Stimulation of FACS-analysed CD4+ and CD8+ human tumour-infiltrating lymphocytes with ionomycin + phorbol-12,13-dibutyrate does not overcome their proliferative deficit.

Human tumour-infiltrating lymphocytes (TIL) were prepared by enzyme digestion from a series of different tumours and were purified on a fluorescence-activated cell sorter (FACS II) according to their CD4+ and CD8+ phenotype. CD4+ and CD8+ TIL were stimulated separately in a low density microculture system with phytohaemagglutinin (PHA) or with ionomycin plus phorbol-12, 13-dibutyrate (PDBu). The PHA-induced proliferation of TIL was highly decreased when compared with control peripheral blood lymphocytes. A decreased proliferation of TIL was also observed when cells were stimulated with ionomycin plus PDBu, a combination which is thought to circumvent early events associated with lymphocyte activation. Some TIL were also plated in limiting dilution where they showed decreased frequencies of proliferating T cell precursors. The data suggest that one component of the inhibition of TIL must be acting 'downstream' of the early events of lymphocyte activation.