The polyomavirus, BK virus (BKV) is a ubiquitous persistent DNA virus infecting humans. It is strongly associated with urogenital tract diseases, including hemorrhagic cystitis (HC) and tubulonephritis in immunocompromised individuals such as hematopoietic cell transplant (HCT) and kidney transplant recipients. In the latter patient population, BKV-associated tubulonephritis and uretal stenosis often leads to nephropathy and allograft rejection. The observed elevation of BKV viruria in these immunocompromised patients implies viral reactivation due to escape from immune regulation. Little is known about this immune control in immunocompetent individuals, but it is likely to be mediated by cytotoxic T lymphocytes (CTL) of the cellular arm of the immune system. The relevant viral MHC-I restricted antigenic epitopes are currently unknown. We propose to identify them, thus making it possible to track cellular immunity to this pathogen, assess individuals at risk, and investigate immunotherapeutic interventions. We will employ, as a model of human immunity, a HLA transgenic (HHD-II) mouse system infected with recombinants of an attenuated poxvirus called Modified Vaccinia Ankara (MVA) expressing full length BKV major T antigen (TAg) in order to elicit candidate A2.1-restricted CTLs. The other BKV-encoded polypeptides (tAg, VP1, VP2 and VP3) will be subsequently investigated if the TAg proves weakly immunogenic. The advantages of MVA for this purpose include its avirulence in mammals, its low vector immunogenicity and low inflammatory response, which may facilitate identification of BKV-specific responses. After in vitro stimulation of splenocytes from the immunized mice with syngeneic murine cells infected with recombinant adenovirus expressing BKV TAg, the expanded CTLs will be tested for BKV-specific cytotoxicity and characterized to determine the epitopes recognized, and CTL clones generated from the bulk population. The epitopes will be fine-mapped in these assays using target cells infected with vaccinia or adenoviruses expressing BKV TAg fragments, or pulsed with panels of peptides corresponding to regions of the BKV TAg containing probable MHC-I restricted cellular epitopes. We will complement the mouse studies with investigation of the levels and character of cellular immunity to BKV in normal volunteers and in patients suffering from HC. An understanding of these putative BKV cellular epitopes will permit the design of assays such as those using MHC-I tetramers for assessment of levels of BKV-specific immune responses in clinical samples. Such information may be useful in guiding clinical management, and lay the foundation for possible future vaccines.
|Zhou, Wendi; Sharma, Madeva; Martinez, Joy et al. (2007) Functional characterization of BK virus-specific CD4+ T cells with cytotoxic potential in seropositive adults. Viral Immunol 20:379-88|
|Sharma, Madeva C; Zhou, Wendy; Martinez, Joy et al. (2006) Cross-reactive CTL recognizing two HLA-A*02-restricted epitopes within the BK virus and JC virus VP1 polypeptides are frequent in immunocompetent individuals. Virology 350:128-36|
|Wang, Zhongde; La Rosa, Corinna; Lacey, Simon F et al. (2006) Attenuated poxvirus expressing three immunodominant CMV antigens as a vaccine strategy for CMV infection. J Clin Virol 35:324-31|
|Krymskaya, Ludmila; Sharma, Madeva C; Martinez, Joy et al. (2005) Cross-reactivity of T lymphocytes recognizing a human cytotoxic T-lymphocyte epitope within BK and JC virus VP1 polypeptides. J Virol 79:11170-8|