Adjuvanticity of an IL-12 fusion protein expressed by recombinant DG-vesicular stomatitis virus Sheri D. Klas, 1 Clinton S. Robison, Michael A. Whitt, and Mark A. Miller * Department of Molecular Sciences, University of Tennessee Health Science Center, 858 Madison Avenue, Memphis, TN 38163, USA Received 3 May 2002; accepted 16 September 2002 Abstract The remarkable immunomodulatory and adjuvant properties of rIL-12 have been well described. Many early studies docu- menting the adjuvanticity of IL-12 were performed using the murine model of Listeria monocytogenes infection. In this report, we describetheconstructionofanattenuatedrecombinantvesicularstomatitisvirus(VSV-DG)thatencodesasingle-chainIL-12fusion protein (IL-12F), and the use of this virus as an expression vector to produce large quantities of IL-12F. VSV-expressed IL-12F (vIL-12F)wasthenco-administeredtomicealongwithapoorlyimmunogeniclisterialantigenpreparationasavaccineregimenand theresultingimmuneresponsesweremonitored.ThevIL-12Fwasfoundtohaveadjuvantpropertiessimilartothoseobservedfor rIL-12. Co-administration of vIL-12F and listerial antigen elicited powerful cell-mediated immune responses that conferred long- lived protective listerial immunity. These studies demonstrated that VSVDG-IL12F-infected cells secrete bioactive single-chain IL-12, and laid the foundation for studies using VSVDG-IL12F as a vector for delivery of IL-12F in vivo. Ó 2002 Elsevier Science (USA). All rights reserved. Keywords: Vaccination; Cytokines; Th1/Th2 cells; IL-12; Listeria monocytogenes; VSV; Immunity 1. Introduction Althoughtherearemanyhumanpathogensforwhich effectivevaccinesexist,vaccinedevelopmentremainsone oftheprimaryfrontiersofbiomedicalresearch.Vaccine formulations comprised of live attenuated pathogens havehistoricallybeenthemosteffective,however,fewof them are considered safe enough for general use in hu- mans. Conversely, the majority of subunit vaccines are safe,yet,veryfewsubunitvaccineshaveproventobeef- fective.Oneoftheprimarygoalsofvaccinedevelopment istoidentifyadjuvantsthateffectivelypromoteimmune responses to subunit vaccine formulations and are safe enoughforuseinhumans.Inrecentyearsithasbecome clearthatoneoftheprimaryfailuresofmany(ifnotmost) ineffective subunit vaccines is that antigen is processed and presented by antigen presenting cells primarily throughtheMHCclassIIpathway,resultinginthede- velopment of antibody-mediated, but not cell-mediated immuneresponses.Therefore,anequallyimportantgoal istodesignvaccineregimensthatelicitthemostappro- priate type of response, namely, to elicit cell-mediated (Th1type)responsesspecificforintracellularpathogens, andantibody-mediated(Th2type)responsesspecificfor extracellular pathogens [1–4]. It has been well documented that the cytokines pro- duced during the initial stages of the immune response to an invading pathogen or vaccine formulation play a critical role in the development of antigen-specific Th cells [3]. Several lines of evidence demonstrated that the ‘‘decision’’ of a Th0 cell to differentiate into a Th1 or Th2 cell is strongly influenced by the balance between IL-12, which promotes development of Th1 cells, and IL-4, which promotes development of Th2 cells [5]. Moreover,manydifferentcytokineshavebeenshownto have immunomodulatory effects that can promote the development of cell-mediated, antigen-specific immune responses when administered as a therapeutic or as an adjuvant component. Cellular Immunology 218 (2002) 59–73 www.academicpress.com * Corresponding author. Fax: 1-901-448-3244. E-mail address: mamiller@utmem.edu (M.A. Miller). 1 Currentaddress:GTx,Inc.,VanVleetResearchBuilding,3North Dunlap, Memphis, TN 38163, USA. 0008-8749/02/$ - see front matter Ó 2002 Elsevier Science (USA). All rights reserved. PII:S0008-8749(02)00575-0