phorbol-12-13-didecanoate and Cell-Transformation--Neoplastic

In this paper, recent studies on the role of cell communication in cancer induction, particularly in two-stage carcinogenesis, were reviewed. Cell communication has been proposed to play an important role in cell growth and differentiation since its discovery. The recent finding that tumor promoters inhibit cell communication supports this possibility. The inhibition of cell communication by phorbol ester tumor promoters was also shown to correlate with enhancement of in vitro carcinogenesis in Balb/c 3T3 cells. This strongly suggests that the blocked cell communication may play a crucial causative role in the process of carcinogenesis. Accumulated evidence indicates that phorbol ester may induce blockage of cell communication through binding to its membrane receptor which is presumably Ca2+/phospholipid-dependent kinase. cAMP enhances cell communication and protects its inhibition by phorbol ester, presumably through activating cAMP-dependent kinase. This indicates the possibility that the two kinases may be key elements for physiological regulation of cell communication. It is proposed that the disturbance of the kinase systems by endogenous and exogenous factors may be responsible for the promotion phase of cancer induction. However, the true physiological role of cell communication in carcinogenesis remains to be demonstrated more directly. Especially, what kinds of molecules can pass through the gap junction and regulate cell functions in a cell community must be challenged in future. Some such molecules were speculatively described in this review.

Topics: Animals; Antigens, Surface; Calcium; Carcinogens; Carrier Proteins; Cell Adhesion Molecules; Cell Communication; Cell Division; Cell Line; Cell Transformation, Neoplastic; Cyclic AMP; Diterpenes; Humans; Intercellular Junctions; Mice; Mice, Inbred BALB C; Permeability; Phorbol Esters; Phospholipids; Protein Kinases; Sodium-Hydrogen Exchangers; Sodium-Potassium-Exchanging ATPase; Terpenes; Tetradecanoylphorbol Acetate

We investigated growth control in mixed cultures of normal and oncogene-transformed mouse fibroblasts. NIH/3T3 transformed by v-myc, polyoma large T, polyoma middle T, v-ras and v-src showed comparable cloning efficiencies in agarized medium. However, when cultivated with an excess of normal cells (Balb/3T3, C3H10T1/2 or primary rat and hamster embryo cells) ras, src, and middle T-transformed cells were able to form "foci" of transformation on the layer of density arrested normal cells, whereas myc- and polyoma large T-transformed cells lacked this ability. Addition of the phorbol ester tumor promoter, phorbol-12,13-didecanoate, rescued proliferation and focus-formation by these nuclear oncogenes-transformed cell lines. Evidence is presented and discussed supporting a main role of intercellular communication between normal and transformed cells in modulating suppression or expression of the transformed phenotype.

Topics: Animals; Carcinogens; Cell Line; Cell Transformation, Neoplastic; Cricetinae; Fibroblasts; Oncogenes; Phorbol Esters; Rats

BALB/c 3T3 cells can be transformed by transfection of an activated cellular oncogene as well as by chemicals. When the cells were transformed by pEJ-ras transfection, a marked increase in Mr 21,000 protein expression was found by Western blotting and immunohistochemical staining, whereas no such increase was detected in cells transformed by methylcholanthrene, suggesting two different molecular mechanisms. By directly microinjecting a fluorescent dye (Lucifer Yellow CH) into individual cells, we measured junctional intercellular communication among and between transformed and surrounding nontransformed cells. In both chemical and oncogene transformation studies, transformed cells and surrounding normal cells have similar capacities for gap-junctional communication, but there was complete lack of communication between transformed and nontransformed cells. When BALB/c 3T3 cells were transformed by methylcholanthrene initiation followed by phorbol ester promotion, again we saw no intercellular communication between transformed and nontransformed cells, suggesting that the observed selective communication block between transformed and nontransformed cells may be a general phenomenon in BALB/c 3T3 cells. These results indicate that selective lack of intercellular communication between transformed and surrounding normal cells may be an important phenomenon that separates transformed cells and nontransformed cells, permitting transformed cells to maintain autonomous growth.

