Effects of interferon on cellular proliferation and apoptosis

Abstract: Interferon (IFN) therapy is today a well established treatment in many diseases, including various malignancies. How IFN exerts antitumor activity is not known, but several mechanisms have been suggested. Previous studies have shown a correlation between the in vitro susceptibility of primary malignant cells to IFN and the clinical response of the patient to IFN therapy, supporting the idea that the antitumor activity of IFN results from direct effects of IFN on the tumor cells. The studies in the present thesis have dealt with the cellular effects of IFN on both malignant and normal cells, with the focus on how IFN modulates proliferation and apoptosis. In addition to their well known antiviral effects, IFNs can exert pleiotropic effects on cells, including potent cell growth inhibition of many cell types. Studies on various tumor cell lines and normal cells, as well as primary tumor cells, have established the antiproliferative effect as an important contributor to the decreased number of tumor cells commonly observed following IFN treatment. The role of apoptosis with regard to IFN's anticellular effects has so far been poorly defined. Using as a model system a number of hematopoietic cell lines, we showed that IFN-[alpha] in vitro is a potent inducer of apoptotic cell death, and that IFN-[alpha] -mediated growth arrest and apoptosis are independent responses to IFN-[alpha]. IFN-[alpha] induces remissions in approximately 15% of patients with multiple myeloma. We have previously shown that IFN-[alpha] exerts a direct cytotoxic effect on myeloma cells from some patients. Analysis of expression of the apoptosis-inhibitory protein, Bcl-2, in pre-treatment bone-marrow samples from patients with myeloma revealed a significant association (p=0.012) between high levels of Bcl-2 and resistance to IFN-[alpha] therapy. These data indicate that over-expression of Bcl-2 may be a cause for resistance to IFN-[alpha] therapy in myeloma, and that one possible mechanism for IFN's antitumor effect in this disease may be induction of apoptosis. In addition to these cytoreductive effects, IFNs have also in some systems been shown to protect malignant cells from apoptosis induced by different stimuli. In a study of p53-induced apoptosis, IFN-[gamma], but not IFN-[alpha], was found to protect cells from apoptosis. In other studies, the molecular mechanism behind IFN-[alpha] induced cell growth arrest was examined. The effect of IFN-[alpha] on expression of members of the cyclin-dependent kinase inhibitor (CKI) families was investigated in sensitive and resistant tumor cell lines, as well as in normal IL-2 - stimulated T-cells. The results demonstrated that IFN-[alpha] is a potent regulator of several CK1s, both from the INK4 family (p15) and the CIP/KIP family (p21 and p27). In sensitive tumor cell lines, a primary response to IFN-[alpha] is induction and binding of p21 to the Gl cyclin dependent kinases (CDKS) CDK4 and CDK2, causing inhibition of these kinases. Secondary events include the increased expression and accumulation of p27 in these G1 CDK complexes, rather than loss of the Gl kinase cyclin components, as well as dephosphorylation of the different pocket proteins. Importantly, in a resistant cell line, p21 protein was not expressed, despite high levels of p21 mRNA following IFN-[alpha] treatment. In normal IL-2 -stimulated T-cells, IFN-[alpha] was found to prevent entry into S-phase, correlating with profound inhibition of IL-2 -induced changes in G 1 regulatory proteins, including the prevention of n-litogen-induced reduction of p27 levels and upregulation of G l cyclins and CDKS.