In vitro effects of 4-hydroxyperoxycyclophosphamide on human immunoregulatory T subset function

Treatment of T cells in vitro with low concentrations of 4-hydroperoxycyclophosphamide (4-HC) is known to result in immunopotentiation of both T and B cell effector function in a manner analogous to that of cyclophosphamide administered in vivo. A previous study demonstrated that augmentation of polyclonal immunoglobulin secretion occurs following pretreatment of autologous collaborating T cells with low concentrations of 4-HC as a result of blockade of suppressor effector induction from suppressor precursor, both of which share the identical T4+,8-phenotype. The present study was undertaken to examine the effects of 4-HC on regulatory T-T interactions in mixed lymphocyte culture (MLC) responses and for allospecific cytotoxic T lymphocyte (CTL) responses. Induction of CTL and MLC proliferation were found to be sensitive to as little as 40 microM 4-HC, whereas CTL effector function was resistant to greater than or equal to 80 microM. CTL effectors were restricted to the T4-,8+ subset and the cells showing sensitivity to low and intermediate 4-HC concentrations were found to be T4+,8-. Secondary MLC and CTL responses displayed a similar 4-HC concentration-dependent inhibition following drug treatment of the T4+ T subset which could only be detected at suboptimal responder to stimulator ratios. This suggests that the mechanisms of CTL induction by a T4+ inducer cell in primary and secondary MLC responses and the sensitivity of induction of 4-HC are qualitatively similar. Pretreatment of T cells with less than or equal to 20 microM 4-HC for one hour prior to Con A activation totally blocked suppressor effector induction both for MLC and CTL function. In contrast, treatment with 80 microM 4-HC following Con A induction was without effect on differentiated T suppressor effector activity. Studies utilizing monoclonal antibody/complement depletion and panning techniques demonstrated that the suppressor precursor and differentiated suppressors for T effector function were restricted to the T4+,8-subset. These results support the hypothesis that regulatory inducer T cell function is significantly more sensitive to the inhibitory effects of low to intermediate concentrations of 4-HC than either the suppressor-cytotoxic precursors themselves or suppressor/cytotoxic effectors. Con A inducible suppressor cell precursor induction (mediated by the T4+,8-subset) demonstrated the greatest sensitivity to 4-HC (less than or equal to 20 microM) followed by inducers of primary and secondary CTL (40-60 microM).(ABSTRACT TRUNCATED AT 400 WORDS)