1. Polyomavirus nephropathy: updated Helmut Hopfer, Basel, Switzerland

2. Agenda SV40 immunohistochemistry and BK viremia PVN treatment: implications for morphology PVN and rejection

3. How to diagnose PVN?

4. BK-BIFQUIT: trial design inter-observer variability SV40 IHC participants score intensity and extent inter-laboratory variability SV40 IHC organizers score intensity and extent SV40 IHC participants SV 40 IHC

5. SV40 IHC: inter-observer variability Substantial agreement (staining intensity and extent of infection) unpublished data, M. Mengel, Edmonton

6. SV40 IHC: inter-laboratory variability Below chance (staining intensity and extent of infection) Substantial agreement (positive vs. negative cores) unpublished data, M. Mengel, Edmonton

7. BK-BIFQUIT: summary BK "best practice": automated stainer, heat induced epitope retrieval (>30 minutes), either citrate or EDTA buffer, monoclonal antibody (PAB416) <1:100 for 25-35 minutes, polymer detection system Scoring of staining intensity and percentage tubules infected is not reproducible Binary categorization of cases as positive/ negative gives acceptable inter-laboratory and inter-observer reproducibility

8. SV40 IHC and BK viremia Number of tubules with SV40+ cells per mm biopsy length significantly correlates with number of BK copies in the blood High number of SV40 negative biopsies: 10'000 copies/ml ~ unpublished data, H. Hopfer, Basel

9. Summary 1 High sampling error < 10’000 c/ml: ~90% negative ≥ 10’000 c/ml: ~40% negative YES / NO scoring of SV40 immunohistochemistry

10. Agenda SV40 immunohistochemistry and BK viremia PVN treatment: implications for morphology PVN and rejection

11. Therapy reduction of immuno- suppression cidofovir? (nephrotoxicity!) leflunomide? Guidelines for screening and therapy Hirsch et al., Am J Transplant 9:S136-S146,2009 Viruria (Decoy cells) Viremia PVN "definite" "presumptive" Reduce IS Monitor viremia Resolved PVN Screening Diagnosis Therapy Resolution

12. Blood ↓ ↑ time after transplantation BK-specific cellular immunity viremia (c/ml) viruria (c/ml) adapted from Comoli et al., Curr Opin Organ Transplant 13:569-574, 2009 BK-specific immunity IFNg SFU/10 5 PBMC Immunosuppression

13. BKV-therapy and course Definite PVNPresumptive PVN Low BK-viremiap-level BKV clearance92%88%100%0.60 months 1st viremia to clearance (median) 8.84.62.90.001 clinical rejection after clearance 8%7%12%0.67 Schaub et al., Am J Transplant 10:2615-2623,2010

14. BKV therapy and morphology decreasing BKVincreasing BKVbefore BKVafter BKV Patients with BKV > 1'000 copies/ml treated with reduction of maintenance immunosuppression no rejection therapy at least 1 surveillance biopsy during BKV Morphological assessment, statistical analysis and correlation with clinical data unpublished data, H. Hopfer, Basel

15. Resolving PVN (decreasing BKV) Residual PVN (cleared BKV)

16. "Tubulitis " and inflammation During decreasing viremia there was a significant increase in the Banff tubulitis score (t) as well as the extent of interstitial inflammatory infiltrate. Persistence of intraepithelial lymphocytes and interstitial inflammation after viral clearance. unpublished data, H. Hopfer, Basel

17. Creatinine course Serum creatinine values overall remained stable (baseline - 1st replication - peak replication - clearance - last follow up) Increase of serum creatinine ≥40 umol/l during decreasing viremia in ~40% of patients, which returned to baseline without additional treatment unpublished data, H. Hopfer, Basel

18. Summary 2 BKV-specific cellular immunity Blood Kidney time after transplantation PVNresolving PVNresidual PVN BK dynamicsincreasingdecreasingcleared viruria viremia BK-specific immunity BK-induced tubular damage BK-induced inflammation anti-BK inflammation and IEL

19. PVN and rejection – a matter of faith? Do you believe in PVN and rejection? Can you distinguish PVN from rejection? How do you treat PVN and rejection?

20. BK-specific, rejection or "innocent"? BK-specific lymphocyte? (anti-BK immune response) HLA-specific lymphocyte? (rejection?) "innocent" lymphocyte? (unspecific infiltrate)

21. Distinction PVN and ICR? SV40 immunohistochemistry? Severity and extent of tubulitis and inflammation? Cellular composition of infiltrate?

22. How to treat PVN and rejection? Individualize decisions in patients with concurrent vascular or humoral rejections PVN is more important than ICR

23. Take-home messages PVN is focal, high number of falsly negative cases Resolving PVN is an anti-viral acute interstitial nephritis Give BK-specific immunity a chance Clinicopathological correlation is key to the correct diagnosis (clinical history, viral dynamics, creatinine course, morphologi- cal findings)