Advances In Molecular Biology: Impact On Rotavirus Vaccine Development

M. K. Estes
1996 Journal of Infectious Diseases  
The first candidate rotavirus vaccine was a live attenuated oral vaccine made by the classical empirical method of serial passage of virus in tissue culture cells. Current tetravalent vaccine candidates that are in the final stages of efficacy testing in the United States were made by genetic reassortment. This article briefly highlights how advances in the basic understanding of the molecular biology of rotaviruses have facilitated vaccine development. New approaches for second-generation
more » ... ond-generation vaccines and improvements in vaccine efficacy based on further exploitation of the tools and knowledge of rotavirus molecular biology and pathogenesis are discussed. The ultimate goal of molecular biology is to understand biologic processes in terms of the chemistry and physics of the macromolecules that participate in these processes. Today, molecular biology encompasses the disciplines of cell biology, biochemistry, biophysics, developmental biology, genetics, and structural biology. Herein, I briefly highlight progress toward understanding rotavirus replication and pathogenesis relative to how this basic knowledge has benefitted vaccine development. The first-generation candidate rotavirus vaccines were live attenuated oral vaccines produced by classical virologic techniques. Studies of the molecular biology of rotaviruses were essential to help interpret the results of vaccine trials and to provide other fundamental information that led to the development of multivalent rather than monovalent vaccines. Table 1 presents a time line of notable molecular biology advances relative to the development of rotavirus vaccines that are currently moving toward licensure. The milestones and references are selective, not inclusive, highlights of the advances that have benefitted vaccine development. The impact of these advances is discussed below. Early Molecular Biology Described the Rotavirus Genome and Proteins The era of molecular biology for rotaviruses began with a description of the rotavirus genome and its proteins, and provided clues to optimize virus cultivation and identify neutralization antigens. The earliest molecular studies of the rotaviruses involved simple characterization of the genome and the proteins associated with virus particles purified from stool. These studies showed that rotaviruses contain 11 segments of double-stranded (ds) RNA. Conclusions from the Financial support: NIH (DK-30144 and AI-24998); initial protein analyses, however, were less clear, primarily because the proteins underwent posttranslational modification (cleavage, glycosylation, and carbohydrate trimming) and because different viruses were studied in different laboratories. A thorough understanding of the molecular properties of the viral proteins (size, posttranslational modification, and location in the viral capsid) required direct studies focused on determining the gene-coding assignments and identification of the functions of the proteins encoded by the genome segments of a single virus strain. The development of methods to synthesize and purify mRNAs from each dsRNA genome segment, the subsequent translation in cell-free systems, and the comparative analysis of proteins made in these systems with those made in virusinfected cells and those present in different forms of purified virus resolved early confusion. Once the viral proteins were clearly described, molecular analysis of key viral antigens and immunogens could proceed. This was facilitated and complemented by studies in which the biologic properties (hemagglutination, protease-enhanced growth of human viruses, and neutralization antigens) were identified by genetic analysis of single-gene reassortants. Cloning and the subsequent sequence analysis of genes of epidemiologically and serologically diverse virus strains brought the beginnings of an understanding of the molecular basis of the antigenic diversity, transmission, pathogenicity, and evolution of rotaviruses. Studies of viral proteins led to the recognition that the outer layer of infectious particles contains two proteins, one of which is susceptible to cleavage by proteolytic enzymes (VP4 is cleaved into VP5 and VP8) and the second of which is a glycoprotein (VP7). Figure 1 illustrates features of the virus structure, gene coding assignments, and localization of the viral proteins and antigens. Finding a glycoprotein in the capsid of this nonenveloped virus was surprising initially. However, this finding was better understood once it was recognized that rotaviruses undergo a unique morphogenetic process in which newly made particles become transiently enveloped as they bud into the endoplasmic reticulum of infected cells. Description of this process led to the discovery that the rotaviruses code for a second nonstructural glycoprotein (now called NSP4), and
doi:10.1093/infdis/174.supplement_1.s37 pmid:8752289 fatcat:b7x3toow2bhgvm6njj6mpfc6k4