Topics: Cell Communication; Cell Transformation, Neoplastic; Cells, Cultured; Methylcholanthrene; Oncogenes; Phenotype; Phorbol Esters; Proto-Oncogene Proteins; Proto-Oncogene Proteins p21(ras); Transfection

Hypoxanthine induces the differentiation of certain transformed cells in vitro, so analyses were undertaken to determine whether this purine metabolite might influence the expression of transformed phenotypes induced in normal cells by chemical agents. Chinese hamster embryo cells and human skin fibroblasts in culture were treated with the promoting agent phorbol-12,13-didecanoate (PDD) with or without prior treatment with 3-methylcholanthrene (MCA), and various phenotypic effects were monitored. Hypoxanthine was found to inhibit significantly the formation of type III foci and the increase in saturation density observed for Chinese hamster cells treated with MCA plus the phorbol ester. Inosine and the hypoxanthine analogue allopurinol could also mediate the effect on saturation density, while xanthosine could not. An increase in the saturation density of human skin fibroblasts, which can be induced by the phorbol ester alone, was also inhibited by hypoxanthine. There was no significant effect on the growth rate or the intracellular nucleotide pools with hypoxanthine-treated cells. The results suggest that a normal purine metabolite, hypoxanthine, can modulate the expression of transformed phenotypes induced in vitro by the known tumor promotor PDD. These observations could help in elucidating the cellular basis for promotion of carcinogenesis.

Topics: Animals; Cell Transformation, Neoplastic; Cells, Cultured; Cricetinae; Dose-Response Relationship, Drug; Humans; Hypoxanthine; Hypoxanthines; Methylcholanthrene; Nucleotides; Phenotype; Phorbol Esters; RNA, Transfer

Before a potentially tumorigenic cell meets a carcinogenic stimulus, its proliferation and differentiation are under tight control along with other neighbouring cells and their functions are well coordinated. In other words, cells in a given tissue form an orderly society and a cancer cell is a rebel against such a society. Therefore, it is important to study how the cellular society is maintained and how carcinogenic stimuli produce chaos in that society. The hypothesis that cell-cell interaction plays an important role in carcinogenesis has recently been reinforced by the finding that the prototype tumor promoter, phorbol esters, inhibits junctional intercellular communication. We present here several lines of evidence which suggest that block of intercellular communication is an important determinant in carcinogenesis, especially during tumor promotion process.

Topics: Animals; Carcinogens; Cell Communication; Cell Transformation, Neoplastic; Cricetinae; Extracellular Space; Homeostasis; Mice; Mice, Inbred BALB C; Neoplasms, Experimental; Phorbol Esters; Protein Kinase C; Skin Neoplasms; Tetradecanoylphorbol Acetate

When vole cells that had been transformed by Rous sarcoma virus were treated with the tumor-promoting phorbol ester 12-O-tetradecanoyl-13-acetate (TPA), specific phosphorylation of pp60v-src was increased. Partial V8 protease mapping indicated that the increased phosphorylation occurred exclusively on serine residues located in the amino terminus of the molecule. Treatment of cells with dimethyl sulfoxide or 4 alpha-phorbol-12,13-didecanoate did not elicit this response. Two-dimensional tryptic phosphopeptide mapping of pp60v-src immunoprecipitated from untreated and TPA-treated cells indicated that a specific tryptic amino-terminal peptide was hyperphosphorylated.

