Introduction: Carcinoembryonic antigen (CEA) is an oncofoetal antigen heterogeneously expressed by colorectal cancer cells, which may provide a useful target for antibody guided tumour localisation and therapy. The efficacy of tumour localisation would be improved by increased, more homogeneous CEA expression. Colorectal cancer cell lines are known to show augmentation of CEA expression on exposure to various chemical agents or to changing physicochemical environments. Aim: The aim of this work was to find chemical or environmental factors which successfully induced increased CEA expression in 3 different colorectal cell lines, Lovo, Ht29 and Colo, which are respectively high, low and non-expressors of CEA. Studies of differentiation were also undertaken, by observing concomitant changes in cell proliferation and morphology. Methods: The three cell lines were cultured in medium containing differentiating agents (Butyric acid, gamma-Interferon and 5-Azacytidine and Theophylline), or a range of commonly used cytotoxic drugs at a set dose, or in conditions of altered environmental factors (pH, temperature, oxygen supply, radiation, and serum content of medium). According to the observed effects of these factors used singly, combinations of chemical and physicochemical factors were also tested for additive or synergistic reactions. The growth, differentiation characteristics and CEA expression parameters were measured by a combination of Electron Microscopy, Immunocytochemistry and Fluorescein Activated Cell Sorting. Membrane CEA expression and total CEA content were compared. In addition, the degree of release of CEA by the growing cells into their medium was measured by Radioimmunoassay. A limited investigation of the genetic events which accompanied observed differentiation and CEA expression was conducted by immunostaining for oncogene products known to be associated with proliferation, apoptosis or tumour progression, e.g.,p53, Bcl-2, K-ras, and c-myc. Results: The differentiating agents were found to induce different effects in the three cell lines, with only the Ht29 cells showing increased CEA levels with each agent. It was noted that CEA expression was seen to rise in the Colo cells with two agents, using the more sensitive FACS analysis (Wilcoxon Rank Sum test). 5-Fluorouracil, hypoxia, serum-depletion and radiation induced increased CEA in some cells and not in others; Chloroquine, Methotrexate and Taxol were universally successful CEA-inducers. Of the combinations. Butyric Acid plus Interferon was particularly effective, causing potentiated augmentation of CEA in Lovo and Ht29 cells. Increases in CEA were frequently accompanied by significant growth inhibition (T-test, Correlation co-efficients -0.4, -0.54 & -0.57 for Lovo, Ht29 and Colo respectively); but were not associated with significant increases in CEA release into the supernatant, as measured by Radio-immunoassay. Electron Microscopy studies showed mixed responses to the various agents, with signs both of cell damage and of improved differentiation. No striking changes in oncogene expression by Ht29 cells exposed to successful CEA-inducers were observed; very slight changes in Bcl-2 or c-myc with 3 of the agents were noted. Conclusions: Apparent non-CEA expressing cells may be induced to express CEA using various agents. Different colorectal cell lines respond to chemical and environmental changes to different extents, via different mechanisms. Potentially useful CEA-inducing agents include Butyric Acid, Interferon, Theophylline, 5-Azacytidine, Chloroquine, 5-Fluorouracil, Methotrexate and Taxol. The combination of Butyric Acid and Interferon is particularly powerful in CEA augmentation. Increases in CEA expression are often accompanied by decreases in cell proliferation, but not by increases in CEA release. Augmentation of CEA expression may be accompanied by morphological features of cell damage or of improved differentiation
