Recombinant proteins that present tumor antigens in woodchuck liver were not available; therefore, a combined cell lysate of neoplastic liver tissues from five chronic WHV carrier Imatinib Mesylate molecular weight woodchucks with HCC was used for cell stimulation. Prior to treatment with SFV-enhIL-12, T-cell responses to tumor antigens were essentially undetectable in chronic WHV carrier woodchucks with HCC, because SIs were below the assay cutoff value of ��3.1 (Fig. (Fig.3A).3A). Following treatment with SFV-enhIL-12, all woodchucks had increases in their T-cell response to tumor antigens, and in two, T-cell response became positive, with SIs of ��3.1. The T-cell response was transient, peaked around the time of maximum tumor remission, and declined thereafter (Fig. (Fig.22 and and3).3).

In contrast, none of the control woodchucks had comparable changes in the tumor antigen-specific T-cell response. Liver antigens (i.e., a combined cell lysate of nonneoplastic liver tissues from the same woodchucks described above) were used as a control stimulator, and T-cell responses against these antigens were not detected in any woodchuck, demonstrating the specificity of the antitumoral T-cell response following treatment with SFV-enhIL-12 (Fig. (Fig.3B3B). FIG. 3. Antitumoral T-cell responses of chronic WHV carrier woodchucks following intratumoral treatment with increasing doses of SFV-enhIL-12. (A) T-cell responses to tumor antigens (i.e., clear supernatant of a combined cell lysate of neoplastic liver tissues …

The results for antitumoral T-cell responses were extended by determining the mRNA expression of leukocyte surface markers and cytokines in PBMC cultures stimulated with tumor antigens (Table (Table2).2). Compared to the pretreatment expression, treatment with SFV-enhIL-12 resulted in increased expression (��2.1-fold above the mRNA expression in unstimulated PBMC cultures but below the assay cutoff value of ��3.1) or positive GSK-3 expression (��3.1) of CD3, CD4, and CD8 in all woodchucks analyzed at 6 weeks posttreatment. At this time point the expression of IFN-�� was positive in tumor antigen-stimulated PBMC cultures from all SFV-treated woodchucks analyzed. Elevated IFN-�� expression correlated with increases in the expression of TNF-��, IL-6, and woodchuck IL-12, and expression of these cytokines was positive in both woodchucks treated with the highest dose of SFV-enhIL-12. Expression of leukocyte surface markers and cytokines was transient, peaked around the time of maximum tumor remission, and declined thereafter (Fig. (Fig.22 and Table Table2).2).