Cyclophosphamide (CY) is commonly used as an immunosuppressive agent; however, it may also enhance the immune response when administered in low doses before antigen exposure. In order to exert these effects, CY requires metabolic activation. Metabolism of Cy gives rise to the reactive phosphoramide mustard (PAM) and acrolein, which primarily bind to DNA and sulfhydryl groups, respectively. Immunosuppression is thought to be mediated by the anti-proliferative effects of PAM, whereas the mechanism and the metabolite responsible for the CY-induced immunoenhancement are unknown. Results of others indicate that the increased immune responses produced by CY are a result of inhibiting the induction of T-suppressor cells (Ts), a T cell subset capable of down-modulating immune responses. The selective effect has been attributed the greater sensitivity of Ts- precursors cells in comparison to activated Ts, B-cells, helper T-cells, and cytolytic T-cells. Suppressor T-cells play n important role in the immune response against tumors and transplanted organs and possibly in autoimmune diseases. Thus, a basic understanding of the mechanism involved in selective modulation of Ts generation by CY may be of great significance in the development of more effective treatments for these conditions. Preliminary results from this laboratory suggest that acrolein may in part mediate the Cy-induced immunoenhancement. In addition, a synergistic enhancement of in vitro immune responses were observed wit exposure of spleen cells to certain combinations of PAM and acrolein. Thus, we first plan to determine the metabolite(s) of CY capable of selectively inhibiting Ts induction. The effects of PAM and/or acrolein exposure on the generation of Ts in well characterized models of hapten- and tumor-specific Ts generation will be investigated. The results of these investigations will allow us to direct studies to determine the biochemical mechanism by which acrolein and/or PAM inhibit Ts generation. Agents which bind sulfhydryl containing cellular molecules will be used to investigate the mechanism of acrolein, whereas anti-proliferative agents will be used to examine the mechanism of PAM. These probes will also be used to determine the mechanism for the synergistic effects produced by PAM and acrolein. The third goal is to determine the characteristics of the target cell involved with Ts-induction which allow these cell types to be more sensitive to acrolein and/or PAM. The levels of intracellular GSH and the proliferative rates of precursor Ts will be compared to B-cells, T-helper cells, and activated Ts by using fluorescent sulfhydryl binding probes and immunostaining methods in conjunction with flow cytometry.
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| Inge, T H; Hoover, S K; Susskind, B M et al. (1992) Inhibition of tumor-specific cytotoxic T-lymphocyte responses by transforming growth factor beta 1. Cancer Res 52:1386-92 |
