6-150) Histology:

Early noninflamatory foci, progression to inflammatory changes with necrosis and fibrosis (-> graft loss 5 month)

No extra renal sites or native kidney involved

Study Mengel et al. 2003 Trofe et al. 2003 Buehrig et al. 2003 Ginevri et al. 2003 Rahaminov et al. 2003 Kang et al. 2003 Ramos et al. 2002

Prevalence 1.1 2.1 2.7 3.0 3.8 3.9 5.1

FK506 57 77 100 66 100 100 91

MMF 87 54 89 66 87 33 97

Graft loss 71 54 38 33 14 100 82

Hirsch et al. 2002

Li et al. 2002 Maiza et al. 2002

6.0
7.0 7.1

60
100 50

40
50 50

0
33 50

Moriyama et al. 2003

10.3

36

NA

22

Natural history of BK nephropathy

Creatinine

Quant PCR

Qual PCR

Diagnosis of PVN
Histology
Intranuclear polyomavirus inclusion bodies in tubular or glomerular parietal cells Cell necrosis F tubular injury / dysfunction May be very focal Alterations are NOT pathognomonic PVN
Immuno histochemistry (large T-Ag) or in-situ hybridisation

Polyomavirus-associated nephropathy (PVAN) Histology

Drachenberg 2005, Human Pathology, 1245-55

Nickeleit, Transplant International 19 (12), 960-973

mild
Drachenberg, 2006, American Journal of Transplantation 4 (12), 2082-2092

severe

BK viremia

FIGURE 3. Correlation between BKV viremia and biopsy findings in 75 patients shedding BKV (see Table 1, footnotes). BKV viremia levels above 10,000 copies/mL were strongly associated with a concurrent biopsy showing PVAN (PP=0.0001). In contrast, JCV viremia was uncommon and of low level (mean 2.0E+03) correlating with the scarce numbers of tubules with positive SV40 staining in the biopsies with JCV PVAN (mean 5%).

Polyomavirus BK Versus JC Replication and Nephropathy in Renal Transplant Recipients: A Prospective Evaluation. Drachenberg, Cinthia; Hirsch, Hans; Papadimitriou, John; Gosert, Rainer; Wali, Ravinder; Munivenkatappa, Raghava; Nogueira, Joseph; Cangro, Charles; Haririan, Abdolreza; Mendley, Susan; Ramos, Emilio Transplantation. 84(3):323-330, August 15, 2007.

Risk Factors for PVAN

Immunosuppressive drugs and combinations
Combinations containing MMF or FK506

Drug concentration
no clear drug level identified

Antilymphocyte preparations
For steroid-resistant graft rejection

Steroids
Only if with unaltered immunosuppression

Other factors increasing the risk

Age Male gender White ethnicity Negative BK serostatus (children)
Positive serostatus not protective

Rare in other transplant settings No risk factors:

Cold ischemia, cadaveric, delayed graft function

Surrogate markers for PVN

Heterogeneity of risk factors Polyomavirus replication single common features for PVN
Urine cytology (Decoy cells) Quantification (Urine) BK DNA / VP1 RNA EM

Problems
Inhibition, degradation, cell pellets vs supernatans

Prevention/Treatment of BKVN

4 wks

12 wks

Cytology/UrinePCR BK viremia BK nephropathy

Prognostic values of diagnostic tests for BKVN

Viscount et al, Transplantation. 2007 Aug 15;84(3):340-5.

Relationship between the levels of BKV viruria and viremia in 204 kidney transplant recipients

Viscount et al, Transplantation. 2007 Aug 15;84(3):340-5.

Managment of PVAN

Modification of Immunosuppression
Graft loss (10 to 80%) No randomized study comparing protocols

Rejection risk 1st year >> 2nd year

Does Reduction in Immunosuppression in Viremic Patients Prevent BK Virus Nephropathy

Study design

prospective study, 2005/6 single centre, France 139 renal tx patients, 123 analysed Recommandation followed, screen for
active infection at 1,3,6,9 and 12 month

positive urine -> qPCR in blood pos blood -> biopsy and reduction in
IS
Almras et al, Transplantation, 2008:1099-14

Calineurin 20% MMF -50% qPCR in blood 4wk and 6wk

Almras et al, Transplantation, 2008:1099-14

 72% of patient with presumptive BKVN

did not progress to BKVN and cleared infection in blood

28%(3/11) did not clear viremia despite

reduction in IS

1/11 (0.9%) subsequently developed

BKVN

50% (1//2) of patients with BKVN

resolved BK nephropathy after reduction of IS
Almras et al, Transplantation, 2008:1099-14

-1.5 log10 from baseline

Almras et al, Transplantation, 2008:1099-14

Cidofovir

very broad spectrum of antiviral activity (Herpes viruses, Adenovirus, Poxviruses, polyomaviruses) poor oral bioavailability

nephrotoxic
mechanism by which cidofovir mediate antiBKV activity not clear (lack of polymerase) cidofovir can inhibit BK in-vitro lipid ester of cidofovir have superior anti BK activity

Potential antivirals for treatment of PVAN

Josephson, M.A., 2006 Transplant Infectious Disease 8 (2), 95-101.

Response of PVAN after cidofovir treatment

Kadambi, 2003,American Journal of Transplantation 3 (2), 186-191

Leflunomide

FDA approved for treatment of rheumatoid arthritis family of drugs called lonitrilamides substantial immune suppressive activity antiviral activity in-vitro against CMV, BK and HSV mechanism not clear

Leflunomide

17 patient, biopsy proven BKN loading (100mg), followed 20-60mg blood level of 50-100 micog/mL Outcome all patient reduction or viral clearance blood+urine (if >40microg/mL 7 blood and 5 from blood + urine clearance serum creatinine stabilized

Williams J, NEJM 2005, 1157

Guidelines for prevention diagnosis and management of BKVN in renal transplant patients

58

Screening for 2 years

59

We suggest screening all KTRs for BKV with quantitative plasma NAT (2C) at least: monthly for the first 36 months after transplantation (2D); then every 3 months until the end of the first posttransplant year (2D); whenever there is an unexplained rise in serum creatinine (2D); and after treatment for acute rejection. (2D) We suggest reducing immunosuppressive medications when BKV plasma NAT is persistently greater than 10 000 copies/mL (107 copies/L). (2D)
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