Modulatory effects of 5‐azacytidine, phorbol ester, and retinoic acid on the malignant phenotype of human lung cancer cells

Abstract

Cloned human cell lines of squamous-cell lung carcinoma and small-cell lung carcinoma were treated with 5-azacytidine (5-azaC), 12-0-tetradecanoyl-phorbol-13 acetate (TPA), retinoic acid (RA), or a combination of these drugs, and the effects on cellular morphology, in vitro growth properties, antigenicity, tumorigenicity and metastatic activity in nude mice were studied. Antigenicity was measured by the expression of major histocompatibility antigens (MHC) and of lung-tumor-associated antigens. 5-AzaC treatment resulted in subclones with shorter population doubling times (from 40-50 hr down to 14-20 hr) and increased cloning efficiencies (from less than 1% to 5-50%). TPA and RA induced loss of proliferative activity in vitro and tumorigenicity in vivo of both lines. This was morphologically associated with the appearance of an abundance of large vaculated cells which by DNA-analysis were all found to be in G0-G1 phase. However, 2 of the 5-azaC-treated subclones were insensitive to both TPA and RA, whereas the remaining subclones (20) all responded like untreated lines to TPA and RA. One of the sublines insensitive to TPA and RA formed metastases in nude mice in contrast to all the other lines used. The density of lung-tumor-associated antigens was significantly reduced by TPA-RA treatment, whereas the expression of MHC antigens was unaffected. In contrast, 5-azaC resulted in some cases in increased density of MHC antigens. The effects of 5-azaC on cellular phenotypes were not directly correlated to the total genomic content of 5-methylcytosine. The data suggest that this experimental system is suitable for studies of phenotypic features of malignant cells as related to cellular differentiation.

Clonal analysis of the malignant properties of B16 melanoma cells treated with the DNA hypomethylating agent 5-azacytidine

“…The lack of correlation between DNA methylation levels and malignant phenotype has been observed by others [31,32]. Several of these clones also exhibited decreased DNA methylation levels, but the correlation was hOt absolute.…”

“…Recent evidence indicates that eukaryotic gene expression is controlled, at least in part, by DNA methylation at specific cytosine residues [4,36,37]. We, and others, have begun to test this hypothesis by experimental manipulation of DNA methylation levels in well-characterized tumor cell populations by treatment with DNA hypomethylating agents and examining subsequent shifts in their malignant properties [20,25,[30][31][32][39][40][41]. We, and others, have begun to test this hypothesis by experimental manipulation of DNA methylation levels in well-characterized tumor cell populations by treatment with DNA hypomethylating agents and examining subsequent shifts in their malignant properties [20,25,[30][31][32][39][40][41].…”

Modulation of growth and proliferation in squamous cell carcinoma by retinoic acid: A rationale for combination therapy with chemotherapeutic agents

“…Mechanistically, inhibition has been attributed to cytostasis rather than cytotoxicity since viability is maintained and growth arrest is reversible (Jeten, 1984). When cellular kinetic analyses have been performed, increases in the duration of S-phase have been reported for 3T3 fibroblasts and several breast carcinoma lines (Schroder et al, 1982;Marth et al, 1985), while GI blocks have been shown for Chinese hamster ovary cells, murine melanoma, human HeLa and several lung carcinoma lines (Haddox and Russell, 1979;Lotan et al, 1981;Dion and Gifford, 1980;Marth et al, 1985;Olsson et al, 1985). When [3H]-thymidine incorporation has been examined, decreased incorporation has correlated with growth inhibition (Haddox and Russel, 1979;Ueda et al, 1980;Schroder et al, 1982).…”

Vitamin A and Retinoid Derivatives for Lung Cancer: A Systematic Review and Meta Analysis

“…Historically, natural retinoids have been used in the treatment of lung cancers since the late 70's when the first human trials were conducted [4], [5], [6], [7] on the basis of antiproliferative effects specific to epithelial tissues and tumors [8], [9], [10], [11]. The development of synthetic retinoids occurred subsequently in an attempt to retain vitamin A's antiproliferative effects specific to epithelial tissue while avoiding dose limiting side effects such as skin and liver toxicity [4], [12].…”