Spatial And Temporal Expression of Herpes Simplex Virus Type 1 Amplicon-Encoded Genes: Implications for Their Use As Immunization Vectors

Abstract

There is great interest in developing new immunization vectors. Helper virus-free herpes amplicons, plasmid-based vectors that encode no viral gene products and have an extremely large coding capacity, are attractive viral vaccine candidates for expressing recombinant proteins in vivo for immunization. Earlier studies in mice, using amplicons encoding the gp120 protein of human immunodeficiency virus (HIV), resulted in strikingly robust cellular immune responses as measured by cytotoxicity and interferon gamma enzyme-linked immunospot assays. To begin to understand how such vectors function in vivo to generate an immune response, we used amplicons encoding reporter constructs including green fluorescent protein (GFP) and luciferase to examine the duration of expression after administration to mice. Luciferase expression, measured with the IVIS system from Xenogen/Caliper Life Sciences (Hopkinton, MA) and by enzymatic assays of tissue extracts, revealed that expression after injection of the HSVluc amplicons peaked earlier than 24 hr after injection into mice. HSVegfp injection resulted in peak accumulation of GFP 24 hr after administration in vivo. Thus, both reporter genes revealed a rather rapid and robust expression pattern of short duration. The short period of expression appears in part to be due to gene silencing. Examination of the cells transduced by amplicons encoding GFP and human B7.1 suggested that the amplicons transduce a variety of cells, including professional antigen-presenting cells. From this and previous work, we conclude that amplicons may engender a potent immune response by directly transducing dendritic cells as well as by cross-priming of antigen produced by other transduced host cells.

HSV-1 amplicon vectors elicit polyfunctional T cell responses to HIV-1 Env, and strongly boost responses to an adenovirus prime

“…This is consistent with the changes in the functional signatures of antigenspecific CD4 + T cells that occur over time, in response to cell differentiation, establishment of immune memory, and changes in antigen load [33,46]. HSV-1 amplicon vectors are known to elicit a sharp, acute burst of antigen exposure that is extinguished within a week following vector delivery [22], while adenovirus vectors may continue to produce low levels of antigen over a protracted period [59]. Thus, the broad similarity in the functional CD4 + T cell profiles elicited by these vectors at both the effector (day 12; Figure 2) and memory phases (day 84; "prime only" groups, Figure 4) is of interest.…”

“…For these experiments, we chose to allow primed mice to 'rest' for 72 days before boosting. Since HSV-1 amplicon vectors induce high and transient levels of transgene (antigen) expression [22], we expected that the overwhelming majority of antigen specific T cells present at the time of the boost would therefore be memory cells. This prediction was consistent with the strikingly polyfunctional nature of the CD4 + T-cell population in mice that received only a priming immunization with HSV-1 amplicon or rAd5 vectors (and no boost).…”

“…These advantages include (i) their ability to elicit strong cellular immune responses to encoded antigens [19][20][21], (ii) their broad host cell tropism and ability to directly transduce antigen presenting cells, including dendritic cells (DC), both in vivo and in vitro [21,22]; (iii) their favorable safety profile and lack of expression of virally-encoded immunomodulatory genes; and (iv) their very large transgene capacity (up to 150 kb) [23,24].…”

Hexamethylene bisacetamide leads to reduced helper virus‐free HSV‐1 amplicon expression titers via suppression of ICP0

“…Transduced vector genome and expression titers were determined at 48 h post-transduction using NIH 3T3 cells, when the CMV promoter in the context of an amplicon vector is maximally expressed (Bowers and Federoff, unpublished observations, and [25]). Removal of HMBA from the amplicon packaging procedure resulted in a marked increase in expression titers, while transduced vector genome titers remained unchanged (Fig.…”

Constitutive and Inducible Innate Responses in Cells Infected by HSV-1- Derived Amplicon Vectors

“…In a study conceived to investigate how amplicon vectors function in vivo to generate an immune response, amplicon vectors expressing either GFP or luciferase reporter proteins were used to examine the duration of expression after administration to mice [46]. Injection with amplicons expressing luciferase revealed that transgene expression peaked earlier than 24 hr after injection in mice.…”