Interferon-α delays S-phase progression in human hepatocellular carcinoma cells via inhibition of specific cyclin-dependent kinases

Abstract

The potential antiproliferative effects of interferon‐α (IFN‐α) in the treatment of hepatocellular carcinoma (HCC) are controversial, and the growth inhibitory mechanisms remain poorly understood. Therefore, the current study was designed to delineate the molecular mechanisms responsible for direct antiproliferative actions of IFN‐α in HCC cells. IFN‐α receptor expression and signal transduction were examined by RT‐PCR, immunoprecipitation, Western analysis, and transient transactivation assays. Effects of IFN‐α on cell growth and cell‐cycle distribution were evaluated based on cell numbers and flow cytometry. Composition and activity of cyclin‐dependent kinase complexes were determined by immunoblotting and histone‐H1‐kinase assays. Expression of IFN‐α receptors was found in all 3 HCC cell lines. IFN‐α binding initiated phosphorylation of Jak1 and Tyk2 kinases leading to Stat1/Stat2 activation, nuclear translocation, and transactivation of an ISRE‐luciferase reporter gene construct. IFN‐α treatment resulted in a time‐ and dose‐dependent reduction of proliferation. Cell cycle analysis of G1‐synchronized, IFN‐α‐treated HCC cells revealed a substantial delay in S‐phase progression but no alteration of G1/S‐phase transition or evidence of apoptotic cell death. Reflecting the time course of S‐phase accumulation, cell cycle‐dependent induction of Cyclin A and Cyclin B was impaired, resulting in reduced activity of Cdk2 and Cdc2 kinases. Furthermore, Cdc25C was selectively down‐regulated. IFN‐α treatment inhibits growth of HCC cells by specifically delaying S‐phase progression, most likely because of inhibition of Cyclin A induction, resulting in decreased activity of the associated Cdk2 and Cdc2 kinases.

Direct anti‐cancer effect of oncostatin M on chondrosarcoma

“…30,31,[40][41][42] This pathway is clearly active in chondrosarcoma cells but we also detected modifications of G2/M proteins, leading to accumulation of inactive cdc2, the kinase necessary for entry into mitosis. 43 In SRC cells, this S/G2/M blockade seems to overwhelm the G0/G1 blockade, possibly because of a delayed induction of p27 kip1 and no induction of p21…”

Interferon α-induced apoptosis on rat preneoplastic liver is mediated by hepatocytic transforming growth factor β1

“…In an etiologically heterogeneous tumor entity such as hepatocellular carcinoma, a detailed knowledge of the pivotal pathways for preneoplastic lesions appears crucial to critically redefine therapeutic treatments. 45 Importantly, delineating the components and targets of IFN-␣ proapoptotic pathways should enhance our understanding of their significance in hepatocarcinoma treatment.…”

Interferon-α Acts on the S/G2/M Phases to Induce Apoptosis in the G1 Phase of an IFNAR2-expressing Hepatocellular Carcinoma Cell Line

“…IFN-␣ was shown to induce G 1 arrest (9) and increase susceptibility to cytotoxic anticancer drugs (10), such as 5-FU, mainly targeting the S phase. However, these effects were evaluated based only on flow cytometry and remain controversial (11)(12)(13). Observation of apoptosis induction by IFN-␣ in living cells and monitoring of cell death at distinct cell cycle stages over time are essential to clarify the mechanisms of the cellular responses induced by IFN-␣.…”