Bryostatin 1 activates T cells that have antitumor activity.

Several strategies have been used to stimulate the growth of tumor-specific T cells in place of tumor antigen. One approach is to use pharmacologic agents to activate the second messenger pathways of T-cell activation. In the present study, we examined the ability of the protein kinase C activator bryostatin 1 (B) plus the calcium ionophore ionomycin (I) to stimulate the growth of lymphocytes obtained from the axillary lymph nodes (DLN) draining a progressively growing intradermal plasmacytoma tumor. Draining lymph node cells were initially cultured with autologous tumor cells and 20 U/ml of interleukin-2 (IL-2) for 7 days. The lymphocytes were then incubated with various concentrations of bryostatin 1 plus 1 microM ionomycin and cultured for an additional 14 days in IL-2. DLN cells initially cultured with autologous tumor and then restimulated with 5 nM bryostatin 1 and 1 microM ionomycin exhibited marked in vitro proliferation and 15-fold expansion of cell numbers over 2 weeks. The cells expanded with B/I were predominantly CD8+ T cells and retained specific in vitro cytotoxicity against autologous tumor. When adoptively transferred to mice with established liver metastases, DLN cells restimulated with B/I-mediated specific tumor regression.

Activation of CD8+ murine T cells from tumor-draining lymph nodes by phorbol dibutyrate plus calcium ionophore.

When lymphocytes from the lymph nodes draining the site of a progressively growing MCA-105 sarcoma are stimulated in vitro with autologous tumor and low-dose interleukin-2 (IL-2), they will grow and develop the ability to lyse autologous tumor cells in vitro; these lymphocytes can also eradicate tumor metastases in vivo. Phorbol esters and calcium ionophores activate signal transduction pathways in T cells and mimic the events triggered by antigen binding. We therefore sought to determine whether large numbers of MCA-105 tumor-specific, therapeutically active T cells could be obtained from MCA-105 draining lymph nodes (DLNs) following a brief exposure to phorbol dibutyrate (PDBu) and ionomycin (Io). DLN cells primarily stimulated with autologous tumor, followed by a secondary stimulation with PDBu-Io and cultured in 20 U/ml IL-2, demonstrated marked expansion of cell numbers during 3 weeks in culture, had moderate cytolytic activity [37% at effector:target ratio (E:T) = 80:1], and were all CD8+ T cells. In contrast, DLN cells stimulated primarily with PDBu-Io and cultured in 20 U/ml IL-2 demonstrated at least 8-10-fold greater growth than antigen-stimulated DLN cells during 3 weeks, were moderately cytolytic (31% at E:T = 80:1), and were a mixed population of CD8+ and CD4+ T lymphocytes. DLN cells that were expanded by either protocol, like cells stimulated repeatedly in vitro with tumor cells, could eliminate MCA-105 pulmonary metastases when given with IL-2 in an adoptive immunotherapy model. DLN cells stimulated primarily with PDBu-Io completely eradicated MCA-105 metastases but had no in vivo antitumor activity against the syngeneic B16 melanoma or MCA-203 sarcoma.(ABSTRACT TRUNCATED AT 250 WORDS)