The lymphnodal clonogenicity and kinetics of metastatic cells disseminated by a transplanted rat carcinoma

Abstract

In contrast to the wealth of data on the cellular transplantation of experimental tumours there are few concerning the further seeding of cells that may be released once tumour growth is established in situ. In particular, the lack of an in vivo clonogenic assay for spontaneous metastatic cells has limited therapeutic studies of this aspect of the cancer problem to the experimental use of clinical endpoints, e.g. prolonged survival or increased survival rates (Van de Velde et al., 1977;De Ruiter et al., 1982;Wondergem et al., 1985). In the 1950s methods for assaying the clonogenicity and transplantation kinetics of primary tumour cells were developed (Hewitt, 1958;Berry & Andrews, 1961) and this produced rapid progress in quantifying the principal factors governing the clinical outcome of primary tumour therapy (Steel, 1977). A similar development in experimental tumour metastasis could be just as important, e.g. in defining the basic principles of adjuvant treatment of patients at significant risk from occult metastases (Salmon, 1977).As indicated by Porter et al. (1973), a limiting dilution (TD50) assay would be helpful in the study of tumour metastasis but, presumably due to the previous lack of a suitable animal model (Carr & Carr, 1981;Fidler & Hart, 1982;Kim, 1984;Vandenris et al., 1985) . More importantly the method adopted was similar to a dilution assay but to avoid the need to isolate metastatic cells from a node, their numbers were assayed by determining the latency of the tumour it formed after transplantation to a fresh host. From the latency and incidence of tumour positive nodes, it is also possible to derive a TD50 curve for metastatic cells and thus provide the basis of an in situ cell survival assay after therapy (Speakman, 1986). In this paper we present and discuss our data on the lymphnodal clonogenicity of untreated metastatic cells. All the data were derived from only a few LMC1 generations using liquid-N2 stored material and no changes in tumour transplantability or growth characteristics were observed. Three types of experiment were performed. The first to determine the TD50 of LMC1 cells derived directly from the primary tumour. The second involved the transfer of axillary, inguinal and para-aortic lymph nodes from tumour-bearing to fresh hosts to determine from the incidence and latency of tumours formed the TD50 of transplanted metastatic cells. In the third, the subcutaneous primary tumour was excised at various times after its implantation, leaving the nodes to form overt metastases and thus determine their incidence and latency in the primary host.