Polyomavirus BK-specific cellular immune response in Kidney transplant recipients

Polyomavirus BK is an emerging pathogen in KT recipients. New potent
immunosuppressive drugs promote reactivation and replication of BKV and progression
towards PVAN. PVAN occurs in up to 10% of the KT recipients with a graft loss in up to
80% of the cases. New potent immunosuppressive drugs, as MMF) and FK506 are risk factors
for developing PVAN. As no proven antiviral drugs are available, the only therapy of choice
is the reduction of immunosuppressiva in order to regain BKV-replication control (H. H.
Hirsch, M. Dickenmann, S. Binggeli, J. Steiger, Schweiz Med Forum 2004; 4:538–541).
BKV-specific cellular and humoral immune response is not well characterized. Recent
findings have shown that BKV-seropositive patients prior to transplantation are not protected
from BKV-replication. In contrast, BKV-specific cellular immune response correlates with
the diagnosis of PVAN (P. Comoli, S. Binggeli, F. Ginevri, H. H. Hirsch, Transplant
Infectious Disease Jun 2006; 8(2):86-94, Review).
The aim of this study was to investigate the interplay of BKV-specific immune response
and BKV-replication in blood samples of KT recipients. We examined the BKV-specific
immune response by ELISpot assay in KT. PBMC of KT recipients were stimulated with
BKV LT-antigen and BKV-VP1 peptide libraries. The BKV-specific immune response was
measured by the detection of IFN-γ by ELISpot assay. From the results of a pilot study with
eight patients we were able to deduce that the dynamics of viral-replication rather than the
viral load correlates with a protective immune response (S. Binggeli, A. Egli, M.
Dickenmann, I. Binet, J. Steiger, H. H. Hirsch, American Journal of Transplantation, Sep
2006; 6(9):2218-9).
To corroborate this previous observation the BKV-specific cellular immunity in 42 KT
recipients and 10 HB were tested. The KT patients were divided into two groups: patient
group 1 with an increasing or stable viral load (inc/hi)1 and patient group 2 with a decreasing
viral load or after resolved PVAN (dec)2. Indeed patients in group 2 showed a significantly
higher immune response upon stimulation with BKV-LT and BKV-VP1 than patients in
group 1 (P=0.003, P=0.001, respectively, Wilcoxon, two-sided). Detailed analysis revealed a
cut-off of >69 SFU/Mio PBMC for BKV LT-antigen, but not for BKV VP1, with
significantly more KT patients from group 2 (dec) than from group 1 (inc/hi). This cut-off has
to be validated in a prospective study and also analyzed whether such a cut-off can be used for
immunosuppressive reduction guidance.
BKV-specific cell expansion was tested in a short-term culture in the presence of either
BKV-LT or -VP1. After 9-day culture, PBMC were restimulated with BKV-LT or -VP1 and
the responses were then compared with responses to direct stimulation (without prior
cultivation). BKV-LT and -VP1 specific cellular immune responses were significantly higher
after 9-day cultivation than after direct stimulation (P=0.002, P=0.003, respectively,
Wilcoxon, two sided).
Due to high sequence homology between JCV and BKV, JCV-LT and -VP1 overlapping
peptide pools were used to test PBMC-cross recognition. JCV-LT and -VP1 responses were
significantly lower than BKV-mediated response (P=0.008, P<0.001, respectively, Wilcoxon,
two-sided). Comparison of JCV- and BKV-specific responses after 9-day culture revealed that
the BKV-VP1 response was significantly higher than the JCV-VP1 (P=0.016, Wilcoxon, two
sided), but no significant difference was observed for LT-antigen (S. Binggeli, A. Egli, S.
Schaub, I. Binet, M. Mayr, J. Steiger, H. H. Hirsch, American Journal of Transplantation, Mar
2007; 7:1-9).
Agnoprotein, a late viral protein, is highly expressed upon infection. We investigated
whether agnoprotein is able to induce a BKV-specific immune response and whether it may
serve as a diagnostic marker. Immunostaining revealed that agnoprotein was highly expressed
in the cytoplasm of infected cells and was only seen in combination with BKV-LT which is
located in the nucleus. Interestingly, BKV-agnoprotein specific cellular and humoral immune
responses were scarcely detected in HB or KT recipients. There are only few published
studies concerning BKV-agnoprotein, and further investigations are necessary to fully
understand the function of agnoprotein during infection. (D. Leuenberger, P. A. Andresen, R.
Gosert, S. Binggeli, E. H. Ström, S. Bodaghi, C Hanssen Rinaldo, H. H. Hirsch, Clinical and
Vaccine Immunology, Aug 2007; 14(8): 959-968).
As no antiviral treatment is available for BKV, the only therapy is the reduction of
immunosuppressive drugs in order to regain immunological control over BKV-replication and
PVAN. However reduction of immunosuppressants upon PVAN diagnosis bears the risk of
rejection or inflammatory response to BKV. It is difficult to distinguish between these two
outcomes because specific markers are yet lacking. Therefore, it is pivotal to record the
clinico-pathological course of the KT patient in order to correctly diagnose the problem as the
therapies are completely different. Measuring the BKV-specific cellular immune response
may support and complement other markers, such as PCR analysis and biopsies, to better
distinguish between rejection and BKV-specific immune response. (S. Schaub, M. Mayr, A.
Egli, S. Binggeli, B. Descoeudres, J. Steiger, M. J. Mihatsch, H. H. Hirsch, Nephrology
Dialysis Transplantation, Aug 2007; 22(8): 2386-90).
Finding the optimal immunosuppressive drug level is crucial for preventing rejection
(under-immunosuppressed) and viral replication (over-immunosuppressed). Our current study
showed a cut-off level of 6.65 ng/ml FK506 drug level in blood, dividing those KT patients
with and without BKV-replication control (ROC-curve: AUC=0.897, sensitivity=78%,
specificity=86%). If this cut-off is validated by a well designed prospective study, it may
serve as a guideline to administrate the optimal drug level. (S. Binggeli, 2007, current results).
BKV-specific epitopes have received considerable attention in the last five years. We
started with the epitope mapping in a kidney patient with the most common HLA-type: HLAA*
01, HLA-B*08. First screening of BKV-LT revealed ten 15aa long peptides with
immunogenic potential. Three of these ten peptides were further investigated for crossrecognition
with the homologous JCV-peptides. Even though response to the three JCVpeptides
was lower, cellular immune response could be clearly detected. It needs further
investigation to find more BKV-specific epitopes and also to test the ability of CD8+ T-cells
to kill BKV-antigen presenting cells. (S. Binggeli, 2007, current results).