Studies on the minimum requirements for in vitro "cure" of tumor cells by cytotoxic T lymphocytes

DBA/2-derived P815 tumor cells form progressively growing tumors when injected into the anterior chamber (AC) of normal BALB/c eyes. By contrast, P815 cells injected into the subconjunctival (SCon) space of BALB/c mice are promptly rejected. Despite these very different outcomes, both AC and SCon tumors elicit an equal clonal expansion of tumor-specific precursor cytotoxic T cells (pTc) within the draining cervical lymph nodes. Moreover, tumor-specific pTc migrate systemically and infiltrate both AC and SCon tumor-containing eyes. The present study explores the hypothesis that pTc generated in response to AC P815 tumors are less efficient "killers" than their SCon counterparts. To address this issue pTc were restimulated in vitro in the presence of exogenous lymphokines to initiate the terminal differentiation of pTc into fully functional cytotoxic T cells (Tc). Then an assay was developed to evaluate the capacity of AC and SCon CD8+ Tc to eliminate P815 tumor cells growing in vitro, i.e., achieve a "cure." This assay established the day tumor cells were completely eliminated (cure), as well as the rate of tumor cell removal. Using this approach it has been learned that pTc harvested from AC and SCon tumor-bearing mice have the same potential for eliminating P815 tumor cells in vitro. Moreover, the efficiency of achieving an in vitro tumor "cure" was found to be remarkably enhanced if exogenous lymphokines, as a surrogate source of T cell "help," were included in the cultures at the time effector lymphocytes were added to the proliferating tumor target cells. This latter finding emphasizes the need for sustained availability of helper factors in order to maximize the efficiency with which Tc can eliminate tumor cells. We interpret the results to mean that successful rejection of tumors in vivo is likely to turn upon (a) arrival of tumor-specific pTc at the tumor site, (b) delivery of "help" that promotes the terminal conversion of these cells into Tc, and (c) sustained presence of "help" which promotes the efficiency of immune elimination of tumor cells. Since equally large numbers of P815-specific pTc enter AC and SCon tumor sites in BALB/c mice, the failure of rejection in the former may be due to an inability of the host to provide immediate and sustained local T cell help. Implications of these studies for the development of immunotherapies for intraocular tumors are discussed.