Topics: Animals; Arvicolinae; Carcinogens; Cell Transformation, Neoplastic; Cells, Cultured; Dimethyl Sulfoxide; ErbB Receptors; Mice; Mice, Inbred BALB C; Oncogene Protein pp60(v-src); Phorbol Esters; Phosphorylation; Protein Kinase C; Protein Kinases; Receptors, Cell Surface; Sarcoma, Avian; Tetradecanoylphorbol Acetate; Viral Proteins

In order to determine whether the tumor-promoting phorbol esters are capable of inducing normal human committed granulocytic-monocytic progenitor cells (CFUc) to proliferate and differentiate in the absence of granulocyte-monocyte colony-stimulating activity (CSA), we studied the effects of these compounds on human granulopoiesis in vitro. We found that when light-density human marrow cells or peripheral blood leukocytes were depleted of adherent cells and then incubated in semisolid tissue culture medium under conditions optimal for CFUc growth, phorbol myristate acetate (PMA) and its congeners produced no measurable stimulatory effect on the proliferation of CFUc in the absence of added CSA. Likewise, when light-density marrow cells that had not been depleted of adherent cells were plated in the cultures, no stimulation of CFUc colony growth resulted from the addition of PMA. However, when light-density peripheral blood leukocytes were used as a target source of CFUc without first subjecting them to adherence separation, enhanced proliferation and differentiation of CFUc were noted in cultures that contained PMA. To investigate the possibility that CSA production by monocytes in these cultures in response to activation by PMA might account for the enhanced colony formation that we observed, we incubated isolated peripheral blood monocytes in short-term liquid suspension cultures and found that in the presence of PMA, large quantities of CSA were secreted into the surrounding medium. Finally, we noted that when marrow cell suspensions were suboptimally stimulated by low concentrations of CSA added to the cultures, the effects of PMA on CFUc proliferation were unpredictable, enhancing colony formation in some cases and inhibiting it in others. Our data indicate that although the tumor-promoting phorbol esters do not appear capable of directly stimulating the proliferation or differentiation of human CFUc in the absence of CSA, they may do so indirectly by causing auxiliary cells such as monocytes to secrete CSA.

Topics: Bone Marrow; Bone Marrow Cells; Cell Transformation, Neoplastic; Colony-Forming Units Assay; Colony-Stimulating Factors; Granulocytes; Hematopoiesis; Hematopoietic Stem Cells; Humans; Macrophages; Monocytes; Phorbol Esters; Phorbols; Tetradecanoylphorbol Acetate

In order to understand the mechanisms of tumor promotion, the relationship between the early effects induced by promoters and the enhancement of transformation was investigated using transformable, promoter-sensitive mouse clone Balb/3T3 A31-1-1. Emphasis was placed on the measurement of all parameters in the same clonal line under the same conditions. The potentials of various derivatives of phorbol ester, indole alkaloid, and polyacetate for enhancing transformation in 3-methylcholanthrene-initiated cells were, in general, in parallel with their potentials for inhibiting phorbol-12, 13-dibutyrate-binding and for inducing early responses such as the reduction of epidermal growth factor (EGF)-binding to cell surface receptors, the increase in glucose uptake and release of arachidonic acid, and the stimulation of DNA synthesis in cells arrested at G0. There was an exception to this general rule; some agents, such as 12-O-tetradecanoylphorbol-13-acetate (TPA) and debromoaplysiatoxin, showed very strong activities to induce most of the early responses, whereas they showed slight or no activity to enhance cell transformation. The cause of this exception was ascribed to their higher susceptibility to metabolic or physical inactivation. However, we found that the continuous suppression of EGF-binding to cell surface receptors by promoters was well correlated with the enhancement of transformation, without exception. Furthermore, continuous elevation or reduction of the number of EGF receptors by various hormones was associated with the suppression or enhancement of cell transformation. The hypothesis was proposed that a continuous decrease in the number of EGF-receptors on the cell surface, or its underlying mechanisms, plays an important role in the enhancement of transformation. The mechanisms by which activation of protein kinase C leads to the enhancement of transformation were discussed, with emphasis on cytoskeletal alteration.

Topics: Animals; Carcinogens; Cell Transformation, Neoplastic; Cells, Cultured; ErbB Receptors; Lyngbya Toxins; Methylcholanthrene; Mice; Phorbol Esters; Receptors, Cell Surface; Tetradecanoylphorbol Acetate