Essential role of nitric oxide and interferon-gamma for tumor immunotherapy with interleukin-10

Several laboratories have reported anti-tumor activity for high levels of i nterleukin-10 (IL-10) expressed as a transgene or administered as recombina nt protein. The authors have reported a positive correlation for nitric oxi de production and anti-tumor activity of IL-10 in a murine model of breast cancer. In the current study they sought evidence of a mechanistic role for nitric oxide in IL-10-mediated tumor growth inhibition. They wanted to det ermine whether pharmacologic inhibition of nitric oxide synthase (NOS) acti vity reverses the therapeutic effect of LL-IO. Administration of either of two NOS inhibitors, aminoguanidine (AC) or L-lysine,N degrees-1-iminoethyl- dihydrochloride, appears to abrogate in part the tumor growth inhibition ob served when IL-10 is overexpressed as a transgene in two murine mammary tum or cell lines. Nitric oxide levels were assessed at the tumor site by measu ring nitrosylated heme levels by electron spin resonance spectroscopy. Nitr ic oxide hemoglobin levels were lower in tumors from aminoguanidine-treated mice. indicating that effective inhibition of nitric oxide production occu rred at the tumor site, previous investigations showed that the inducible f orm of NOS protein (iNOS), but nor constitutive NOS, was expressed at highe r levels in IL-10-expressing tumors. Because iNOS is regulated at the trans criptional level, the authors compared iNOS mRNA levels in IL-IO and contro l tumors. Northern analysis revealed strong iNOS message expression in all six IL-10-expressing tumors examined, whereas message was faintly detected in parental or 66-neo rumors. The inducible form of NOS is responsive to in duction by interferon-gamma (IFN-gamma). The role of IFN-gamma in IL-10-med iated tumor inhibition and iNOS mRNA induction was determined. When tumors were transplanted to IFN-gamma mutant mice, the tumor-inhibitory activity o f IL-10 was lost. Furthermore, iNOS mRNA was no longer induced in the absen ce of host expression of IFN-gamma. These data indicate that nitric oxide c ontributes to the anti-tumor activity of IL-10 and that expression of iNOS in this context depends on IFN-gamma.