Characterisation of clonal variants in a human lung carcinoma cell line: investigations into control of growth and differentiation

Analysis of many human tumours reveals the presence of two or more heterogenous cell subpopulations. Lung cancer in particular exhibits a large degree of intrinsic heterogeneity. DLKP, a human lung cell line established from a tumour which was histologically diagnosed as a ‘poorly differentiated squamous carcinoma’, appears to consist of three morphologically distinct populations. In this study, three clones apparently corresponding to these populations were established from the parental DLKP cells. The growth patterns of these isolated populations in monolayer culture, soft agar, spinner flasks and serum-free medium were investigated and it was found that in all but the latter assay, the parental DLKP cells grew faster than each of the clones and that the growth of the clones themselves varied under the different assay conditions. One clone appears to behave similarly to a multipotent, stem cell-like population, capable of giving rise to the two other clonal morphologies. Variation between the clones was also seen in their respective chromosome numbers and in their ability to adhere to extracellular matrix proteins. An immunohistological characterisation of the DLKP cells and clones was carried out. While each cell line was found to be negative for the expression of keratin and all other epithelial markers examined, the presence of neuron-specific enolase, protein gene product 9.5 and neurofilament reactivity indicated a degree of neuroendocrine (NE) differentiation. These results suggest that the DLKP cell line should be classified as either variant small cell lung carcinoma (SCLC-V) or non-small cell lung carcinoma with NE differentiation (NSCLC-NE). Studies employing the differentiation-inducing agents, 5-bromodeoxyuridine (BrdU) and retinoic acid (RA), resulted in induction of keratin expression in DLKP cells and in each of the clonal subpopulations. A similar effect was found in another keratinnegative lung carcinoma cell line in response to BrdU and this agent also appeared to increase keratin expression in cell lines which inherently expressed keratin intermediate filaments. Multidrug resistance (MDR) is a phenomenon which plays a significant role in the fatality of lung cancer. The proliferative capabilities of DLKP were compared with those of an MDR variant cell line, DLKP-A, and no significant differences were found. In addition, a clonal subpopulation of DLKP-A were transfected with a ribozyme capable of targetting and reducing the expression of p-glycoprotein, a membrane efflux pump involved in drug resistance. Drug resistance was successfully reversed in transfected cells to levels approaching those of parental DLKP sensitive cells