Monofunctional and Polyfunctional CD8+ T Cell Responses to Human Herpesvirus 8 Lytic and Latency Proteins

Human herpesvirus 8 (HHV-8) is the etiological agent of Kaposi''s sarcoma, primary effusion lymphoma, and multicentric Castleman''s disease. It is postulated that CD8⁺ T cell responses play an important role in controlling HHV-8 infection and preventing development of disease. In this study, we investigated monofunctional and polyfunctional CD8⁺ T cell responses to HHV-8 lytic proteins gB (glycoprotein B) and K8.1 and latency proteins LANA-1 (latency-associated nuclear antigen-1) and K12. On the basis of our previous findings that dendritic cells (DC) reveal major histocompatibility complex (MHC) class I epitopes in gB, we used a DC-based system to identify 2 novel epitopes in gB, 2 in K8.1, 5 in LANA-1, and 1 in K12. These new HHV-8 epitopes activated monofunctional and polyfunctional CD8⁺ T cells that produced various combinations of gamma interferon, interleukin 2, tumor necrosis factor alpha, macrophage inhibitory protein 1β, and cytotoxic degranulation marker CD107a in healthy HHV-8-seropositive individuals. We were also able to detect HHV-8-specific CD8⁺ T cells in peripheral blood samples using HLA A*0201 pentamer complexes for one gB epitope, one K8.1 epitope, two LANA-1 epitopes, and one K12 epitope. These immunogenic regions of viral lytic and latency proteins could be important in T cell control of HHV-8 infection.Human herpesvirus 8 (HHV-8), also referred to as Kaposi''s sarcoma-associated herpesvirus, is a gammaherpesvirus that causes Kaposi''s sarcoma (KS), primary effusion lymphoma, and multicentric Castleman''s disease. The importance of developing effective prevention and treatment for HHV-8 infection is evident in that KS, a neoplasm of endothelial origin, continues to be the most common cancer among human immunodeficiency virus (HIV)-infected patients (8). KS is also the leading cause of cancer in children in sub-Saharan Africa (7). Although the incidence of KS in HIV-infected persons declined with the advent of antiretroviral therapy (ART) (10), KS can occur in persons on ART with suppressed HIV infection and high CD4⁺ T cell counts (25).The immune responses responsible for controlling HHV-8 infection and preventing KS are not clear. CD8⁺ T cell immunity likely plays a significant role in HHV-8 infection, as these cells have been shown to be crucial in controlling infection caused by the other human gammaherpesvirus, i.e., Epstein-Barr virus (EBV) (11, 14). In support of this hypothesis, our laboratory (40-42) and others (4-6, 12, 19, 23, 26-28, 31, 32, 36, 37, 43, 44) have shown that CD8⁺ T cells produce gamma interferon (IFN-γ) in response to HHV-8 immunodominant epitopes presented by major histocompatibility complex class I (MHC-I) in HHV-8-seropositive individuals. Little is known whether T cells produce other immune mediators in response to HHV-8 infection. Indeed, polyfunctional T cells, i.e., single cells producing two or more immune mediators, have been linked to control of HIV and other persistent infections (1, 24, 29, 33) and could play a role in controlling HHV-8 infection. In one recent study, HHV-8 epitope-specific, polyfunctional T cells were detected in patients with multicentric Castleman''s disease, but these cells did not differ in number from those in healthy controls (13). Another study has found that patients with controlled KS had HHV-8-specific CD8⁺ T cells that secreted IFN-γ and tumor necrosis factor alpha (TNF-α) but that patients with progressive disease had weaker and less polyfunctional CD8⁺ T cells (2).HHV-8 epitope-specific monofunctional and polyfunctional T cell immunity could be important in development of HHV-8 vaccines that induce T cell responses that target these viral epitopes. In the present study, we therefore investigated CD8⁺ T cell responses to two HHV-8 lytic proteins, gB (glycoprotein B) and K8.1, and two latency proteins, LANA-1 (latency associated nuclear antigen-1) and K12. We previously showed that optimal induction of T cell reactivity to the HHV-8 protein gB required 1 week of stimulation with peptide-loaded, autologous, mature, monocyte-derived dendritic cells (DC) (40). Using this enhanced DC-T cell stimulation system, we now have revealed several new epitopes for these four lytic and latency HHV-8 proteins in healthy HHV-8-seropositive individuals, which induce both monofunctional and polyfunctional CD8⁺ T cells. These regions of HHV-8 could be critical in understanding HHV-8 immunopathogenesis and in vaccine development.