Kevin Staveley-O'Carroll

PubMed2005

Targeting of vaccinia virus using biotin-avidin viral coating and biotinylated antibodies.

INTRODUCTION: To test a general method for altering the tropism of viral vectors, we conjugated targeting antibody to the surface of recombinant vaccinia virus with a biotin-avidin-biotin linker and assessed the resulting infectivity in target cells and controls. MATERIALS AND METHODS: We biotinylated a vaccinia viral vector and used avidin to crosslink the biotinylated viral surface to a biotinylated antibody specific for a molecule on the surface of a target cell. In an in vitro model system, we coated a recombinant vaccinia construct containing the E. coli beta-galactosidase gene with antibody to the murine class I MHC molecule Db. Target cells were B78H1 murine melanoma cells transduced with either the Db gene or, as a control, the Kb gene. Infectivity was assessed by staining target cells with x-gal to demonstrate expression of virally delivered beta-galactosidase. This technique was also assessed in a second system with vaccinia/beta-gal targeted to the murine B7.2 molecule. The infectivity of the resulting construct was assessed for murine SA1 fibrosarcoma cells transfected with the B7.2 gene and for wild-type, B7.2-negative SA1. Experiments were repeated in each system with similar results. RESULTS: This strategy demonstrated antibody-mediated viral targeting in both the B78H1 and the SA1 models. Importantly, addition of the targeting coat diminished the infectivity of the modified vaccinia for control cells but preserved infectivity for targeted cells. In the B78H1 system, Db-targeted vaccinia consistently had 2- to 3-fold greater infectivity for B78H1Db than B78H1Kb. Increasing the number of avidin molecules used per virion in the synthesis of the viral coat led to greater selectivity but decreased overall infectivity. In the SA1 system, B7.2-targeted vaccinia demonstrated completely ablated infectivity for control SA1 cells, but maintained infectivity for target SA1/B7.2 cells. CONCLUSIONS: Recombinant viral vectors such as vaccinia may be coated with biotin/avidin and linked to biotinylated antibodies to preferentially target specific cell types in vitro. Such an approach may be useful in targeting recombinant lytic viruses to tumors for destruction and in immune up-regulation in vivo. Similarly, this approach may enhance nonlytic viruses for gene therapy applications.

Animals↗

PubMed2005

Developing the young academic surgeon.

In the past, the process of developing the young academic surgeon was arguably less strategic, one that was often not deliberately managed and monitored, leading in some cases to academic drift and disillusionment. Once upon a time it was assumed that greatness was genetic and that the next triple threat would emerge when a pre-programmed set of genes was turned on. Today, as the complexities and vicissitudes of our work increase, it is practically impossible for even the most gifted young person to be successful without careful attention to career development. Faculty development must be deliberate and strategic--every junior faculty member is unique and will require a customized career development plan that is well thought out, linked to measurable goals, monitored routinely and buttressed by effective mentoring. This approach will require time and commitment--precious commodities that are in short supply as the demands on our time are only escalating. By recruiting the right people (those who fit with the organization's values and goals) and providing the right environment, we can optimize the growth and satisfaction of our young faculty and, in so doing, create departments that are leaders in carrying out our missions of research, education and patient care. We cannot afford to have our young people fail--it is simply too costly, both from a financial and a human perspective.

Academic Medical Centers↗

PubMed2004

Developing the young academic surgeon.

In the past, the process of developing the young academic surgeon was arguably less strategic, one that was often not deliberately managed and monitored, leading in some cases to academic drift and disillusionment. Once upon a time it was assumed that greatness was genetic and that the next triple threat would emerge when a pre-programmed set of genes was turned on. Today, as the complexities and vicissitudes of our work increase, it is practically impossible for even the most gifted young person to be successful without careful attention to career development. Faculty development must be deliberate and strategic--every junior faculty member is unique and will require a customized career development plan that is well thought out, linked to measurable goals, monitored routinely and buttressed by effective mentoring. This approach will require time and commitment--precious commodities that are in short supply as the demands on our time are only escalating. By recruiting the right people (those who fit with the organization's values and goals) and providing the right environment, we can optimize the growth and satisfaction of our young faculty and, in so doing, create departments that are leaders in carrying out our missions of research, education and patient care. We cannot afford to have our young people fail--it is simply too costly, both from a financial and a human perspective.

Academic Medical Centers↗

PubMed2003

In vivo ligation of CD40 enhances priming against the endogenous tumor antigen and promotes CD8+ T cell effector function in SV40 T antigen transgenic mice.

The ability to initiate and sustain CD8(+) T cell responses to tumors in vivo is hindered by the development of peripheral T cell tolerance against tumor-associated Ags. Approaches that counter the onset of T cell tolerance may preserve a pool of potentially tumor-reactive CD8(+) T cells. Administration of agonist Ab to the CD40 molecule, expressed on APCs, can enhance immunization approaches targeting T lymphocytes in an otherwise tolerance-prone environment. In this report, the effects of anti-CD40 administration on priming of naive CD8(+) T cells against an endogenous tumor Ag were investigated. Line 501 mice express the SV40 large T Ag oncoprotein as a transgene from the alpha-amylase promoter, resulting in the development of peripheral CD8(+) T cell tolerance to the H-2-D(b)-restricted immunodominant epitope I of T Ag by 6 mo of age, before the appearance of osteosarcomas. We demonstrate that naive epitope I-specific TCR transgenic (TCR-I) T cells undergo peripheral tolerance following adoptive transfer into 6-mo-old 501 mice. In contrast, administration of agonistic anti-CD40 Ab led to increased expansion of TCR-I T cells in 501 mice, the acquisition of effector function by TCR-I T cells and the establishment of T cell memory. Importantly, this enhanced priming effect of anti-CD40 administration did not require immunization and was effective even if administered after naive TCR-I T cells had encountered the endogenous T Ag. Thus, anti-CD40 administration can block the onset of peripheral tolerance and enhance the recruitment of functionally competent effector T cells toward an endogenous tumor Ag.

Adoptive Transfer↗